Biological properties of vitamins of the B-complex, part 2 – vitamins B6 and B7 (biotin, vitamin H)

Nutrition Research Reviews cambridge.org/nrr Review Article Cite this article: Dias P, Siatka T, Vopršalová M, Moravcová M, Pourová J, Přívratská N, Krčmová LK, Javorská L, and Mladěnka P (2025). Biological properties of vitamins of the B-complex, part 2 –vitamins B 6 and B 7 (biotin, vitamin H). Nutrition Research Reviews, page 1 of 34. doi: 10.1017/S0954422425100097 Received: 9 September 2024 Revised: 20 March 2025 Accepted: 19 May 2025 Keywords: biotin; essential; physiological; pyridoxine; toxicity Abbreviations: AI, adequate intake; CoA, coenzyme A; GABA, gamma-aminobutyric acid; IL, interleukin; MCC, methylcrotonyl-CoA carboxylase; PLP, pyridoxal 50-phosphate; PLPBP, pyridoxal phosphate–binding protein; PNPO, pyridoxine phosphate oxidase; SMVT, sodium-dependent multivitamin transporter; TNF-α, tumour necrosis factor α; TNSALP, tissue non-specific alkaline phosphatase Corresponding author: Přemysl Mladěnka; Email: [email protected] © The Author(s), 2025. Published by Cambridge University Press on behalf of The Nutrition Society. This is an Open Access article, distributed under the terms of the Creative Commons Attribution licence (https:// creativecommons.org/licenses/by/4.0/), which permits unrestricted re-use, distribution and reproduction, provided the original article is properly cited. Biological properties of vitamins of the B-complex, part 2 –vitamins B 6 and B 7 (biotin, vitamin H) Patrícia Dias1, TomášSiatka2, Marie Vopršalová1, Monika Moravcová1, Jana Pourová1, Nikola Přívratská3 , 4, Lenka Kujovská Krčmová3 , 4, Lenka Javorská3and Přemysl Mladěnka1 1Department of Pharmacology and Toxicology, Faculty of Pharmacy in Hradec Králové, Charles University, Hradec Králové, Czech Republic; 2Department of Pharmacognosy, Faculty of Pharmacy in Hradec Králové, Charles University, Hradec Králové, Czech Republic; 3Department of Clinical Biochemistry and Diagnostics, University Hospital Hradec Králové, Hradec Králové, Czech Republic and 4Department of Analytical Chemistry, Faculty of Pharmacy in Hradec Králové, Charles University, Hradec Králové, Czech Republic Abstract Vitamins B 6 (that is, pyridoxin and its analogues) and B 7 (that is, biotin or vitamin H) are essential molecules for many physiological processes. In addition to their well-known involvement in several enzymatic reactions, recent discoveries revealed their participation in other processes, for example, in gene expression via epigenetic processes, such as biotinylation of proteins in the case of biotin. Plants, fungi, archaea and most bacteria synthesise both vitamins, whereas animals and humans lack enzymes for their biosynthesis and depend on their exogenous supply. At least in the case of biotin, human gastrointestinal microbiota can likely partly satisfy the need. Both vitamins are water soluble and require a transporter for efficient absorption after oral administration; they can be rapidly excreted; hence, they are considered largely non-toxic. In addition to physiological and kinetic aspects of vitamin B 6 and biotin, this review, which is based on a search in PubMed up to 2023, covers sources of these vitamins, the impact of food treatment on their content, causes and symptoms of deficiency and specific mutations related to their function. Currently available literature on the analytical determination of these vitamins in biological fluids, possible pharmacological uses and symptoms of toxicity, although rare, are also included. Introduction Vitamins of the B-complex represent water-soluble molecules with essential roles in humans. The present review is a follow-up to our previous manuscript, in which we summarised the biological properties of the vitamins B 1 ,B 2 ,B 3 and B 5 (1). Herein, we centre on vitamins B 6 and B 7 (biotin) to provide a comprehensive summary of sources, properties, physiological functions, disorders that result from their deficiency and scientific information, which has been often overlooked since their discovery. We sought to cover all significant studies on the topic, including current trends and potential directions for future research. Such a review has been previously missing in the available literature. Methods PubMed was used as the bibliography database, and eligible publications were selected from 1938 to 2023. The following keywords were added to the query box: (vitamin B 6 AND properties) and (vitamin B 6 AND sources) and (vitamin B 6 AND pharmacokinetics), (vitamin B 6 AND physiological function), (vitamin B 6 AND pharmacological uses), (vitamin B 6 AND toxicity). Instead of vitamin B 6 , similar combinations were used with pyridoxine, vitamin B 7 and biotin. The eligibility criteria were as follows: peer-reviewed journal articles or book chapters published in the English language. There were no exclusion criteria for the search. Vitamin B 6 An introduction to vitamin B 6 Vitamin B 6 , ordinarily but imprecisely known as pyridoxine, is a general term for water-soluble pyridine derivatives with the same physiological role. This vitamin comprises six related compounds –vitamers (Fig. 1a), that is, pyridoxine (or pyridoxole, an alcohol), pyridoxal (an aldehyde), pyridoxamine (an amine) and their 50-phosphate esters, such as pyridoxal 50https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press phosphate (PLP), pyridoxamine 50-phosphate and pyridoxine 50-phosphate. PLP is the biologically active form of vitamin B 6 because it is a cofactor of most vitamin B 6 -dependent enzymes in the organism(2,3). Pyridoxine was discovered in 1934 by Hungarian physician Paul György and colleagues, and was isolated in pure form shortly thereafter. Humans must acquire it from their diet. Moreover, PLP can be recycled from food and degraded vitamin B 6 in the salvage pathway when the vitamin undergoes interconversion inside cells and yields different forms, including active PLP (Fig. 1b)(4,5). Pyridoxine, pyridoxal and pyridoxamine are converted to their phosphorylated forms by the pyridoxine/pyridoxamine/pyridoxal kinase, while phosphatases hydrolyze phosphorylated vitamin B 6 vitamers. Pyridoxine 50-phosphate and pyridoxamine 50-phosphate are further oxidised to the active form, PLP, by the enzyme pyridoxine (pyridoxamine) phosphate oxidase. Sources of vitamin B 6 Natural sources of vitamin B 6 Plants, fungi, archaea and most bacteria synthesise pyridoxine, whereas animals and humans lack enzymes for its biosynthetic pathway and rely solely on the exogenous supply of the vitamin(3,6– 27). Vitamin B 6 is widely distributed in foods of plant and animal origin. Whole grains, bananas, potatoes, pulses, nuts, beef, pork, poultry, organ meats and fish are good sources for humans(28–50). Some herbs and spices (for example, garlic, curry, and ginger)(51), some gluten-free pseudocereals (for example, amaranth)(52)and royal jelly are also rich in vitamin B 6 (37,53). In animal-derived foods, vitamin B 6 is usually present in phosphorylated forms (mainly of pyridoxal and pyridoxamine) and, to a lesser extent, in the free form(23,37,54–56). There is limited information on the bioavailability of vitamin B 6 from animal products in humans. The bioavailability is estimated to be generally high and, in many cases, almost Fig. 1. Chemical structures of vitamin B 6 , including its active forms, and vitamin B 7 . (a) Structure of the vitamers of B 6 .(b) Vitamin B 6 salvage pathway. PK, pyridoxine/pyridoxamine/pyridoxal kinase; PNPO, pyridoxine phosphate oxidase. (c) Chemical structure of D(þ)- biotin. The biotin molecule is composed of two rings: an imidazolidinone ring (blue) and a tetrahydrothiophene group (red) attached to a valeric acid moiety as a side chain (yellow). 2 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press complete. However, thermal processing reduces it by 25–30%; and the reaction between pyridoxal and pyridoxal phosphate with the ε-amino group of protein-bound lysine may be responsible for the decreased bioavailability(57–61). In plant-derived foods, the vitamin usually occurs as both free pyridoxine and in a glycosylated form, particularly as pyridoxine-βD -glucoside, whose proportion can range depending on the plant species, from 5% to 75% of the total vitamin content(23,28,54,57,62–68). The glucoside is only partly cleaved enzymatically by hydrolases in the small intestine, and its bioavailability is about 50% and 75% lower than that of free pyridoxine in humans and rats, respectively, that is to say, that apparently the capability of utilising the glycosylated form is species specific. The contribution of pyridoxine-βD -glucoside to the total vitamin B 6 intake in the average human diet is around 15%, hence different types of vegetarian diet do not pose a risk for vitamin B 6 deficiency. This fact is also supported by findings from a population-based survey comparing the vitamin B 6 status among vegetarians, pescatarians, flexitarians and meat-eaters. However, individuals with a marginal intake of total vitamin B 6 would be more prone to reduced nutritional status due to this incomplete bioavailability(28,29,46,54,57,58,62,64,69–83). The absolute bioavailability of vitamin B 6 from a mixed diet is estimated to be about 75%(29,44,84–86). Vitamin B 6 is also synthesised in significant quantities by the microbiota of the human large intestine, and this could represent a secondary exogenous source of the vitamin. Indeed, the existence of a specific carrier-mediated mechanism for pyridoxine uptake in human colonocytes was demonstrated. Conversely, it is likely that a large portion of the vitamin produced by microbiota is taken up by non-synthesising microbes. The extent of the contribution of microbially produced vitamin B 6 to overall body levels is unclear as there are no human studies to provide evidence for it(22,28,29,37,87–93). Amounts of vitamin B 6 in some selected foodstuffs are shown in Table 1. Antivitamins B 6 The diet can also contain antivitamin B 6 that either compete for reactive sites of vitamin B 6 -requiring enzymes or directly inactivate the vitamin(37,94). The best-known antivitamin B 6 is probably ginkgotoxin (4’-O-methylpyridoxine), which occurs in different tissues of the tree Ginkgo biloba, with the highest concentrations being present in seeds. Ingestion of ginkgotoxin can lead to abdominal pain, epileptiform convulsions and loss of consciousness due to the aforementioned interference with vitamin B 6 .As seeds are a food source in Southeast Asia, including China, Japan and Korea, and extracts from leaves are used in pharmaceutical products worldwide, they represent a potential health risk(3,95–112). Indeed, ginkgotoxin and its derivatives found in the African trees of the genus Albizia (for example, A. tanganyicensis,A. versicolor, A. julibrissin and A. lucida) are the cause of poisoning of livestock (cattle and sheep): one of the most important agricultural problems in South Africa(3,101,113). Flaxseed contains the vitamin B 6 antagonists, 1-aminoD -proline, and its precursor, the dipeptide linatine. Their possible deleterious effects through the consumption of flaxseed deserve attention in individuals with moderate vitamin B 6 status(94,114–118). Gyromitrin (N-methyl-N-formylhydrazone) from the toxic mushroom Gyromitra esculenta (genus Gyromitra is also known as false morrel) is converted to (mono) methylhydrazine after ingestion, which is able to inhibit pyridoxal kinase and hence depletes vitamin B 6. Intoxication usually occurs about 10 h after the ingestion of fresh or dried mushrooms. It gives rise to poisoning symptoms such as confusion and seizures. Table 1. Vitamin B 6 content in selected foodstuffs Food Vitamin B 6 content (μg/100 g) References Oat 120–960 (47,650,651) Wheat 127–407 (45,47,138,139,142,190,203,272,650–652) Rice, brown 123–563 (142,143,148,149) Rice, white 93–161 (142,143,148,149) Maize 307–620 (47,142,145,650,651) Rye 202–290 (47,138,650,651) Barley 231–1100 (47,138,650,651) Millet 380 (650) Sorghum 170−590 (653) Soybean 267–550 (80,654,655) Lentil 540 (476) Peanut 260–350 (654,656,657) Macadamia nut 218–300 (196,658) Pistachio nut 1032–1700 (47,196,654,658) Hazelnut 378–600 (47,196,658) Walnut 443–540 (47,196,654,658) Almond 100–188 (80,196,654,658,661) Garlic 1240 (476) Potato 140–345 (47,142,188) Carrot 60–206 (47,80,188) Cabbage 65–140 (47,188) Tomato 60–65 (47,188) Broccoli 130–190 (47,188) Cauliflower 140–170 (47,188) Spinach 120–227 (47,188) Orange 83–88 (47,80) Avocado 290 (662) Strawberry 30 (47,188) Apple 56–104 (47,80,188) Pear 14–40 (47,188) White bread 16–80 (80,143,190,663,664) Brown bread 79–170 (80,143,190,663,664) Pork 370–540 (31,39,142,476,665) Beef 264–579 (31,38,39,142,476) Chicken breast 330–811 (476,666,667) Liver, beef 840–1080 (39,476) Liver, pork 690 (39,476) Tuna 430 (668) Sardines 310 (668) Baker’s yeasts 430 (476) Oyster mushroom 100–110 (476,669) Button mushroom 50–77 (208,476,669) (Continued) Biological properties of vitamins of the B-complex 3 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press Interestingly, during cooking, methylhydrazine volatilises, and poisoning occurs also after inhalation of these vapours(118–122). Similarly, agaritine containing a hydrazinic moiety in its structure is a toxic principle of various Agaricus species, for example, the edible button mushroom Agaricus bisporus(118,121,123). The content of both toxins in fungi may be decreased by processing, such as boiling in water, drying and freezing(121,124,125). Other natural vitamin B 6 antagonists, which are of little significance to human nutrition, are toxic non-proteinogenic amino acids occurring in some leguminous plants: mimosine in Mimosa and Leucaena species, and canavanine and canaline in Canavalia species(118,126–132). Effects of food processing on vitamin B 6 content Food processing is the transformation of agricultural products into foods for human consumption. Primary processing is the conversion of the inedible raw products into food ingredients. Secondary processing involves the conversion of food ingredients into edible foods. Tertiary processed foods are commercially prepared foods. Products from primary processes make up the major part of the human diet as they are either consumed raw or used as ingredients in secondary and tertiary processes(133). Food processing may alter the vitamin B 6 content(134,135). A rough overview of the major data on vitamin B 6 losses in some food groups due to processing is presented in Supplementary Table S1 in the Supplementary Data. More data on specific foods, information on conditions and comments are in the text below. Milling and refining of cereals The primary processing of cereals (milling and refining) that separates the bran and germ, which are rich in micronutrients, from starchy endosperm causes a considerable loss of vitamin B 6 (136–141). Milling reduces the value of the vitamin B 6 content in maize by 65–75%(137,142–146). The vitamin B 6 content decreases by 66–89% in white wheat flour, compared with wholegrain flour(45,136–138,142–144,147). The content of vitamin B 6 is likewise 64% and 79.5% lower in refined than in wholegrain rye and sorghum flour, respectively.(144). Vitamin B 6 losses in nonparboiled and parboiled white rice are 42–86% and 12–26%, respectively, compared with brown rice. The decline in vitamin B 6 in parboiled rice is lower, in contrast to the non-parboiled one, because a part of the vitamin diffuses from the vitamin-rich outer bran layer into the endosperm during the parboiling process that takes place before milling(68,137,142,143,148–152). The secondary processing of cereals, such as breadmaking, rice cooking and nixtamalisation of maize, brings on additional vitamin B 6 losses. They are discussed later (‘Processing of plant-based foods’). Properties of vitamin B 6 and mechanisms of vitamin loss during food processing Vitamin B 6 loss during processing and storage of food can occur in several ways. Being soluble in water, leaching is one of the principal causes. Vitamin B 6 in foods is stable under acidic conditions but unstable in neutral and alkaline environments, particularly when exposed to heat or light. The acidic aqueous solutions of vitamin B 6 may be heated without decomposition, as vitamin B 6 is destroyed by ultraviolet radiation in neutral or alkaline solutions but not in acidic solutions. Vitamin B 6 is normally stable to oxygen. Of the several vitamers, pyridoxine is far more stable than pyridoxal and pyridoxamine. Therefore, the processing losses of vitamin B 6 tend to be highly variable, with plant-derived foods (containing mostly pyridoxine) losing little of the vitamin, and animal products (containing mostly pyridoxal and pyridoxamine) associated with higher losses(37,59,87,132,134,135,147,153–166). Processing of animal-based foods Boiling, stewing, roasting and frying reduce the vitamin B 6 content by 55%, 33–58%, 30% and 40–45%, respectively, in pork; by 60– 77%, 55–57%, 40% and 55–58%, respectively, in beef; and by 40– 58%, 40–47%, 50% and 45–56%, respectively, in chicken, depending on cooking temperature and time(161,167–171). In whole meat dishes, including cooking liquid, gravy, juice or soup, about 15–20% more vitamin B 6 remains, owing to retention of the vitamin that leached into the water phase(168,170,172–174). Fried breaded meats contain 5–35% more vitamin B 6 than those without breading, which may assist in trapping the liquid and, therefore, decreasing the loss of water-soluble vitamins(170,175). About 9% of vitamin B 6 was lost from pork and beef when the drip exuding from the frozen meat during thawing was discarded(176,177). The cooking loss of vitamin B 6 in fish meat (gilthead seabream, anchovy and Atlantic bonito) was 55–85% and 60–89% when grilled and baked, respectively, due to thermal degradation and leakage of the vitamin in the lost water(178). Heat-induced reduction of vitamin B 6 in milk is usually 5–20%, 5–10%, 5–20%, 10–50% and 40% for boiled, pasteurised, ultra-high temperature treated, sterilised and condensed milk, respectively, compared with raw milk(134,170,179–184). Hard cooked, poached, scrambled, baked and fried eggs lose 20– 23%, 15%, 10%, 10% and 10% of vitamin B 6 during cooking, respectively(170,185,186). Processing of plant-based foods Boiling, steaming and frying lead usually to a vitamin B 6 loss of 30– 35%, 15% and 10%, respectively, in vegetables alone, and to that of about 10% when taking the total dish into account(168,170).In chickpeas, microwave cooking, autoclaving and boiling caused a decline of 19%, 34% and 42% in vitamin B 6 content, respectively(187). The amount of vitamin B 6 in potatoes is reduced by 30– 57%, 21% and 10% during boiling, baking and deep frying, respectively(161,169,188). The way of cooking rice influences the content of vitamin B 6 . In different rice varieties, the boiling cooking method (cooking rice with extra water and then eliminating the water) led to vitamin losses of 3–74%, compared with the traditional cooking method (cooking with a constant amount of water without removing the water)(189). During breadmaking, the vitamin B 6 content decreased on average by 33% and 62% in whole and white wheat bread, respectively, in comparison with whole and white wheat flour(190,191). Similar Table 1. (Continued ) Food Vitamin B 6 content (μg/100 g) References Milk 35–60 (48,142,467,670) Yoghurt 87–100 (467,670) Cheese, cheddar 69 (476) Eggs 130–241 (48,142,185) 4 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press results were obtained during rye sourdough bread production(192). Toasting wheat bread induced an increase in vitamin B 6 by 75% due to its release from glycosidic bound forms(191). Effects of extrusion techniques on vitamin B 6 retention in cereal grains showed a reduction of 0–23% and of 65% in maize grits and oat whole grains, respectively(193). Drying of tarhana, a traditional Turkish fermented cereal food, resulted in vitamin B 6 losses of 3%, 16% and 23% at temperatures of 50 °C, 60 °C and 70 °C, respectively(194). A decrease in vitamin B 6 content in nuts varied from 2–7.5% in almonds, up to 4–34% in pistachio nuts after roasting(195,196). Alkali-processing of corn grains to masa (nixtamalisation) resulted in a loss of 23% of vitamin B 6 (145). The highly variable content of vitamin B 6 in beer is affected by several factors, including raw materials and the brewing process(197–199). Germination is an effective way to improve the nutrition value of edible seeds: increases of 54%, 78% and 26% in vitamin B 6 content occurred in germinated lentils(200), rough rice(201)and faba beans(202), respectively. Conversely, vitamin B 6 levels decreased by 11%, 13% and 50% in germinated wheat(203), brown rice(201)and sorghum(204), respectively, after germination. Food preservation and storage Canning, a food conservation method, brought on a vitamin B 6 reduction of 46%, 34%, 31% and 18% in mushrooms, whole peeled tomatoes, white asparagus and lentils compared with their respective unprocessed products(205). Ionising irradiation, a method used for food preservation, has a low effect on vitamin B 6 ; losses ranging from zero in wheat to about 15% in fish were observed(206,207). The amount of vitamin B 6 in button mushrooms significantly declined by 23% and 45% after 6 and 12 months, respectively, during frozen storage at −20 °C(208). The content of vitamin B 6 decreased gradually in aseptically packaged ultra-high temperature treated milk during storage at room temperature, resulting in a 96% loss after 20 weeks(183). No remarkable changes and a 20% decline in vitamin B 6 content happened in vacuum-packaged broccoli au gratin and salmon, respectively, stored at room temperature, either on the Earth or exposed to spaceflight for 880 d; the vitamin content in flight samples did not degrade faster than that of ground controls(209). The investigation of the influence of storage conditions on vitamin B 6 retention in a freeze-dried tuna mornay meal (containing tuna, vegetables and pasta) fortified with that vitamin showed a mean decrease of 14% in the vitamin following storage at temperatures of 1 °C, 30 °C and 40 °C for up to 24 months(210). The vitamin B 6 losses in meals in two hospital foodservice systems, the cook/hot-hold system, where food is held hot from the time of cooking to service, and the cook/chill system, where the cooked food is chilled, stored and reheated, have also been summarised and compared(211). Industrial production of vitamin B 6 Pyridoxine hydrochloride, which is mainly used in pharmaceutical preparations, dietary supplements and as an additive in food and feed, is manufactured by chemical synthesis(29,37,60,84,87,158,212–219). All present-day industrial vitamin B 6 syntheses use the Diels–Alder reaction of a diene (4,5-substituted oxazoles) and a dienophile (alkyldioxepins) as a key step(158,220–225). An alternative to the current chemical processes might be environmentally sustainable bioprocesses based on microbial vitamin B 6 fermentation, which is of great interest to the biotechnological industry. Several attempts have been made to construct overproducing strains by genetic engineering of microorganisms such as Sinorhizobium meliloti,E. coli and Bacillus subtilis. Unfortunately, production levels are too low and are not cost effective. Therefore, major metabolic engineering efforts are still required for developing fermentation processes that could outcompete the chemical synthesis of vitamin B 6 . The main bottlenecks are insufficient activities of some enzymes in the biosynthetic pathway and accumulation of toxic intermediate metabolites(226–237). Food fortification and biofortification with vitamin B 6 Food fortification is defined as the practice of deliberately adding an essential micronutrient to food that is commonly consumed by the general population with the intention of improving the nutritional quality of the food supply and providing a public health benefit with minimal risk to health(238–240). Foods fortified with vitamin B 6 , similarly to dietary supplements, constitute an additional dietary source of the vitamin(60,150,241–246).Overall,vitaminB 6 deficiency is rare in the general healthy population(8,29,44,150,243,246–250).It may be a concern in high-income as well as low-income countries in certain groups(142), such as older adults(245,251–254), people of low socio-economic status and those experiencing food insecurity(142,241,244,245,250). As for 2022, some countries, mostly but not solely located in Africa, have mandatory fortification of wheat flour (most often), maize flour and/or rice with vitamin B 6 (Nicaragua, Panama, Cuba, Peru, Jordan, Palestine, Nigeria, Chad, Ethiopia, Kenya, Uganda, Rwanda, Burundi, Tanzania, Mozambique, Zimbabwe and South Africa)(255–257).Thereisa voluntary fortification with vitamin B 6 in many other countries, such as the USA, the Dominican Republic, Eswatini, India, Bangladesh,Myanmar,theUKandcountriesoftheEuropean Union; the vitamin is added to various foods, such as atta, maida, rice, breakfast cereals, beverages and cereal-based foods for infants and young children(28,60,142,150,217,255,256,258–265). Biofortification is a process of increasing the density of micronutrients (vitamins and minerals) in a crop and comprises (sensu stricto, that is, omitting agronomic practices) conventional plant breeding and genetic engineering approaches. It differs from fortification because it aims to make plant foods naturally more nutritive rather than adding nutrients to the foods during food processing. Biofortification is an ideal strategy to improve nutrition for rural and poor communities that rely on subsistence farming for nutrition or may not have access to diverse diets, supplements and fortified foods. Biofortification complements existing interventions and may help by increasing the daily adequacy of micronutrient intake among the most vulnerable micronutrient deficient people(142,239,266–268). Vitamin B 6 is de novo synthesised by plants, and therefore, biofortification could be a promising route to enhance food quality by increasing the vitamin levels in plants in the future(8,269–271). Analysis of the natural diversity of vitamin B 6 content in wheat, rice and potato germplasm has shown limited variation, so breeding strategies do not seem to be adequate to increase the vitamin content in those crops(68,142,272,273), in contrast to maize, where remarkable wide ranges in vitamin B 6 levels among various genotypes were recently reported(274). Most efforts to date have used genetic engineering approaches. Biosynthesis of vitamin B 6 is primarily controlled by two enzymes, making vitamin B 6 biofortification an attractive target for plant geneticists. Overexpression of genes encoding one or both enzymes leads to the enhanced accumulation of vitamin B 6 in transgenic plants compared with the untransformed ones: 0·86– 1·25-fold in tobacco plants, 1·45–4-fold in Arabidopsis seeds, 0·16– 34·96-fold in wheat seeds, 1·6–3·9-fold in rice seeds, 3–16-fold in cassava roots and 1·07–1·5-fold in potato tubers. Interestingly, Biological properties of vitamins of the B-complex 5 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press enhancing vitamin B 6 levels in plants may also positively affect their tolerance to environmental stress(27,142,268–270,275–280). All biofortification attempts revealed the feasibility of raising the vitamin B 6 amounts in plants. So far, the vitamin B 6 contents in transgenic plants are low and highly variable. Regardless, more research for understanding the regulatory mechanisms that control genes involved in the biosynthesis and metabolism of vitamin B 6 in plants is needed(225,269). Pharmacokinetics of vitamin B 6 The total content of vitamin B 6 in the adult human body is about 170 mg(281).B 6 vitamers are absorbed in the upper small intestine (jejunum) from diet and/or oral supplements. In addition to the dietary sources of the vitamin, humans might also receive vitamin B 6 from bacterial microbiota in the large intestine as mentioned earlier(88,282,283). All vitamin B 6 analogues, that is, pyridoxine, pyridoxamine, and pyridoxal, are present in the diet. Phosphorylated forms undergo dephosphorylation by means of phosphatases prior to absorption into epithelial cells and prior to release into the portal system. Phosphorylated forms are poorly diffusible and, in fact, they are trapped in cells and a dephosphorylation step is necessary for their efflux. The bioavailability of vitamin B 6 from supplements is about 95%, whereas the bioavailability of pyridoxin, pyridoxal and pyridoxamine is similar. The presence of fibre in plant sources reduces bioavailability by 5–10%, while the presence of pyridoxine glucoside reduces bioavailability by 75–80%. On average, the bioavailability of vitamin B 6 from a mixed diet can be estimated to be about 75%. In fact, absorption in the intestine is mediated both via passive diffusion (that is, a large amount is readily absorbable without cell saturation) and a carrier mediated mechanism (that is, a saturable mechanism). In humans, there is carrier-mediated transport of B 6 vitamers via the vitamin B 1 (thiamine) transporters THTR1 and THTR2, which belong to the SLC19A2 and SLC19A3 families(284). The maximum concentration (C max ) of pyridoxine is usually achieved within 5·5h (285,286). In the liver, all forms of dephosphorylated vitamin B 6 are rephosphorylated and finally converted to pyridoxal 50-phosphate in hepatocytes. Several enzymes, such as ATP-dependent pyridoxine/pyridoxamine/ pyridoxal kinase, phosphatases and flavin mononucleotide– dependent pyridoxine phosphate oxidase (PNPO) are involved in these reactions. PNPO converts pyridoxine 50-phosphate (PNP) and pyridoxamine 50-phosphate (PMP) into pyridoxal 50-phosphate (PLP) (Fig. 1b). Pyridoxal phosphate further binds to albumin in the liver, and it is released into the circulation, where it forms approximately 60% of total circulating B 6 ,withlesseramountsofallthree dephosphorylated forms. After dissociation from albumin and dephosphorylation by alkaline phosphatase, free pyridoxal is taken up by erythrocytes and then trapped inside cells in the form of PLP(287–292). Plasma PLP is the most common parameter for determination of vitamin B 6 status. Its usual concentration is more than 30 nM in adults(5). PLP is utilised as a cofactor of many enzymes related to a row of metabolic pathways(293,294), as will be discussed later. Circulatory PLP passes into breast milk, and also crosses physiological barriers such as the placental and blood–brain barriers. The same mechanism, as in other organs, is described for brain entry and storage, that is, initial dephosphorylation in the blood–brain barrier by means of tissue non-specific alkaline phosphatase (TNSALP), followed by uptake and entrapping of the vitamin in neurons after phosphorylation to PLP(295). The major inactive metabolite of PLP is 4-pyridoxic acid. It is formed in the liver and excreted in the urine (Fig. 2). Urinary excretion of this metabolite greater than 3 mmol/d can be used as a marker of adequate short-term vitamin B 6 status. Its half-life appears to be 15–20 d(296). There is not a large storage of vitamin B 6 in tissues, probably owing to the fact that humans require only small amounts of vitamin B 6 from food sources, since the biologically active form, PLP, can be formed not only by interconversion from different B 6 vitamers but also using the cofactors from degraded enzymes in the salvage pathway. Physiological function of vitamin B 6 The active form of vitamin B 6 , PLP, acts as a coenzyme in more than 140 different enzymatic reactions necessary for vital cellular processes(8). This function is enabled by the highly reactive aldehyde group of PLP, that forms Schiff bases with the εamino groups of lysine residues at the active centres of PLP-dependent Fig. 2. Pharmacokinetics of vitamin B 6 . The figure summarises the pharmacokinetics of vitamin B 6 in the human body. PN, pyridoxine; PNP, pyridoxine 50-phosphate; PL, pyridoxal; PLP, pyridoxal 50-phosphate; PM, pyridoxamine; PMP, pyridoxamine 50-phosphate; TNSALP, tissue non-specific alkaline phosphatase; BB, blood–brain barrier. 6 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press enzymes. Conversely, binding to lysine residues on some hormonal receptors is responsible for transcriptional modulation. Moreover, the aldehyde group can react with other amino acids in proteins, especially with cysteine or histidine(297). PLP is involved in various pathways, such as: •Some steps during the metabolism of amino acids, for example, transamination, decarboxylation and racemisation processes. Metabolic transformation of sulphur-containing amino acids, for example, the conversion of methionine to cysteine through the key intermediate homocysteine or S-adenosylmethionine. Elevated levels of circulating homocysteine in the blood are associated with an increased risk of cardiovascular diseases, and S-adenosylmethionine is a methyl donor for many methylation reactions, for example, methylation of proteins, DNA and RNA, and others(298–301). In addition, cysteine synthesised by this transsulfuration pathway is an important contributor to glutathione synthesis, which plays a role in oxidative stress and the antioxidant defense system. •Some processes during carbohydrate metabolism, for example, the degradation of stored carbohydrates such as glycogenolysis, as PLP is a cofactor for glycogen phosphorylase. PLP also plays a role in the reactions that generate glucose from amino acids in the process known as gluconeogenesis(302–304). •Lipid metabolism, especially biosynthesis of sphingolipids, which are important for myelin formation and their breakdown(305). •Biosynthesis of many neurotransmitters, particularly the formation of serotonin from tryptophan and the synthesis of epinephrine (adrenaline), norepinephrine (noradrenaline) and dopamine (3,4-dihydroxyphenethylamine) from phenylalanine and tyrosine. PLP also controls the formation and regulation of the inhibitory transmitter γ-aminobutyric acid (GABA) in the brain, and the neuromodulator serine(84,306–309). •Catabolism of tryptophan and its conversion to niacin, that requires kynureninase, which also necessitates vitamin B 6 (310–313). •Biosynthesis of tetrapyrroles (for example, haem). PLP is needed for the enzymatic reaction using succinyl-CoA and glycine to generate δ-aminolevulinic acid, an intermediate precursor in tetrapyrrole biosynthesis(314,315). •Immune and inflammatory pathways, especially regulation of cytokine production, particularly interferons and interleukin 6(316,317). Besides the role of PLP as a cofactor in biochemical reactions, vitamin B 6 also plays other important roles in non-enzymatic functions, for example: •PLP inhibits enhancement in gene expression by steroid and thyroid hormones, and vitamins A and D by binding to lysine residues in hormone–receptor complexes(318,319). •Antioxidative activity by scavenging reactive oxygen species and chelating of redox-active metal ions(320–322). Vitamin B 6 deficiency and related disorders Severe vitamin B 6 deficiency resulting from inadequate intake (especially from dietary deficit) is rare in the healthy general population. Hypovitaminosis is usually found in association with other B vitamin deficiencies, such as those of folic acid (vitamin B 9 ) and vitamin B 12 . As aforementioned, it should be emphasised that dietary vitamin B 6 deficiency can occur in elderly people (aged 65 years and over)(323). Secondary vitamin B 6 deficiency is mostly a result of genetic disorders or drug interactions(324,325). Owing to the involvement of vitamin B 6 in many metabolic pathways, a lack of sufficient amounts of vitamin B 6 vitamers causes various biochemical changes and may lead to significant health problems. In particular, PLP is essential in the synthesis and metabolism of amino acids and neurotransmitters. Loss of function of the PLP-dependent enzyme glutamate decarboxylase leads to decreased levels of the inhibitory neurotransmitter GABA. Vitamin B 6 deficiency in humans is associated with seborrheic dermatitis and cheilosis (including cracks at the corners of the mouth), glossitis with ulceration, anaemia, sensory polyneuropathy, depression, decreased immune function and increased risk of cardiovascular diseases. In children, characteristic symptoms of deficiency are abnormalities in hearing and seizures.(84)Seizures are the results of an imbalance between excitatory (glutamate) and inhibitory (GABA) neurotransmitters(326–328). In the population, there are certain groups of people at increased risk of vitamin B 6 inadequacy. People with impaired absorption, especially due to malabsorption syndromes (usually associated with Crohn’s disease and ulcerative colitis) and after bariatric surgery, have low vitamin B 6 levels. Patients with renal disease, predominantly with chronic renal insufficiency undergoing dialysis, and liver disease tend to have low plasma PLP concentrations. Also, alcoholics need vitamin B 6 supplementation because alcohol is metabolised to acetaldehyde, which decreases PLP formation in cells and competes with PLP for protein binding. Additional groups at risk of vitamin inadequacy despite adequate dietary intakes are not solely elderly persons but also those with autoimmune disorders (for example, rheumatoid arthritis), those who are obese and in pregnancy, or those who are taking oral contraceptives(329–332). Analytical methods for the detection of vitamin B 6 are summarised in Table 2. More details are shown in Supplementary Data Table S2, which evaluates individual specific methodologies with the relevant citations from which the information was obtained. Pyridoxine-dependent epilepsy Pyridoxine-dependent epilepsy (pyridoxine-dependent seizures, vitamin B 6 -responsive epilepsy) is a rare inherited metabolic disease characterised by recurrent seizures with their onset usually in prenatal, neonatal and postnatal periods or in childhood. Seizures are caused primarily by low levels of GABA due to PLP deficiency, nevertheless, other abnormalities are involved, for example, low levels of adenosine and methionine cycle defects. This type of epilepsy responds to high intravenous doses of vitamin B 6 , either as pyridoxine or as its active form PLP, but are resistant to conventional antiepileptic drugs(333). Decreased PLP availability in this disease is caused by mutations in some genes involved in vitamin B 6 metabolism, for example: •Mutations in ALDH7A1, a gene encoding antiquitin, the enzyme with α-aminoaddipic semialdehyde dehydrogenase activity, involved in lysine degradation. Antiquitin deficiency leads to the accumulation of the toxic lysine intermediates αaminoadipic semialdehyde and 1-piperideine-6-carboxylic acid, inactivating PLP by chemical complexation(334–336). •Mutations in the ALDH4A1 gene occurring in metabolic disease hyperprolinaemia II causes the formation of Biological properties of vitamins of the B-complex 7 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press Table 2. Summary of analytical methods for the assessment of vitamins B 6 and B 7 in biological fluids Technique Sensitivity (nM) Analytes Matrix Advantages Disadvantages Ref. Publication year LC-MS 0·1–127·51 ×103B 6 ,B 6 -PL, B 6 -P5P, B 6 -PM, B 7 , B 6 -PA, B 6,7,9 and its vitamers, metabolites and others * Human milk * Serum * Whole blood * Plasma * Mice brain samples * Tears * Faeces Usually short analysis time, small sample volume (30–250 μl), using MRM in detection, simple methods for various matrices Some methods have complicated sample preparation (breast milk) and complicated gradient elution, some methods use SIM and are not fully validated (671,353)2012–2024 HPLC-FLD 0·3–20 B 6 -P5P, B 6 -PL, B 6 -PA and others * Whole blood * Cerebrospinal fluid * Serum Methods use small sample volume (100– 250 μl) and some of them simple derivatisation procedure Methods have no IS included, usually complicated sample preparation, long analysis time with post column derivatisation (672,357)2004–2020 HPLC-PDA 2D-LC-UV 0·1–7·29 ×103B 6 , B 6 -PL, B 6 -P5P, B 6 -PA and others * Urine * Plasma * Animal plasma Methods use small sample volume (60 μl) Methods do not have optimal recovery, there is a long analysis time, poor sensitivity, derivatisation, complicated sample preparation, and no IS (673,359)2014–2023 MLC-PDA 0·177 ×103B 6 * Plasma Method has simple sample preparation No IS is used (674)2021 Sensors/ nanodots / CL/FLD/ECD 5–9·06 ×103B 6 ,B 6 -PL, B 7 , and others * Serum * Urine * Plasma * Whole blood * Artificial urine Usually simple sample preparation, small sample volume (10 μl) and small solvent consumption, low price, some methods use common screen-printed carbon electrode Standard addition method is not suitable in clinical analysis, necessity of electrode, nanocomposite or carbon nanosheet preparation, technique is research only –not commercially available, electrodes are prepared in laboratory, higher detection limits compared with modified electrodes, some methods use large sample volumes (10 ml) (675,370)2018–2023 Microbiological test kits 2·87−14·57 B 6 -P5P B 7 * Serum Small sample volume (50 μl) High price (working in duplicate recommended), usually long analysis time (24 h), ATBs in patients’ sample could influence results (676,372) HPLC-FLD kits 1·62–4·21 B 6 -P5P, and others * Plasma * Whole blood * Serum Small sample volume (100–300 μl) No IS is used, long analysis time, different extraction procedures for each vitamin, different analysis conditions (temperature etc.), high price for small sample series (677,374)2021 LC-MS/MS kits 1·5–6·96 B 6 -PL, B 6 -P5P, and others * Whole blood Methods use IS, MRM, there is short analysis time, small sample volume (50 μl) combined with simple sample preparation High price for small sample series (678)2021 ELISA kits 0·13−51·16 ×10−3B 7 * Serum * Plasma * Urine Methods use small sample volume (50–250 μl). One kit is suitable for various matrices, high sensitivity Methods are for research only, cross-reactivity with analogues, time consuming methods with high price for small sample series (679,377)2021 B 6 , pyridoxine; B 6 -PL, pyridoxal; B 6 -P5P, pyridoxal-5-phosphate; B 6 -PM, pyridoxamine; B 6 -PA, pyridoxic acid; B 7 , biotin; B 9 , folic acid; ATB, antibiotic; CL, chemiluminescence; ECD, electrochemical detection; ELISA, enzyme-linked immunosorbent assay; FLD, fluorescence detection; HPLC, high performance liquid chromatography; IS, internal standard; LC-MS, coupling of liquid chromatography and mass spectrometry; MLC, micellar liquid chromatography; MRM, multiple reaction monitoring; MS, mass spectrometer; MS/MS, tandem mass spectrometry; PDA, photodiode array detection; SIM, selected ion monitoring; 2D-LC, two-dimensional liquid chromatography. 8 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press pyrroline-5-carboxylate, a compound structurally similar to 1-piperideine-6-carboxylic acid, that also leads to the inactivation of PLP(337,338). •Mutations in the PNPO gene influencing PLP recycling and synthesis(339–341). •Mutations in the pyridoxal phosphate-binding protein (PLPBP) gene (formerly called proline synthetase co-transcribed homolog). PLPBP protects PLP from damage by intracellular phosphatases(342–345). •Mutations in the ALPL gene encoding tissue non-specific alkaline phosphatase (TNSALP) in the metabolic disorder hypophosphatasia(346). •Mutations in the PIGV,PIGO and PGAP2 genes responsible for the development of hyperphosphatasia with seizures and neurologic deficit (Mabry syndrome). These genes play a crucial role in the production of the glycosylphosphatidylinositol anchor that binds TNSALP to the cell membrane. Mutations result in the production of non-functional glycosylphosphatidylinositol anchors and the subsequent release of TNSALP into the blood(347). These metabolic diseases associated with defects in vitamin B 6 are summarised in Table 3. Clinically used drugs as antivitamins for B 6 In addition to natural antivitamins for B 6 , there are also certain clinically used drugs that have the same effect. Drugs such as theophylline (a bronchodilator used in the treatment of respiratory diseases, for example, asthma) and caffeine (psychostimulant) directly inhibit pyridoxal kinase, the enzyme involved in activation of PLP. In the case of caffeine, such effects are probable solely in intoxication. The result is a PLP deficiency with accompanying reduction in PLP-dependent enzyme activities. Known consequences include neurotoxic reactions, for example, peripheral neuropathy, restlessness, agitation, tremors and seizures(348–350).It should be mentioned that standardised extracts from Ginkgo biloba are easily available and used in the therapy of a number of conditions, such as peripheral circulatory disturbances, dizziness and tinnitus, etc.(98,351,352). Hydrazine derivatives, beyond the aforementioned gyromitrin, also include the antituberculosis drug isoniazid (isonicotinic acid hydrazide). Administration of this drug, particularly in overdose, results in not only the inhibition of pyridoxal kinase by the isoniazid metabolite (hydrazone) but also the inactivation of PLP by other isoniazid metabolites (hydrazines and hydrazides), that form, for example, isonicotinilhydrazide, a compound that is easily excreted in the urine(353).Another antituberculosis drug, cycloserine, reacts with PLP forming covalent complexes that might inhibit pyridoxal kinase(354). Another group of drugs, including penicillamine and levodopa, form complexes with PLP, but they do not inhibit pyridoxal kinase(355,356). In addition, antiepileptic drugs (phenytoin, valproic acid, and carbamazepine) increase the metabolism of vitaminB 6 vitamers, resulting in low PLP plasma levels(357). Dietary recommendation and pharmacological use of vitamin B 6 Vitamin B 6 is available in both multivitamin preparations with other B vitamins and as a single vitamin preparation. Oral tablets or solutions for parenteral (intravenous, intramuscular) administration are the most common forms; they usually contain pyridoxine hydrochloride or sometimes PLP. In adults, the current recommended dietary allowances range between 1·3–2·0 mg/d. During pregnancy, lactation and in the elderly, this requirement is increased(286). Recommendations for pyridoxine intake according to age and gender are listed in Table 4. As a supplement, vitamin B 6 is used especially in cases of its deficiency, which may be due to insufficient intake or increased need, as specified earlier. As a medication, pyridoxine or PLP are given prophylactically or therapeutically to patients with pyridoxine-dependent epilepsy. In newborns with hereditary syndrome, it is necessary to administer this vitamin in the first week of life to prevent mental retardation or anaemia, and lifelong therapy is necessary. In the literature, however, there is a lack of congruence regarding dose recommendations. The optimal dosage should Table 3. Inborn metabolic disorders related to pyridoxine dependent seizures Disease (synonyms) Genetic defect Other symptoms except seizures Antiquitin deficiency (pyridoxine-dependent seizures) ALDH7A1 Developmental delay, intellectual disability, abdominal distention Hyperprolinaemia II (pyrroline carboxylate dehydrogenase deficiency) ALDH4A1 Developmental delay, mental retardation PNPO deficiency (pyridoxamine5´- phosphate oxidase deficiency) PNPO Developmental delay, sideroblastic anaemia, microcephaly, feeding difficulties PLPBP deficiency PLPBP Developmental delay, intellectual disability, microcephaly, anaemia Hypophosphatasia ALPL Impaired calcification of bones/teeth, anaemia, respiratory insufficiency Hyperphosphatasia (Mabry syndrome) PIGV, PIGO, PGAP2 Mental retardation, intellectual disability, facial dysmorphism, brachytelephalangy, anal stenosis Table 4. Recommendations for vitamin B 6 intake by gender and age(286) Individuals Condition, age Dose (mg/d) Men 19–50 years 1·3 >51 years 1·7 Women 19–50 years 1·3 >51 years 1·5 pregnancy 1·9 lactation 2·0 Children up to 6 months 0·1 7–11 months 0·3 1–3 years 0·5 4–8 years 0·6 9–13 years 1·0 Adolescent male 14–18 years 1·3 Adolescent female 14–18 years 1·2 Biological properties of vitamins of the B-complex 9 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press increases pancreatic glucokinase activity in rat pancreatic islet primary cultures, and 100 nM biotin duplicated the activity observed in controls. Also, glucokinase mRNA levels increased by ~80% after incubation with 1 μM biotin during 24 h(600). Moreover, a study using rat hepatocytes demonstrated that the addition of biotin (1 μM) to the culture medium induces a threefold increase in the content of cGMP and a fourfold increase in glucokinase activity and mRNA levels(601). Thus, both pancreatic and hepatic glucokinase are regulated by biotin in a positive manner(600,601). Regarding the underlying mechanisms involved in gene regulation, biotinyl-AMP, which is the intermediary product formed by the action of holocarboxylase synthetase, is thought to be responsible for the gene regulatory functions of biotin. BiotinylAMP activates the soluble guanylate cyclase with a subsequent increase in intracellular concentrations of cGMP and activation of protein kinase G(602,603). This signal transduction pathway is involved in the regulation of genes involved in biotin homeostasis and function, including biotin-dependent carboxylases and holocarboxylase synthetase, SMVT and others (for example, the asialoglycoprotein receptor and oncogenes). Holocarboxylase synthetase mRNA levels in the liver, kidney, muscle and brain of rats fed a biotin-deficient diet were significantly lower compared with the controls. Conversely, pyruvate and propionyl CoA carboxylase mRNA levels were not altered, while the amounts of Fig. 5. Physiological function of biotin. (a) Schematic representation of the biotin cycle. Free biotin binds covalently to five apocarboxylases: propionyl-CoA carboxylase (PCC), methylcrotonyl-CoA carboxylase (MCC), pyruvate carboxylase (PC) and acetyl-CoA carboxylases (ACC-1 and ACC-2), by the action of biotin holocarboxylase synthetase. This step requires ATP and gives rise to active holocarboxylases, which are important in amino acid catabolism, the synthesis and oxidation of fatty acids and gluconeogenesis. When needed, holocarboxylases can be proteolysed to biocytin. Then, biotinidase allows the release of free biotin. Adapted from(378,586).(b) Simplified scheme of a human mitochondrion and biotin-dependent carboxylases, their role and location (cytosol, outer membrane and matrix). ACC-1, acetyl–CoA carboxylase 1; ACC-2, acetyl-CoA carboxylase 2; PCC, propionyl– CoA carboxylase; MCC, methylcrotonyl-CoA carboxylase; PC, pyruvate carboxylase; aa, amino acids (valine, isoleucine, methionine, threonine). Adapted from(649). 16 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press these enzymes were lower.(604)Biotin was also identified as the factor required for the expression of the asialoglycoprotein receptor in the human liver cancer cell line HepG2(605). Moreover, in vitro studies demonstrated that expression of oncogenes (N-myc, c-myb, N-ras and raf) correlate positively with biotin concentrations. A pharmacological concentration of biotin (10 nM) increased the expression of N-myc to 120%, whereas a very low biotin concentration (25 pM) decreased it to 53% compared with the controls containing biotin at a physiological concentration (250 pM)(606). Moreover, some studies indicate that biotin-dependent genes are clustered in specific chromosomes(607). Biotin as anti-inflammatory and immunomodulator Recent studies also reported a link between biotin and immune and inflammatory functions. Mice not-absorbing biotin due to knockout of the SMVT gene revealed chronic inflammation in the cecum(608). Biotin-deficient human monocyte–derived dendritic cells demonstrated a higher secretion of cytokines such as tumour necrosis factor α(TNF-α), IL-12p40, IL-23 and IL-1β(609). Biotinidase deficiency, an inborn disorder characterised by impaired biotin bioavailability and recycling, can be associated with weakened immunity that manifests as recurrent infections and dermatitis(610). Indeed, biotin deficiencies are often associated with skin manifestations(611). The underlying pathophysiological mechanisms could be alterations in the role of biotin-dependent carboxylases, such as acetyl–CoA carboxylase 1 or propionyl-CoA carboxylase, interfering with fatty acid metabolism and the cutaneous immune system. Moreover, immune and inflammatory functions of biotin cannot be explained solely by its involvement in carboxylation but also via its effects on transcriptional factors such as nuclear factor κB and Sp1/3(599). Nuclear factor κB regulates genes involved in inflammation and innate and adaptive immune response. Sp1 and Sp3 have been associated with the expression of the gene encoding the cytokine IL-10(612). Laboratory assessment of biotin status Methods for measurement of biotin are summarised in Table 2 (more details are shown in Supplementary Data Table S2). Indicators of biotin status could be helpful in the diagnosis of conditions associated with biotin deficiency. Stratton et al. identified lymphocyte propionyl-CoA carboxylase (PCC) activity as an indicator of biotin deficiency in human subjects(613). However, due to analytical issues, the PCC assay is not adequate to assess biotin status in large population studies. Another marker is the plasma level of 3-hydroxyisovaleryl carnitine. This might be an early and sensitive indicator of biotin deficiency in humans(614). Moreover, urinary 3-hydroxyisovaleryl carnitine could also be used(615). Biotin deficiency and related disorders Frank biotin deficiency cases were reported in people who consume raw egg white for long periods, in cases of parenteral nutrition, and inborn errors of metabolism that cause biotin wasting(379,616). Nutritional biotin deficiency and inherited disorders associated with gene mutations encoding holocarboxylase synthetase or biotinidase give rise to a pathological state called multiple carboxylase deficiency. The pathophysiological mechanisms include: (1) in the case of holocarboxylase synthetase deficiency, a decrease in the affinity of holocarboxylase synthetase for biotin with consequent impairment in the formation of holocarboxylases at physiological biotin levels, and (2) in the case of biotinidase deficiency, alterations in biotin release from its protein conjugates and hence its recycling (Fig. 5a). This is followed by its loss in urine as biocytin. The estimated incidence of biotinidase deficiency is ~ 1:60 000–80 000 of newborns, whereas holocarboxylase synthetase deficiency is estimated to be less than 1:200 000 of newborns. Both are autosomal recessive disorders. Clinical manifestations of biotinidase deficiency include seizures, hypotonia, lack of coordinated movement and balance impairment, respiratory problems, hearing and vision loss, skin rashes, hair loss and retarded cognitive and physical development. Holocarboxylase synthetase deficiency symptoms include severe metabolic acidosis, lethargy, hypotonia, vomiting, seizures, hypothermia, unconsciousness, and even coma and death. All these clinical manifestations from both disorders respond well to early treatment through biotin supplementation(617). The dose needed for the treatment is not excessive, and the onset of the effect is relatively rapid. In a case report of two Chinese infants with late-onset holocarboxylase synthetase deficiency, 30 mg/d biotin treatment in the initial phase solved the metabolic disorders within 48h. Moreover, in the following period, biotin supplementation improved the patient’s clinical condition(618). Biotin-thiamine-responsive basal ganglia disease is a rare autosomal recessive neurometabolic disorder. Formerly, it was Table 7. Biotin-dependent carboxylases, their location in cells and roles Biotin-dependent carboxylases Location Roles References Acetyl-CoA carboxylase 1 (ACC-1) Cytosol Carboxylation of acetyl-CoA to malonyl-CoA. ACC-1 isoform is expressed in lipogenic tissues such as the liver and kidney (686) Acetyl-CoA carboxylase 2 (ACC-2) Outer mitochondrial membrane Carboxylation of acetyl-CoA to malonyl-CoA. ACC-2 isoform is expressed in skeletal muscles and the heart, tissues where fatty acid oxidation is important (686) Methylcrotonyl-CoA carboxylase (MCC) Mitochondrial matrix Catabolism of leucine and carboxylation of 3-methylcrotonyl-CoA to 3-methylglutaconylCoA (687) Pyruvate carboxylase (PC) Mitochondrial matrix Catalysis of the transformation of pyruvate to oxaloacetate (688) Propionyl-CoA carboxylase (PCC) Mitochondrial matrix Catalysis of the conversion of propionyl-CoA to methylmalonyl-CoA (689) Biological properties of vitamins of the B-complex 17 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press called biotin-responsive basal ganglia disease and described as a subacute encephalopathy, with confusion, dysarthria and dysphagia with occasional supranuclear facial nerve palsy or external ophthalmoplegia that can progress to severe quadriparesis and even death. Symptoms of biotin-thiamine-responsive basal ganglia disease disappear within a few days with biotin treatment (5–10 mg/kg/d), and relapse occurs within 1 month if biotin is discontinued(619). Recent studies have shown that regimens for curing this condition include both biotin and thiamine to treat and prevent acute crises and relapses(620). Recent clinical studies have focused on cases of marginal biotin deficiency. Their incidence is higher than was assumed in the past(615). Logically, the absence of symptoms commonly present in biotin deficiency is not a suitable marker for vitamin B 7 marginal deficiency. Biotin deficiency is teratogenic in several animal species. In mice, egg-induced biotin deficiency caused a higher incidence of cranial malformations and shortening of the long limb bones(621).It need not be emphasised that other vitamins of the B complex, such as folic acid, have already been shown to be essential(622). Hence, the roles and kinetics of biotin in pregnancy deserve attention. A cross-sectional study of normal human gestation reported an increased excretion of 3-hydroxyisovaleric acid in early and late pregnancy. However, there was a paradoxical increase in biotin excretion late in pregnancy, suggesting that biotin status in pregnancy was not reduced(623).Conversely, a longitudinal study with women from early to late pregnancy found evidence that biotin status decreases during pregnancy. By late pregnancy, approximately half of the participants showed less than the lower limit of normal biotin excretion rates(624). Pharmacological use of biotin Pharmacologic doses of biotin are used for treating patients with disorders of biotin metabolism as mentioned in the previous chapter. Holocarboxylase synthetase deficiency can be treated with 10 mg biotin/d with children showing improvement in their condition, while biotinidase deficiency can be treated with a dose of 5–20 mg biotin daily(380). Considering the potential neuroprotective role of biotin, its use in the treatment of neurological diseases could be beneficial. In an open-label study with twenty-three patients with primary and secondary progressive multiple sclerosis, treatment with high-dose biotin (100–300 mg/d) from 2 to 36 months revealed an improvement in several symptoms. Overall, the clinical improvement was delayed by 2–8 months, and 300 mg biotin/d, a 10 000 times higher dose than the recommended daily intake, generated the best clinical response(625). Some mechanisms were suggested to be responsible: (1) activation of pyruvate carboxylase, propionyl– CoA carboxylase and methylcrotonyl-CoA carboxylase may lead to an increase in ATP production in neurons, and (2) activation of acetyl–CoA carboxylases may lead to myelin repair.(625)A doubleblind, placebo-controlled study with 154 patients with primary or secondary progressive multiple sclerosis receiving 100 mg of biotin orally, thrice daily or placebo for 12 months corroborated the previous findings(626). In contrast, in an observational prospective study of 178 patients, again with primary and secondary progressive multiple sclerosis, high-dose biotin did not show a clear improvement in disability and quality of life(627). Regarding dietary biotin supplements, they frequently appear as combinations of the B-complex vitamins or multivitamin complexes. Its main indications are to fortify hair, nails and skin. Although several reports have demonstrated clinical improvement after biotin supplementation in cases of biotin deficiency, research demonstrating its efficacy in hair and nail growth in healthy individuals is limited(628). Biotin is synthesised de novo in plants, fungi and microorganisms, and this property might be used from a therapeutic point of view. For instance, Mycobacterium tuberculosis needs to biosynthesise this vitamin for its pathogenicity during all stages of the life cycle. For this reason, inhibitors of biotin biosynthetic enzymes could be a potential target for the development of novel antibiotics against tuberculosis(629). Toxicity of biotin Owing to being water soluble, excessive amounts of biotin are known to be easily excreted. Hence, it seems this vitamin is relatively non-toxic. Moreover, the maximum daily dose is unlikely to cause adverse side effects in the general population, that is, the tolerable upper intake level, has not yet been established(630,631). Regardless, there are some concerns about high-dose biotin. A case report of a 54-year-old woman with progressive multiple sclerosis reported an aggravation of the neurologic state, accompanied by lipid storage in muscle, after 5 months of treatment with three times 100 mg biotin/d. Symptomsdisappeared in a few months after biotin withdrawal(632). In addition, animal experiments with mice fed with a biotin-supplemented diet (97·7 mg free biotin/kg) over 8 weeks revealed alterations in the testis(633). In addition, experiments with rats fed 5000 and 8000 mg biotin/kg diets for 28 d showed a decrease in testis weight(634). These results are in disagreement with in vitro fertilisation studies in which biotin supplementation to sperm wash medium (2·44 mg/ml) improved the fertilising ability of mice spermatozoa(635). Interferences with tests Elevated blood levels of biotin cause interference in streptavidin– biotin hormone immunoassays(636–638). In competitive assays (for example, triiodothyronine, thyroxine, steroid hormones and 25hydroxyvitamin D), there are falsely increased hormones concentrations, whereas in sandwich assays (for example, glycoprotein regulating hormones), falsely decreased hormones concentrations have been reported(639). The degree of interference is dependent on plasma biotin concentration, and it is significant at concentrations of 30 μg/L or more(637). False hyperthyroidism is the most frequently misdiagnosed endocrine disorder(514,637,640–642). Likewise, false high 25–hydroxyvitamin D serum levels were detected in patients receiving high dose of biotin (>100 mg). Logically, several concerns arise, since this is a crucial laboratory test in multiple sclerosis patients receiving vitamin D supplementation(637,643,644). Biotin interference in cardiac troponin assays has also been reported(645,646). Although, one analysis using a Roche assay led to the conclusion that this interference is rare, and its probability is even lower than other confounders such as blood sample hemolysis and simple biological variation of cardiac troponin(647). The International Federation for Clinical Chemistry Committee on Cardiac Biomarkers (IFCC-CB) reported a cardiac troponin assay interference table for hemolysis and biotin to guide healthcare professionals and clinicians whenever there is an inconsistency between cardiac biomarker results and the clinical situation(645). Biotin supplementation has also been linked to alterations in hepatitis B-virus (HBV), hepatitis C-virus (HCV) and human immunodeficiency virus (HIV)-related serological markers(648).A 18 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press study in which ten healthy volunteers vaccinated against hepatitis B were administered a single oral dose of biotin (100 mg) revealed anti-HB levels below the cutoff value for four of the ten participants. Moreover, around 80–90% of false positive results for anti-HBe and anti-HBc occurred. On the contrary, in HIV and HCV serology testing, biotin caused false negative results(648). Conclusions The current review summarised the literature on two B-group vitamins, B 6 and B 7 (also known as H or biotin). Historically, research interest in these two water-soluble vitamins commenced at the beginning of the twentieth century and allowed for the identification of numerous characteristics and essential roles in multiple physiological functions (for example, neurological and metabolic processes). Novel discoveries, however, revealed their much larger physiological roles beyond their participation in multiple enzymatic reactions. Humans lack biosynthetic pathways for vitamins B 6 and B 7 and, therefore, must obtain them from exogenous sources (for example, foods and supplements). These vitamins are also synthesised by microbiota in the human large intestine, and it is likely that a part of such produced vitamins, at least in the case of biotin, can be absorbed and used by humans. Cases of deficiency of vitamins B 6 and B 7 are rare. However, inherited disorders associated with gene mutations require prompt and lifelong treatment with these vitamins, starting at early life stages (that is, in newborns). Moreover, vitamin B 6 deficiency can follow administration of several clinically used drugs (for example, isoniazid) or poisoning with Gyromitra mushroom and Ginkgo biloba seeds. Beyond cases of deficiency, both vitamins have been used or tested in several other conditions. For instance, pyridoxine has been used for prevention of vomiting in pregnancy, while biotin has been recently tested in controlled trials of patients with primary and secondary progressive multiple sclerosis. Last but not least, both vitamins are considered relatively non-toxic when an adequate intake is followed. Recent research has brought novel discoveries linking both vitamin B 6 and biotin with anti-inflammatory effects in particular. However therapeutic use of both vitamins in various inflammatory disorders still needs much more research. In the case of biotin, further investigation of its role in gene expression regulation through both transcription factors and epigenetic processes is necessary. Supplementary material. To view supplementary material for this article, please visit https://doi.org/10.1017/S0954422425100097 Acknowledgments. We thank Alberto Gomes for his help in formatting the figures. Authorship. P.D. wrote the biological section on biotin. T.S. was responsible for sources of both vitamins. M.V. wrote the biological section on vitamin B 6 . M.M. and J.P. contributed to critical assessment of the paper. N.P., L.K.K. and L.K. prepared and critically assessed chemical and analytical issues. P.M. was responsible for conceptualisation, critical review, finalisation and obtained funding for the open access publication. All authors participated in the critical revision of the article. Financial support. This open-access review paper was supported by the project New Technologies for Translational Research in Pharmaceutical Sciences (NETPHARM), project ID CZ.02.01.01/00/22_008/0004607, and is co-funded by the European Union, and the ErasmusþProgramme of the European Union, Key Action 2: Strategic Partnerships, Project no. 2020-1CZ01-KA203-078218.M.M. sends thanks to Charles University (SVV 260 663). L.J., K.M. and L.K.K. send thanks to MH-CZ-DRO (UHHK, 00179906). Competing interests. The authors have no relevant interests to declare. Ethical standards. Not applicable. Consent for Publication. Not applicable. Availability of data and materials. The following supporting information is available through the online version of this article at the publisher’s website. References 1. Hrubša M, Siatka T, Nejmanová I, et al. (2022) Biological properties of vitamins of the B-complex, part 1: Vitamins B1, B2, B3, and B5. Nutrients 14, 484. 2. McCormick D (2006) Vitamin B6. In B Bowman & R Russell, (eds), Present Knowledge in Nutrition. 9 ed. Washington, DC: International Life Sciences Institute, pp 269–277. 3. Mooney S, Leuendorf J-E, Hendrickson C & Hellmann H (2009) Vitamin B6: a long known compound of surprising complexity. Molecules 14, 329–351. 4. Eliot A & Kirsch J (2004) Pyridoxal phosphate enzymes: mechanistic, structural, and evolutionary considerations. Annu Rev Biochem 73, 383–415. 5. Leklem JE (1990) Vitamin B-6: a status report. J Nutr 120 (suppl_11), 1503–1507. 6. Tanaka T, Tateno Y & Gojobori T (2005) Evolution of vitamin B6 (pyridoxine) metabolism by gain and loss of genes. Mol Biol Evol 22, 243–250. 7. Hellmann H & Mooney S (2010) Vitamin B6: a molecule for human health? Molecules 15, 442–459. 8. Parra M, Stahl S & Hellmann H (2018) Vitamin B6 and its role in cell metabolism and physiology. Cells 7, 84. 9. Tambasco-Studart M, Titiz O, Raschle T, Forster G, Amrhein N & Fitzpatrick TB (2005) Vitamin B6 biosynthesis in higher plants. Proc Natl Acad Sci 102, 13687–13692. 10. Titiz O, Tambasco-Studart M, Warzych E, et al. (2006) PDX1 is essential for vitamin B6 biosynthesis, development and stress tolerance in Arabidopsis. Plant J 48, 933–946. 11. Mooney S & Hellmann H (2010) Vitamin B6: killing two birds with one stone? Phytochemistry 71, 495–501. 12. Fitzpatrick TB (2011) Vitamin B6 in plants: more than meets the eye. Adv Bot Res 59,1–38. 13. Fitzpatrick TB, Amrhein N, Kappes B, Macheroux P, Tews I & Raschle T (2007) Two independent routes of de novo vitamin B6 biosynthesis: not that different after all. Biochem J 407,1–13. 14. Roje S (2007) Vitamin B biosynthesis in plants. Phytochemistry 68, 1904–1921. 15. Gerdes S, Lerma-Ortiz C, Frelin O, et al. (2012) Plant B vitamin pathways and their compartmentation: a guide for the perplexed. J Exp Bot 63, 5379–5395. 16. Leuendorf JE, Osorio S, Szewczyk A, Fernie AR & Hellmann H (2010) Complex assembly and metabolic profiling of Arabidopsis thaliana plants overexpressing vitamin B6 biosynthesis proteins. Mol Plant 3, 890–903. 17. Mittenhuber G (2001) Phylogenetic analyses and comparative genomics of vitamin B6 (pyridoxine) and pyridoxal phosphate biosynthesis pathways. J Mol Microbiol Biotechnol 3,1–20. 18. Kondo H, Nakamura Y, Dong Y-X, Nikawa J-I & Sueda S (2004) Pyridoxine biosynthesis in yeast: participation of ribose 5-phosphate ketol-isomerase. Biochem J 379,65–70. 19. Sakai A, Kita M & Tani Y (2004) Recent progress of vitamin B6 biosynthesis. J Nutr Sci Vitaminol 50,69–77. 20. Tramonti A, Nardella C, di Salvo ML, et al. (2021) Knowns and unknowns of vitamin B6 metabolism in Escherichia coli. EcoSal Plus 9, eESP-0004. Biological properties of vitamins of the B-complex 19 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press 21. Begley TP, Chatterjee A, Hanes JW, Hazra A & Ealick SE (2008) Cofactor biosynthesis—still yielding fascinating new biological chemistry. Curr Opin Chem Biol 12, 118–125. 22. Magnusdottir S, Ravcheev D, de Crecy-Lagard V & Thiele I (2015) Systematic genome assessment of B-vitamin biosynthesis suggests cooperation among gut microbes. Front Genet 6, 148. 23. Liu Z, Farkas P, Wang K, Kohli MO & Fitzpatrick TB (2022) B vitamin supply in plants and humans: the importance of vitamer homeostasis. Pl J 111, 662–682. 24. Stach K, Stach W & Augoff K (2021) Vitamin B6 in health and disease. Nutrients 13, 3229. 25. Meir Z & Osherov N (2018) Vitamin biosynthesis as an antifungal target. J Fungi 4, 72. 26. Scott DE, Ciulli A & Abell C (2007) Coenzyme biosynthesis: enzyme mechanism, structure and inhibition. Nat Prod Rep 24, 1009–1026. 27. Chen H & Xiong L (2009) Enhancement of vitamin B6 levels in seeds through metabolic engineering. Plant Biotechnol J 7, 673–681. 28. da Silva VR & Gregory JF (2020) Chapter 13 –vitamin B6. In BP Marriott, DF Birt, VA Stallings & AA Yates, (eds), Present knowledge in nutrition, 11th ed. London, UK: Academic Press, pp 225–237. 29. EFSA (2016) Dietary reference values for vitamin B6. EFSA J14, e04485. 30. Chawla J and D Kvarnberg (2014) Hydrosoluble vitamins. In J Biller & JM Ferro (eds), Handbook of clinical neurology, neurologic aspects of systemic disease part II, 1st ed. Amsterdam, Netherlands: Elsevier. 120, pp 891–914. 31. Wyness L, Weichselbaum E, O’connor A, et al. (2011) Red meat in the diet: an update. Nutr Bull 36,34–77. 32. Pereira PMdCC & Vicente AFdRB (2013) Meat nutritional composition and nutritive role in the human diet. Meat Sci 93, 586–592. 33. Iglesia I, Mouratidou T, Gonzalez-Gross M, et al. (2017) Foods contributing to vitamin B6, folate, and vitamin B 12 intakes and biomarkers status in European adolescents: the HELENA study. Eur J Nutr 56, 1767–1782. 34. Kraemer K, Semba RD, Eggersdorfer M & Schaumberg DA (2012) Introduction: the diverse and essential biological functions of vitamins. Ann Nutr Metab 61, 185–191. 35. Kennedy DO (2016) B vitamins and the brain: mechanisms, dose and efficacy—a review. Nutrients 8, 68. 36. Geiker NRW, Bertram HC, Mejborn H, et al. (2021) Meat and human health—current knowledge and research gaps. Foods 10, 1556. 37. McDowell LR (2000) Vitamin B6. In LR McDowell (author), Vitamins in animal and human nutrition, 2nd ed. Ames, USA: Iowa state university Press, pp 385–417.* 38. Li C (2017) The role of beef in human nutrition and health. In ME Dikeman (ed), Ensuring safety and quality in the production of beef. 1st ed., Vol 2. Cambridge, UK: Burleigh Dodds Science Publishing, pp 329–338. 39. Ahmad RS, Imran A & Hussain MB (2018) Nutritional composition of meat. In MS Arshad (ed), Meat science and nutrition. London, UK: IntechOpen Limited. https://www.intechopen.com/chapters/61245 Available from: http://dx.doi.org/10.5772/intechopen.77045. 40. Hall APB, Moore JG & Morgan AF (1956) Vitamin content of avocados. Calif Agric 10,13–14. 41. Mielgo-Ayuso J, Aparicio-Ugarriza R, Olza J, et al. (2018) Dietary intake and food sources of niacin, riboflavin, thiamin and vitamin B6 in a representative sample of the Spanish population. The ANIBES study. Nutrients 10, 846. 42. Neufingerl N & Eilander A (2020) Nutrient intake and status in adults consuming plant-based diets compared to meat-eaters: a systematic review. Nutrients 14, 29. 43. Yoshino K, Inagawa M, Oshima M, et al. (2005) Trends in dietary intake of folate, vitamins B6, and B12 among Japanese adults in two rural communities from 1974 through 2001. J Epidemiol 15,29–37. 44. Jungert A, Linseisen J, Wagner K-H, Richter M & German Nutrition Society (2020) Revised DA-CH reference values for the intake of vitamin B6. Ann Nutr Metab 76, 213–222. 45. Shewry PR & Hey SJ (2015) The contribution of wheat to human diet and health. Food Energy Secur 4, 178–202. 46. Waldmann A, Dörr B, Koschizke J, Leitzmann C & Hahn A (2006) Dietary intake of vitamin B6 and concentration of vitamin B6 in blood samples of German vegans. Public Health Nutr 9, 779–784. 47. Köseoğlu SZA (2020) Determination and evaluation of the pyridoxal, pyridoxine, and pyridoxamine forms of vitamin B-6 in plant-based foods in terms of healthy vegetarian nutrition. Prog Nutr 22, e2020015. 48. Yaman M (2019) Determination and evaluation in terms of healthy nutrition of the pyridoxal, pyridoxine and pyridoxamine forms of vitamin B6 in animal-derived foods. Eur J Sci Technol, 2019, 611–617. https://de rgipark.org.tr/tr/download/article-file/683971. 49. Melanson KJ, Angelopoulos TJ, Nguyen VT, et al. (2006) Consumption of whole-grain cereals during weight loss: effects on dietary quality, dietary fiber, magnesium, vitamin B-6, and obesity. J Am Diet Assoc 106, 1380–1388. 50. Planells E, Sanchez C, Montellano M, Mataix J & Llopis J (2003) Vitamins B6 and B12 and folate status in an adult Mediterranean population. Eur J Clin Nutr 57, 777–785. 51. EFSA Panel on Dietetic Products, Nutrition and Allergies (2016) Allergies. Dietary reference values for vitamin B6. EFSA J 14, e04485. 52. Rybicka I & Gliszczynska-Swiglo A (2017) Gluten-free flours from different raw materials as the source of vitamin B1, B2, B3 and B6. J Nutr Sci Vitaminol 63, 125–132. 53. Kamyab S, Gharachorloo M, Honarvar M & Ghavami M (2020) Quantitative analysis of bioactive compounds present in Iranian royal jelly. J Apic Res 59,42–52. 54. Ollilainen V-M (1999) HPLC analysis of vitamin B6 in foods. Agric Food Sci 8, 515–619. 55. Ndaw S, Bergaentzle M, Aoude-Werner D & Hasselmann C (2000) Extraction procedures for the liquid chromatographic determination of thiamin, riboflavin and vitamin B6 in foodstuffs. Food Chem 71, 129–138. 56. Schmidt A, Schreiner M & Mayer H (2017) Rapid determination of the various native forms of vitamin B6 and B2 in cow’s milk using ultra-high performance liquid chromatography. J Chromator A 1500,89–95. 57. Reynolds RD (1988) Bioavailability of vitamin B-6 from plant foods. Am J Clin Nutr 48, 863–867. 58. Ink SL, Gregory III JF & Sartain DB (1986) Determination of pyridoxine beta-glucoside bioavailability using intrinsic and extrinsic labeling in the rat. J Agric Food Chem 34, 857–862. 59. Gregory III J, Ink S & Sartain D (1986) Degradation and binding to food proteins of vitamin B-6 compounds during thermal processing. J Food Sci 51, 1345–1351. 60. EFSA (2023) Scientific opinion on the tolerable upper intake level for vitamin B6. EFSA J 21, e08006. 61. Kabir H, Leklem JE & Miller LT (1983) Comparative vitamin B-6 bioavailability from tuna, whole wheat bread and peanut butter in humans. J Nutr 113, 2412–2420. 62. Gregory III JF (1998) Nutritional properties and significance of vitamin glycosides. Annu Rev Nutr 18, 277–296. 63. Gregory III JF & Ink SL (1987) Identification and quantification of pyridoxine beta-glucoside as a major form of vitamin B6 in plant-derived foods. J Agric Food Chem 35,76–82. 64. Gregory III JR (1997) Bioavailability of vitamin B-6. Eur J Clin Nutr 51, S43–S8. 65. Kabir H, Leklem J & Miller LT (1983) Measurement of glycosylated vitamin B6 in foods. J Food Sci 48, 1422–1425. 66. Kall MA (2003) Determination of total vitamin B6 in foods by isocratic HPLC: a comparison with microbiological analysis. Food Chem 82, 315–327. 67. Bachmann T, Maurer A & Rychlik M (2020) Development of a LC-MS/ MS method using stable isotope dilution for the quantification of individual B6 vitamers in fruits, vegetables, and cereals. Anal Bioanal Chem 412, 7237–7252. 68. Mangel N, Fudge JB, Gruissem W, Fitzpatrick TB & Vanderschuren H (2022) Natural variation in vitamin B1 and vitamin B6 contents in rice germplasm. Front Plant Sci 13, 856880. 69. Gregory III JF, Trumbo PR, Bailey LB, Toth JP, Baumgartner TG & Cerda JJ (1991) Bioavailability of pyridoxine-50-β-d-glucoside determined in humans by stable-isotopic methods. J Nutr 121, 177–186. 20 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press 70. Gregory III J (2012) Accounting for differences in the bioactivity and bioavailability of vitamers. Food Nutr Res 56, 5809. 71. Bird RP (2018) The emerging role of vitamin B6 in inflammation and carcinogenesis. Adv Food Nutr Res 83, 151–194. 72. SokačDG, KovačS, Buši´c V, Martin CR & FuračJV (2012) The chemistry and biochemistry of vitamin B6: synthesis of novel analogues of vitamin B6. In Preedy VR (ed), B vitamins and folate: chemistry, analysis, function and effects. Cambridge, UK: The Royal Society of Chemistry. pp 135–145. 73. Nakano H & McMahon LG & Gregory III JF (1997) Pyridoxine-50-βD - glucoside exhibits incomplete bioavailability as a source of vitamin B-6 and partially inhibits the utilization of co-ingested pyridoxine in humans. J Nutr 127, 1508–1513. 74. Armada LJ, Mackey AD & Gregory III JF (2002) Intestinal brush border membrane catalyzes hydrolysis of pyridoxine-50-β-d-glucoside and exhibits parallel developmental changes of hydrolytic activities toward pyridoxine-50-β-d-glucoside and lactose in rats. J Nutr 132, 2695–2699. 75. Roth-Maier DA, Kettler SI & Kirchgessner M (2002) Availability of vitamin B6 from different food sources. Int J Food Sci Nutr 53, 171–179. 76. Tsuge H, Maeno M, Hayakawa T & Suzuki Y (1996) Comparative study of pyridoxine-α,β-glucosides, and phosphopyridoxyl-lysine as a vitamin B6 nutrient. J Nutr Sci Vitaminol 42, 377–386. 77. Mackey AD, Lieu SO, Carman C & Gregory III JF (2003) Hydrolytic activity toward pyridoxine-50-βD -glucoside in rat intestinal mucosa is not increased by vitamin B-6 deficiency: effect of basal diet composition and pyridoxine intake. J Nutr 133, 1362–1367. 78. Hansen CM, Leklem JE & Miller LT (1996) Vitamin B-6 status indicators decrease in women consuming a diet high in pyridoxine glucoside. J Nutr 126, 2512–2518. 79. Schorgg P, Bärnighausen T, Rohrmann S, Cassidy A, Karavasiloglou N & Kühn T (2021) Vitamin B6 status among vegetarians: findings from a population-based survey. Nutrients 13, 1627. 80. Basu TK & Donaldson D (2003) Intestinal absorption in health and disease: micronutrients. Best Pract Res Clin Gastroenterol 17, 957–979. 81. EFSA (2017) Dietary reference values for nutrients. Summary Report EFSA Supporting Publication 14, p e15121. 82. Yaman M, Çatak J, Uğur H, et al. (2021) The bioaccessibility of watersoluble vitamins: a review. Trends Food Sci Technol 109, 552–563. 83. Strain J, Hughes C, Pentieva K, Ward M, Hoey L & McNulty H (2017) The B-vitamins. In HK Biesalski, A Drewnowski, JT Dwyer, J Strain, P Weber & M Eggersdorfer (eds), Sustainable nutrition in a changing world, 1st ed. Cham, Switzerland: Springer International Publishing AG, pp 185–205. 84. Spinneker A, Sola R, Lemmen V, Castillo M, Pietrzik K & Gonzalez-Gross M (2007) Vitamin B6 status, deficiency and its consequences-an overview. Nutr Hosp 22,7–24. 85. Yaman M & Mızrak ÖF (2019) Determination and evaluation of in vitro bioaccessibility of the pyridoxal, pyridoxine, and pyridoxamine forms of vitamin B6 in cereal-based baby foods. Food Chem 298, 125042. 86. Tarr JB, Tamura T & Stokstad EL (1981) Availability of vitamin B6 and pantothenate in an average American diet in man. Am J Clin Nutr 34, 1328–1337. 87. Combs Jr G & McClung J (2017) Chapter 14-vitamin B6. In GF Combs & JP McClung (authors), The vitamins, 5th ed. Fundamental Aspects in Nutrition and Health, London, UK: Academic Press. pp 351–370. 88. Said HM (2011) Intestinal absorption of water-soluble vitamins in health and disease. Biochem J 437, 357–372. 89. Said ZM, Subramanian VS, Vaziri ND & Said HM (2008) Pyridoxine uptake by colonocytes: a specific and regulated carrier-mediated process. Am J Physiol-Cell Physiol 294, C1192–C1197. 90. Uebanso T, Shimohata T, Mawatari K & Takahashi A (2020) Functional roles of B-vitamins in the gut and gut microbiome. Mol Nutr Food Res 64, 2000426. 91. Yoshii K, Hosomi K, Sawane K & Kunisawa J (2019) Metabolism of dietary and microbial vitamin B family in the regulation of host immunity. Front Nutr 6, 48. 92. Rowland I, Gibson G, Heinken A, et al. (2018) Gut microbiota functions: metabolism of nutrients and other food components. Eur J Nutr 57,1–24. 93. Rodionov DA, Arzamasov AA, Khoroshkin MS, et al. (2019) Micronutrient requirements and sharing capabilities of the human gut microbiome. Front Microbiol 10, 1316. 94. Mayengbam S, House J & Aliani M (2016) Investigation of vitamin B inadequacy, induced by exposure to the anti-B factor 1-amino D-proline, on plasma lipophilic metabolites of rats: a metabolomics approach. Eur J Nutr 55, 1213–1223. 95. Wada K, Ishigaki S, Ueda K, Sakata M & Haga M (1985) An antivitamin B6, 4’-methoxypyridoxine, from the seed of Ginkgo biloba L. Chem Pharm Bull 33, 3555–3557. 96. Zou M, Zhang W, Wu R, Jiang H, Cao F & Su E (2021) Removal of ginkgotoxin from the Ginkgo biloba seeds powder by adopting membrane separation technology. J Clean Prod 280, 124452. 97. Kosaki Y, Naito H, Nojima T & Nakao A (2020) Epileptic seizure from ginkgo nut intoxication in an adult. Case Rep Emerg Med 2020,ID 5072954. 98. Azuma F, Nokura K, Kako T, Kobayashi D, Yoshimura T & Wada K (2020) An adult case of generalized convulsions caused by the ingestion of Ginkgo biloba seeds with alcohol. Intern Med 59, 1555–1558. 99. Kobayashi D, Yoshimura T, Johno A, Ishikawa M, Sasaki K & Wada K (2015) Decrease in pyridoxal-50-phosphate concentration and increase in pyridoxal concentration in rat plasma by 40-O-methylpyridoxine administration. Nutr Res 35, 637–642. 100. Kobayashi D, Yoshimura T, Johno A, Sasaki K & Wada K (2011) Toxicity of 40-O-methylpyridoxine-50-glucoside in Ginkgo biloba seeds. Food Chem 126, 1198–1202. 101. Leistner E & Drewke C (2010) Ginkgo biloba and ginkgotoxin. J Nat Prod 73,86–92. 102. Kajiyama Y, Fujii K, Takeuchi H & Manabe Y (2002) Ginkgo seed poisoning. Pediatrics 109, 325–327. 103. Arenz A, Klein M, Fiehe K, et al. (1996) Occurrence of neurotoxic 40-Omethylpyridoxine in Ginkgo biloba leaves, Ginkgo medications and Japanese Ginkgo food. Planta Med 62, 548–551. 104. Buss K, Drewke C, Lohmann S, Piwonska A & Leistner E (2001) Properties and interaction of heterologously expressed glutamate decarboxylase isoenzymes GAD65kDa and GAD67kDa from human brain with ginkgotoxin and its 5´-phosphate. J Med Chem 44, 3166–3174. 105. Kästner U, Hallmen C, Wiese M, Leistner E & Drewke C (2007) The human pyridoxal kinase, a plausible target for ginkgotoxin from Ginkgo biloba.FEBS J 274, 1036–1045. 106. Samuels N, Finkelstein Y, Singer SR & Oberbaum M (2008) Herbal medicine and epilepsy: proconvulsive effects and interactions with antiepileptic drugs. Epilepsia 49, 373–380. 107. Mei N, Guo X, Ren Z, Kobayashi D, Wada K & Guo L (2017) Review of Ginkgo biloba-induced toxicity, from experimental studies to human case reports. J Environ Sci Health, Part C 35,1–28. 108. Zhu J-P, Gong H, Labreche F, et al. (2021) In vivo toxicity assessment of 40-O-methylpyridoxine from Ginkgo biloba seeds: growth, hematology, metabolism, and oxidative parameters. Toxicon 201,66–73. 109. Zhu J-P, Gong H, Wu C-E, Fan G-J, Li T-T & Wang J-H (2021) 40-Omethylpyridoxine: preparation from Ginkgo biloba seeds and cytotoxicity in GES-1 cells. Toxins (Basel) 13, 95. 110. Zhang W, Shi M, Zhang F, Cao F & Su E (2021) A facile method to determine the native contents of 40-O-methylpyridoxine and 40-Omethylpyridoxine-50-glucoside in Ginkgo biloba seeds. J Agric Food Chem 69, 14270–14277. 111. Lim HB & Kim D (2018) Effect of heat treatment on 40-Omethylpyridoxine (MPN) content in Ginkgo biloba seed extract solution. J Sci Food Agric 98, 5153–5156. 112. Gong H, Wu C-E, Fan G-J, Li T-T, Wang J-H & Wang T (2018) Determination and comparison of 40-O-methylpyridoxine analogues in Ginkgo biloba seeds at different growth stages. J Agric Food Chem 66, 7916–7922. 113. Steyn PS, Vleggaar R & Anderson LA (1987) Structure elucidation of two neurotoxins from Albizia tanganyicensis. S Afr J Chem 40, 191–192. 114. Mayengbam S, Raposo S, Aliani M & House JD (2015) Oral exposure to the anti-pyridoxine compound 1-amino d-proline further perturbs Biological properties of vitamins of the B-complex 21 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press homocysteine metabolism through the transsulfuration pathway in moderately vitamin B6 deficient rats. J Nutr Biochem 26, 241–249. 115. Mayengbam S, Raposo S, Aliani M, House JD (2016) A vitamin B-6 antagonist from flaxseed perturbs amino acid metabolism in moderately vitamin B-6-deficient male rats. J Nutr 146,14–20. 116. Mayengbam S, Yang H, Barthet V, Aliani M & House JD (2014) Identification characterization, and quantification of an anti-pyridoxine factor from flaxseed using ultrahigh-performance liquid chromatography–mass spectrometry. J Agric Food Chem 62, 419–426. 117. Sasaoka K, Ogawa T, Moritoki K & Kimoto M (1976) Antivitamin B-6 effect of 1-aminoproline on rats. Biochim Biophys Acta Gen Sub 428, 396–402. 118. Klosterman HJ (1974) Vitamin B6 antagonists of natural origin. J Agric Food Chem 22,13–16. 119. Erogul M, Ernsting K & Savitt D (2001) Gyromitra mushrooms. In MA McGuigan, A. Anderson & A Woolf A (eds), Clin Toxicol Rev 23. Boston, USA: Massachusetts/Rhode Island Poison Control System. 120. White J, Weinstein SA, De Haro L, et al. (2019) Mushroom poisoning: a proposed new clinical classification. Toxicon 157,53–65. 121. Lagrange E & Vernoux J-P (2020) Warning on false or true morels and button mushrooms with potential toxicity linked to hydrazinic toxins: an update. Toxins (Basel) 12, 482. 122. Horowitz KM & Kong EL & Horowitz BZ (2017) Gyromitra mushroom toxicity. StatPearls Publishing. Available from: https://www.ncbi.nlm.nih. gov/books/NBK430685/. 123. Berdanier CD (2007) Food constituents. In CD Berdanier, JT Dwyer & EB Feldman (eds), Handbook of nutrition and food. Boca Raton, USA: CRC Press, pp 3–35. 124. Arshadi M, Nilsson C & Magnusson B (2006) Gas chromatography–mass spectrometry determination of the pentafluorobenzoyl derivative of methylhydrazine in false morel (Gyromitra esculenta) as a monitor for the content of the toxin gyromitrin. J Chromator A 1125, 229–233. 125. Roupas P, Keogh J, Noakes M, Margetts C & Taylor P (2010) Mushrooms and agaritine: a mini-review. J Funct Foods 2,91–98. 126. Klosterman HJ (1979) Vitamin B6 antagonists of natural origin. Methods Enzymol 62, 483–495. 127. Negi VS, Pal A & Borthakur D (2021) Biochemistry of plants N– heterocyclic non-protein amino acids. Amino Acids 53, 801–812. 128. Honda MD & Borthakur D (2019) Mimosine concentration in Leucaena leucocephala under various environmental conditions. Trop Grass-Forra Trop 7, 164–172. 129. Oogai S, Fukuta M, Inafuku M & Oku H (2022) Isolation and characterization of mimosine degrading enzyme from Arthrobacter sp. Ryudai-S1. World J Microbiol Biotechnol 38, 172. 130. Rosenthal GA (1981) A mechanism of L-canaline toxicity. Eur J Biochem 114, 301–304. 131. Rosenthal GA (1997) L-canaline: a potent antimetabolite and anti-cancer agent from leguminous plants. Life Sci 60, 1635–1641. 132. Gregory III J & Kirk J (1981) The bioavailability of vitamin B6 in foods. Nutr Rev 9,1–8. 133. Özilgen S & Özilgen M (2017) General template for the FMEA applications in primary food processing. In B Hitzmann (ed), Measurement, modeling and automation in advanced food processing. Cham: Springer International Publishing, pp 29–69. 134. Berry Ottaway P (2010) Stability of vitamins during food processing and storage. In LH Skibsted, J Risbo & ML Andersen (eds), Chemical deterioration and physical instability of food and beverages. Cambridge, UK: Woodhead Publishing, pp 539–560. 135. Godoy HT, Amaya-Farfan J & Rodriguez-Amaya DB (2021) Degradation of vitamins. In DB Rodriguez-Amaya & J Amaya-Farfan (eds), Chemical changes during processing and storage of foods. London, UK: Academic Press, pp 329–383. 136. Slavin JL, Jacobs D & Marquart L (2000) Grain processing and nutrition. Crit Rev Food Sci Nutr 40, 309–326. 137. Thielecke F, Lecerf J-M & Nugent AP (2021) Processing in the food chain: do cereals have to be processed to add value to the human diet? Nutr Res Rev 34, 159–173. 138. Călinoiu LF & Vodnar DC (2018) Whole grains and phenolic acids: a review on bioactivity, functionality, health benefits and bioavailability. Nutrients 10, 1615. 139. Batifoulier F, Verny M-A, Chanliaud E, Remesy C & Demigné C (2006) Variability of B vitamin concentrations in wheat grain, milling fractions and bread products. Eur J Agron 25, 163–169. 140. Lebert L, Buche F, Sorin A & Aussenac T (2022) The wheat aleurone layer: optimisation of its benefits and application to bakery products. Foods 11, 3552. 141. Brouns F, Hemery Y, Price R & Anson NM (2012) Wheat aleurone: separation, composition, health aspects, and potential food use. Crit Rev Food Sci Nutr 52, 553–568. 142. Titcomb TJ & Tanumihardjo SA (2019) Global concerns with B vitamin statuses: biofortification, fortification, hidden hunger, interactions, and toxicity. Compr Rev Food Sci Food Saf 18, 1968–1984. 143. Garg M, Sharma A, Vats S, et al. (2021) Vitamins in cereals: a critical review of content, health effects, processing losses, bioaccessibility, fortification, and biofortification strategies for their improvement. Front Nutr 8, 586815. 144. Hegedüs M, Pedersen B & Eggum BO (1985) The influence of milling on the nutritive value of flour from cereal grains. 7. Vitamins and tryptophan. Plant Foods Hum Nutr 35, 175–180. 145. Gwirtz JA & Garcia-Casal MN (2014) Processing maize flour and corn meal food products. Ann N Y Acad Sci 1312,66–75. 146. Dunn ML, Jain V & Klein BP (2014) Stability of key micronutrients added to fortified maize flours and corn meal. Ann N Y Acad Sci 1312,15–25. 147. Henry C & Heppell N (2002) Nutritional losses and gains during processing: future problems and issues. Proc Nutr Soc 61, 145–148. 148. Tiozon RJN, Fernie AR & Sreenivasulu N (2021) Meeting human dietary vitamin requirements in the staple rice via strategies of biofortification and post-harvest fortification. Trends Food Sci Technol 109,65–82. 149. Kyritsi A, Tzia C & Karathanos VT (2011) Vitamin fortified rice grain using spraying and soaking methods. LWT –Food Sci Technol 44, 312– 320. 150. de Pee S (2014) Proposing nutrients and nutrient levels for rice fortification. Ann N Y Acad Sci 1324,55–66. 151. Bhar S, Bose T, Dutta A & Mande SS (2022) A perspective on the benefits of consumption of parboiled rice over brown rice for glycaemic control. Eur J Nutr 61, 615–624. 152. Schroeder HA (1971) Losses of vitamins and trace minerals resulting from processing and preservation of foods. Am J Clin Nutr 24, 562–573. 153. Aboul-Enein HY & Loutfy MA (1984) Pyridoxine hydrochloride. In Analytical profiles of drug substances,13, In K Florey (ed), Analytical profiles of drug substances,13. Orlando, USA: Academic Press, pp 447– 486. 154. Schnellbaecher A, Binder D, Bellmaine S & Zimmer A (2019) Vitamins in cell culture media: stability and stabilization strategies. Biotechnol Bioeng 116, 1537–1555. 155. Yessaad M, Bernard L, Bourdeaux D, Chennell P & Sautou V (2018) Development of a stability indicating method for simultaneous analysis of five water-soluble vitamins by liquid chromatography. Pharm Technol Hosp Pharm 3, 207–218. 156. Juhász M, Takahashi S, Kitahara Y & Fujii T (2012) Thermal decomposition of pyridoxine: an evolved gas analysis-ion attachment mass spectrometry study. Rap Comm Mas Spect 26, 759–764. 157. Ferguson TI, Emery S, Price-Davies R & Cosslett AG (2014) A review of stability issues associated with vitamins in parenteral nutrition. e-SPEN J 9, e49–e53. 158. Bonrath WPK, Zhang QM, Pauling H & Weimann BJ (2020) Vitamins, 10. Vitamin B6. In G Bellussi, M Bohnet, J Bus, K Drauz, H Greim, KP Jackel, U Karst, A Kleemann, G Kreysa, T Laird, W Meier, E Ottow, M Roper, J Scholtz, K Sundmacher, R Ulber, U Wietelmann (eds), Ullmann’s encyclopedia of industrial chemistry, 7th ed. Weinheim, Germany: WileyVCH Verlag GmbH & Co. KGaA, pp 1–16. 159. Gregory III J & Kirk J (1978) Assessment of storage effects on vitamin B6 stability and bioavailability in dehydrated food systems. J Food Sci 43, 1801–1808. 22 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press 160. Gregory III J & Leatham K (1990) Lack of vitamin B6 activity of 6hydroxypyridoxine. J Food Sci 55, 1143–1146. 161. Shibata K, Yasuhara Y & Yasuda K (2001) Effects of cooking methods on the retention of vitamin B6 in foods, and the approximate cooking loss in daily meals. J Home Econ Jpn 52, 1187–1197. 162. McCormick DB (1978) Nature and activity of vitamin B6 bound in heatprocessed foods. Nutr Rev 36, 346–348. 163. Lešková E, Kubíková J, Kováčiková E, Košická M, Porubská J & Holčíková K (2006) Vitamin losses: retention during heat treatment and continual changes expressed by mathematical models. J Food Compost Anal 19, 252–276. 164. FuliaşA, Vlase G, Vlase T, Oneţiu D, Doca N & Ledeţi I (2014) Thermal degradation of B-group vitamins: B 1 ,B 2 and B 6 .J Therm Anal Calorim 118, 1033–1038. 165. Riaz MN, Asif M & Ali R (2009) Stability of vitamins during extrusion. Crit Rev Food Sci Nutr 49, 361–368. 166. Ribeiro DO, Pinto DC, Lima LMT, Volpato NM, Cabral LM & de Sousa VP (2011) Chemical stability study of vitamins thiamine, riboflavin, pyridoxine and ascorbic acid in parenteral nutrition for neonatal use. Nutr J10,1–9. 167. Mašková E, Rysova J, Fiedlerova V & Holasova M (1994) Vitamin and mineral retention in meat in various cooking methods. Potravinarske Vedy-UZPI (Czech Republic) 12, 407–416. 168. Bognár A (1993) Studies on the influence of cooking on the vitamin B6 content of food. In U Schlemmer (ed) Bioavailability ’93: nutritional, chemical and food processing implications of nutrient availability Proceedings, Part 1/2. Karlsruhe: Bundesforschungsanstalt für Ernährung S, pp 346–351. 169. Bognár A (1998) Comparative study of frying to other cooking techniques influence on the nutritive value. Grasas Aceites 49, 250–260. 170. Bognár A (2002) Tables on weight yield of food and retention factors of food constituents for the calculation of nutrient composition of cooked foods (dishes). Karlsruhe, Germany: Bundesforschungsanstalt für Ernährung. 171. Çatak J & Çaman R (2020) Pyridoxal pyridoxamine, and pyridoxine cooking loss: characterizing vitamin B6 profiles of chicken meats before and after cooking. J Food Process Preserv 44, e14798. 172. USDA (2007) USDA Table of Nutrient Retention Factors, Release 6: Beltsville, USA. https://www.ars.usda.gov/arsuserfiles/80400530/pdf/re tn06.pdf. 173. Engler PP & Bowers JA (1976) B-vitamin retention in meat during storage and preparation. A review. J Am Diet Assoc 69, 253–257. 174. Meyer BH, Mysinger M & Wodarski L (1969) Pantothenic acid and vitamin B6 in beef. J Am Diet Assoc 54, 122–125. 175. Olds S, Vanderslice J & Brochetti D (1993) Vitamin B6 in raw and fried chicken by HPLC. J Food Sci 58, 505–507. 176. Pearson A, Burnside J, Edwards H, Glasscock R, Cunha T & Novak A (1951) Vitamin losses in drip obtained upon defrosting frozen meat. Food Res 16,85–87. 177. Pearson A, West R & Luecke R (1959) The vitamin and amino acid content of drip obtained upon defrosting frozen pork. Food Res 24, 515–519. 178. Çatak J, Çaman R & Ceylan Z (2020) Critical vitamin assessment: pyridoxal, pyridoxamine, and pyridoxine levels for three species of raw and cooked fish samples. J Aquat Food Prod Technol 29, 981–989. 179. Muehlhoff E, Bennett A & McMahon D (2013) Milk and dairy products in human nutrition. Rome, Italy: FAO, pp 43–64. 180. Ryley J & Kajda P (1994) Vitamins in thermal processing. Food Chem 49, 119–129. 181. Asadullah K-u-N, Tarar OM, Ali SA, Jamil K & Begum A (2010) Study to evaluate the impact of heat treatment on water soluble vitamins in milk. J Pak Med Assoc 60, 909–912. 182. Rolls BA & Porter JWG (1973) Some effects of processing and storage on the nutritive value of milk and milk products. Proc Nutr Soc 32, 9–15. 183. Oamen E, Hansen A & Swartzel K (1989) Effect of ultra-high temperature steam injection processing and aseptic storage on labile water-soluble vitamins in milk. J Dairy Sci 72, 614–619. 184. Amador-Espejo G, Gallardo-Chacon J, Nykänen H, Juan B & Trujillo A (2015) Effect of ultra high-pressure homogenization on hydro-and liposoluble milk vitamins. Food Res Int 77,49–54. 185. Roe M, Church S, Pinchen H & Finglas P (2013) Nutrient analysis of eggs: Analytical report. Norwich: Institute of Food Research. 186. Öhrvik V, Carlsen MH, Källman A & Martinsen TA (2015) Improving food composition data by standardizing calculation methods. Copenhagen: Nordisk Ministerråd. 187. Alajaji SA & El-Adawy TA (2006) Nutritional composition of chickpea (Cicer arietinum L.) as affected by microwave cooking and other traditional cooking methods. J Food Compos Anal 19, 806–812. 188. Roe M, Church S, Pinchen H & Finglas P (2013) Nutrient analysis of fruit and vegetables; analytical report. Norwich: Institute of Food Research. 189. Rezaei M, Alizadeh Sani M, Amini M, et al. (2022) Influence of cooking process on the content of water-soluble B vitamins in rice marketed in Iran. Food Sci Nutr 10, 460–469. 190. Batifoulier F, Verny M-A, Chanliaud E, Rémésy C & Demigné C (2022) Effect of different breadmaking methods on thiamine, riboflavin and pyridoxine contents of wheat bread. J Cer Sci 42, 101–108. 191. Nurit E, Lyan B, Pujos-Guillot E, Branlard G & Piquet A (2016) Change in B and E vitamin and lutein, β-sitosterol contents in industrial milling fractions and during toasted bread production. J Cer Sci 69, 290–296. 192. Mihhalevski A, Nisamedtinov I, Hälvin K, Ošeka A & Paalme T (2013) Stability of B-complex vitamins and dietary fiber during rye sourdough bread production. J Cer Sci 57,30–38. 193. Athar N, Hardacre A, Taylor G, Clark S, Harding R & McLaughlin J (2006) Vitamin retention in extruded food products. J Food Compost Anal 19, 379–383. 194. EkıncıR (2005) The effect of fermentation and drying on the watersoluble vitamin content of tarhana, a traditional Turkish cereal food. Food Chem 90, 127–132. 195. Bull´o M, Juanola-Falgarona M, Hernández-Alonso P & Salas-Salvad´oJ (2015) Nutrition attributes and health effects of pistachio nuts. Br J Nutr 113, S79–S93. 196. Stuetz W, Schlörmann W & Glei M (2017) B-vitamins, carotenoids and α-/γ-tocopherol in raw and roasted nuts. Food Chem 221, 222–227. 197. Romanini E, Rastelli S, Donadini G, Lambri M & Bertuzzi T (2021) Pyridoxine and folates during small and large scale brewing. J Inst Brew 127, 135–139. 198. Rossi F, Spigno G, Luzzani G, et al. (2021) Effects of the intake of craft or industrial beer on serum homocysteine. Int J Food Sci Nutr 72,93–98. 199. Bertuzzi T, Mulazzi A, Rastelli S, Donadini G, Rossi F & Spigno G (2020) Targeted healthy compounds in small and large-scale brewed beers. Food Chem 310, 125935. 200. Zhang H, De Silva D, Dissanayaka D, Warkentin TD & Vandenberg A (2021) Validated B vitamin quantification from lentils by selected reaction monitoring mass spectrometry. Food Chem 359, 129810. 201. Moongngarm A & Saetung N (2010) Comparison of chemical compositions and bioactive compounds of germinated rough rice and brown rice. Food Chem 122, 782–788. 202. Khalil A & Mansour E (1995) The effect of cooking, autoclaving and germination on the nutritional quality of faba beans. Food Chem 54, 177–182. 203. Žili´c S, Basi´c Z, Hadži-Taškovi´c Šukalovi´c V, Maksimovi´c V, Jankovi´cM & Filipovi´c M (2014) Can the sprouting process applied to wheat improve the contents of vitamins and phenolic compounds and antioxidant capacity of the flour? Int J Food Sci Technol 49, 1040–1047. 204. Pinheiro SS, Anunciação PC, de Morais Cardoso L, et al. (2021) Stability of B vitamins, vitamin E, xanthophylls and flavonoids during germination and maceration of sorghum (Sorghum bicolor L.). Food Chem 345, 128775. Biological properties of vitamins of the B-complex 23 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press 205. Martin-Belloso O & Llanos-Barriobero E (2001) Proximate composition, minerals and vitamins in selected canned vegetables. Eur Food Res Technol 212, 182–187. 206. Kilcast D (1994) Effect of irradiation on vitamins. Food Chem 49, 157–164. 207. Woodside J (2015) Nutritional aspects of irradiated food. Stewart Postharvest Rev 11,1–6. 208. BernaśE & Jaworska G. (2016) Vitamins profile as an indicator of the quality of frozen Agaricus bisporus mushrooms. J Food Compos Anal 49, 1–8. 209. Zwart S, Kloeris V, Perchonok M, Braby L & Smith S (2009) Assessment of nutrient stability in foods from the space food system after long-duration spaceflight on the ISS. J Food Sci 74, H209–H217. 210. Coad R & Bui L (2020) Stability of vitamins B1, B2, B6 and E in a fortified military freeze-dried meal during extended storage. Foods 9, 39. 211. Williams PG (1996) Vitamin retention in cook/chill and cook/hot-hold hospital foodservices. J Am Diet Assoc 96, 490–498. 212. Fischesser J, Fritsch H, Gum AG, Karge R, Keuper R (2009) Manufacture of vitamin B 6 . Patent No. US 7.495,101 B2. 213. Bachmann T & Rychlik M (2018) Synthesis of [13C3]-B6 vitamers labelled at three consecutive positions starting from [13C3]-propionic acid. Molecules 23, 2117. 214. European Commission (2006) Commission Directive 2006/141/EC of 22 December 2006 on infant formulae and follow-on formulae and amending Directive 1999/21/EC. Off J Eur Union 49,1–33. 215. EFSA (2021) Assessment of a feed additive consisting of vitamin B6 (pyridoxine hydrochloride) for all animal species for the renewal of its authorisation (Kaesler Nutrition GmbH). EFSA J 19, e06612. 216. Bailey RL, Gahche JJ, Lentino CV, et al. (2011) Dietary supplement use in the United States, 2003–2006. J Nutr 141, 261–266. 217. Berendsen AA, van Lieshout LE, van den Heuvel EG, Matthys C, Péter S & de Groot LC (2016) Conventional foods, followed by dietary supplements and fortified foods, are the key sources of vitamin D, vitamin B6, and selenium intake in Dutch participants of the NU-AGE study. Nutr Res 36, 1171–1181. 218. Bird JK, Barron R, Pigat S & Bruins MJ (2022) Contribution of base diet, voluntary fortified foods and supplements to micronutrient intakes in the UK. J Nutr Sci 11, e51. 219. Gum AG (2006) Manufacture of vitamin B 6 . Patent No. WO 2006/066806 A1. 220. Eggersdorfer M, Laudert D, Letinois U, et al. (2012) One hundred years of vitamins—a success story of the natural sciences. Angew Chem Int Ed 51, 12960–12990. 221. Bonrath W & Netscher T (2005) Catalytic processes in vitamins synthesis and production. Appl Catal A: Gen 280,55–73. 222. Zou Y, Shi X, Zhang G, Li Z, Jin C & Su W (2013) Improved “oxazole” method for the practical and efficient preparation of pyridoxine hydrochloride (vitamin B6). Org Process Res Dev 17, 1498–1502. 223. Shtyrlin YG, Petukhov A, Strelnik A, et al. (2019) Chemistry of pyridoxine in drug design. Russ Chem Bull 68, 911–945. 224. Dumond YR & Gum AG (2003) Silane reduction of 5-hydroxy-6-methylpyridine-3, 4-dicarboxylic acid diethyl ester: synthesis of vitamin B6. Molecules 8, 873–881. 225. Ledesma-Amaro RS, Jiménez A & Revuelta JL (2013) Microbial production of vitamins. In McNeil B, Archer D, Giavasis I & Harvey L (eds), Microbial production of food ingredients, enzymes and nutraceuticals. Sawston: Woodhead Publishing, pp 571–594. 226. Acevedo-Rocha CG, Gronenberg LS, Mack M, Commichau FM & Genee HJ (2019) Microbial cell factories for the sustainable manufacturing of B vitamins. Curr Opin Biotechnol 56,18–29. 227. Wang Y, Liu L, Jin Z & Zhang D (2021) Microbial cell factories for green production of vitamins. Front Bioeng Biotechnol 9, 661562. 228. Commichau FM, Alzinger A, Sande R, et al. (2014) Overexpression of a non-native deoxyxylulose-dependent vitamin B6 pathway in Bacillus subtilis for the production of pyridoxine. Metab Eng 25,38–49. 229. Commichau FM, Alzinger A, Sande R, et al. (2015) Engineering Bacillus subtilis for the conversion of the antimetabolite 4-hydroxy-L-threonine to pyridoxine. Metab Eng 29, 196–207. 230. Rosenberg J, Ischebeck T & Commichau FM (2017) Vitamin B6 metabolism in microbes and approaches for fermentative production. Biotechnol Adv 35,31–40. 231. Rosenberg J, Yeak KC & Commichau FM (2018) A two-step evolutionary process establishes a non-native vitamin B6 pathway in Bacillus subtilis. Environ Microbiol 20, 156–168. 232. Yamamura E-T (2019) Bioconversion of pyridoxine to pyridoxamine through pyridoxal using a Rhodococcus expression system. J Biosci Bioeng 127,79–84. 233. Hoshino T, Ichikawa K, Nagahashi Y & Tazoe M (2004) Microorganism and process for preparing vitamin B 6 . Patent No. WO 2004/029228 A1. 234. Ichikawa K, Hoshino T & Tazoe M (1997) Fermentative production of vitamin B 6 . Patent No. EP 0 765 938 A2. 235. Tazoe M, Ichikawa K & Hoshino T (1999) Production of vitamin B6 in Rhizobium. Biosci Biotechnol Biochem 63, 1378–1382. 236. Kim J, Kershner JP, Novikov Y, Shoemaker RK & Copley SD (2010) Three serendipitous pathways in E. coli can bypass a block in pyridoxal-50phosphate synthesis. Mol Syst Biol 6, 436. 237. Richts B & Commichau FM (2021) Underground metabolism facilitates the evolution of novel pathways for vitamin B6 biosynthesis. Appl Microbiol Biotechnol 105, 2297–2305. 238. Cardoso RV, Fernandes Â, Gonzaléz-Paramás AM, Barros L & Ferreira IC (2019) Flour fortification for nutritional and health improvement: a review. Food Res Int 125, 108576. 239. Malik KA & Maqbool A (2020) Transgenic crops for biofortification. Front Sustain Food Syst 4, 571402. 240. Allen L, De Benoist B, Dary O & Hurrell R (2006) Guidelines on food fortification with micronutrients: WHO/FAO, Geneva, Switzerland: WHO Press. 241. Bird JK, Murphy RA, Ciappio ED & McBurney MI (2017) Risk of deficiency in multiple concurrent micronutrients in children and adults in the United States. Nutrients 9, 655. 242. Das JK, Salam RA, Mahmood SB, et al. (2019) Food fortification with multiple micronutrients: impact on health outcomes in general population. Cochrane Database Syst Rev,2019, CD011400. https:// www.cochranelibrary.com/cdsr/doi/10.1002/14651858.CD011400.pub2/ epdf/full . 243. Tucker KL, Olson B, Bakun P, Dallal GE, Selhub J & Rosenberg IH (2004) Breakfast cereal fortified with folic acid, vitamin B-6, and vitamin B-12 increases vitamin concentrations and reduces homocysteine concentrations: a randomized trial. Am J Clin Nutr 79, 805–811. 244. Duvenage SS & Schönfeldt HC (2007) Impact of South African fortification legislation on product formulation for low-income households. J Food Compos Anal 20, 688–695. 245. Tang Y, Xu M & Shiu-Ming K (2018) Factors contributing to the high prevalence of vitamin B6 deficiency in US: a systematic review. J Hum Nutr 2,58–64. 246. Ho C-L, Quay TA, Devlin AM & Lamers Y (2016) Prevalence and predictors of low vitamin B6 status in healthy young adult women in metro Vancouver. Nutrients 8, 538. 247. Newman JC, Malek AM, Hunt KJ & Marriott BP (2019) Nutrients in the US diet: naturally occurring or enriched/fortified food and beverage sources, plus dietary supplements: NHANES 2009–2012. J Nutr 149, 1404–1412. 248. Kim Y-N & Cho Y-O (2014) Evaluation of vitamin B6 intake and status of 20to 64-year-old Koreans. Nutr Res Pract 8, 688–694. 249. Pfeiffer CM, Sternberg MR, Schleicher RL, Haynes BMH, Rybak ME & Pirkle JL (2013) The CDC’s second national report on biochemical indicators of diet and nutrition in the u.s. population is a valuable tool for researchers and policy makers. J Nutr 143, 938S–947S. 250. Brown MJ, Ameer AM, Daley SF, et al. (2023) Vitamin B6 deficiency. StatPearls [Internet]. Treasure Island, FL: StatPearls Publishing. https:// www.ncbi.nlm.nih.gov/books/NBK470579/. 251. Morris MS, Picciano MF, Jacques PF & Selhub J (2008) Plasma pyridoxal 50-phosphate in the US population: the National Health and Nutrition Examination Survey, 2003–2004 12. Am J Clin Nutr 87, 1446–1454. 252. Bates CJ, Pentieva KD, Prentice A, Mansoor MA & Finch S (1999) Plasma pyridoxal phosphate and pyridoxic acid and their relationship to plasma 24 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press homocysteine in a representative sample of British men and women aged 65 years and over. Br J Nutr 81, 191–201. 253. Porter K, Hoey L, Hughes CF, Ward M & McNulty H (2016) Causes consequences and public health implications of low B-vitamin status in ageing. Nutrients 8, 725. 254. ter Borg S, Verlaan S, Hemsworth J, et al. (2015) Micronutrient intakes and potential inadequacies of community-dwelling older adults: a systematic review. Br J Nutr 113, 1195–1206. 255. FFI. Food Fortification Initiative (2022) https://www.ffinetwork.org/cou ntry-profiles. 256. GFDx. Global Fortification Data Exchange (2022) https://fortificationda ta.org/map-number-of-nutrients/. 257. Swanepoel E, Havemann-Nel L, Rothman M, et al. (2019) Contribution of commercial infant products and fortified staple foods to nutrient intake at ages 6, 12, and 18 months in a cohort of children from a low socioeconomic community in South Africa. Matern Child Nutr 15, e12674. 258. EU Parliament (2006) Regulation (EC) No 1925/2006 of the European Parliament and of the Council of 20 December 2006 on the addition of vitamins and minerals and of certain other substances to foods. Off J Eur Union 50,26–38. 259. European Commission (2006) Commission directive 2006/125/EC on processed cereal-based foods and baby foods for infants and young children. Off J Eur Union L339/16. 260. FSSAI (Food Safety and Standards Authorityof India) (2018) Food Safety and Standards (Fortification of Foods) Regulations. In: The Gazette of India: Extraordinarry (PART 111, Section 4). New Delhi, India: Ministry of Health and Family Welfare (MoHFW), Government of India. 261. Tapola N, Karvonen H, Niskanen L & Sarkkinen E (2004) Mineral water fortified with folic acid, vitamins B6, B12, D and calcium improves folate status and decreases plasma homocysteine concentration in men and women. Eur J Clin Nutr 58, 376–385. 262. Hennessy A, Walsh E, Walton J & Flynn A (2011) The contribution of fortified foods to micronutrient intake in Irish adults aged 18–64 years. Proc Nutr Soc 70, E112. 263. Sharma S & Litonjua A (2014) Asthma, allergy, and responses to methyl donor supplements and nutrients. J Allergy Clin Immunol 133, 1246– 1254. 264. Czeizel AE & Merhala Z (1998) Bread fortification with folic acid, vitamin B12, and vitamin B6 in hungary. Lancet 352, 1225. 265. Hannon EM, Kiely M & Flynn A (2018) The impact of voluntary fortification of foods on micronutrient intakes in Irish adults. Br J Nutr 97, 1177–1186. 266. Bouis H (2018) Reducing mineral and vitamin deficiencies through biofortification: progress under HarvestPlus. Hidden hunger: strategies to improve nutrition quality. World Rev Nutr Diet 118, 112–122. 267. Bouis HE & Saltzman A (2017) Improving nutrition through biofortification: a review of evidence from HarvestPlus, 2003 through 2016. Glob Food Sec 12,49–58. 268. Strobbe S & Van Der Straeten D (2018) Toward eradication of B-vitamin deficiencies: considerations for crop biofortification. Front Plant Sci 9, 443. 269. Vanderschuren H, Boycheva S, Li K-T, Szydlowski N, Gruissem W & Fitzpatrick TB (2013) Strategies for vitamin B6 biofortification of plants: a dual role as a micronutrient and a stress protectant. Front Plant Sci 4, 143. 270. Fudge J, Mangel N, Gruissem W, Vanderschuren H & Fitzpatrick TB (2017) Rationalising vitamin B6 biofortification in crop plants. Curr Opin Biotechnol 44, 130–137. 271. Blancquaert D, De Steur H, Gellynck X & Van Der Straeten D (2017) Metabolic engineering of micronutrients in crop plants. Ann N Y Aca Sci 1390,59–73. 272. Shewry PR, Van Schaik F, Ravel C, et al. (2011) Genotype and environment effects on the contents of vitamins B1, B2, B3, and B6 in wheat grain. J Agric Food Chem 59, 10564–10571. 273. Mooney S, Chen L, Kühn C, Navarre R, Knowles NR & Hellmann H (2013) Genotype-specific changes in vitamin B6 content and the PDX family in potato. BioMed Res Int 2013, 389723. 274. Azam FMS, Lian T, Liang Q, Wang W, Zhang C & Jiang L (2022) Variation of vitamin B contents in maize inbred lines: potential genetic resources for biofortification. Front Nutr 9, 1029119. 275. Herrero S & Daub ME (2007) Genetic manipulation of vitamin B-6 biosynthesis in tobacco and fungi uncovers limitations to up-regulation of the pathway. Plant Sci 172, 609–620. 276. Raschke M, Boycheva S, Crèvecoeur M, et al. (2011) Enhanced levels of vitamin B6 increase aerial organ size and positively affect stress tolerance in Arabidopsis.Plant J 66, 414–432. 277. Mohsin S, Irfan M, Saeed A, Malik KA & Maqbool A (2022) Enhanced expression of PDX1 accumulates vitamin B6 in transgenic wheat seeds. J Cer Sci 107, 103502. 278. Mangel N, Fudge JB, Li KT, et al. (2019) Enhancement of vitamin B6 levels in rice expressing Arabidopsis vitamin B6 biosynthesis de novo genes. Plant J 99, 1047–1065. 279. Li K-T, Moulin M, Mangel N, et al. (2015) Increased bioavailable vitamin B6 in field-grown transgenic cassava for dietary sufficiency. Nat Biotechnol 33, 1029–1032. 280. Bagri DS, Upadhyaya DC, Kumar A & Upadhyaya CP (2018) Overexpression of PDX-II gene in potato (Solanum tuberosum L.) leads to the enhanced accumulation of vitamin B6 in tuber tissues and tolerance to abiotic stresses. Plant Sci 272, 267–275. 281. Coburn SP, Lewis DL, Fink WJ, Mahuren JD, Schaltenbrand WE & Costill DL (1988) Human vitamin B-6 pools estimated through muscle biopsies. Am J Clini Nutr 48, 291–294. 282. Hamm MW, Mehansho H & Henderson LM (1979) Transport and metabolism of pyridoxamine and pyridoxamine phosphate in the small intestine of the rat. J Nutr 109, 1552–1559. 283. Said HM (2004) Recent advances in carrier-mediated intestinal absorption of water-soluble vitamins. Annu Rev Physiol 66, 419–446. 284. Yamashiro T, Yasujima T, Said HM & Yuasa H (2020) pH-dependent pyridoxine transport by SLC19A2 and SLC19A3: implications for absorption in acidic microclimates. J Biol Chem 295, 16998–17008. 285. Surtees R, Mills P & Clayton P (2006) Inborn errors affecting vitamin B6 metabolism. Future Neurol 1, 615–620. 286. EFSA (2016) Panel on dietetic products, nutrition and allergies, NDA. Dietary reference values for vitamin B6. EFSA J 14, 4485. 287. Anderson BB, Fulford-Jones CE, Child JA, Beard ME & Bateman CJ (1971) Conversion of vitamin B6 compounds to active forms in the red blood cell. J Clin Invest 50, 1901–1909. 288. Bohney JP, Fonda ML & Feldhoff R (1992) Identification of Lys190 as the primary binding site for pyridoxal 50-phosphate in human serum albumin. FEBS Lett 298, 266–268. 289. Fonda ML, Trauss C & Guempel UM (1991) The binding of pyridoxal 50phosphate to human serum albumin. Arch Biochem Biophys 288,79–86. 290. Ink SL & Henderson L (1984) Effect of binding to hemoglobin and albumin on pyridoxal transport and metabolism. J Biol Chem 259, 5833– 5837. 291. Merrill Jr AH & Henderson JM (1990) Vitamin B6 metabolism by human liver. Ann N Y Acad Sci 585, 110–117. 292. Merrill Jr AH, Henderson JM, Wang E, McDonald BW & Millikan WJ (1984) Metabolism of vitamin B-6 by human liver. J Nutr 114, 1664–1674. 293. Di Salvo ML, Contestabile R & Safo MK (2011) Vitamin B6 salvage enzymes: mechanism, structure and regulation. Biochim Biophys Acta (BBA)-Proteins Proteomics 1814, 1597–1608. 294. Lumeng L & Li T (1975) Characterization of the pyridoxal 50-phosphate and pyridoxamine 50-phosphate hydrolase activity in rat liver. Identity with alkaline phosphatase. J Biol Chem 250, 8126–8131. 295. Surtees R, Mills P & Clayton P (2006) Inborn errors affecting vitamin B 6 metabolism. Future Neurol 1, 615–620. 296. Stanulovi´c M, Jeremi´c V, Leskovac V & Chaykin S (1976) New pathway of conversion of pyridoxal to 4-pyridoxic acid. Enzyme 21, 357–369. 297. Phillips RS (2015) Chemistry and diversity of pyridoxal-50-phosphate dependent enzymes. Biochim Biophys Acta (BBA)-Proteins Proteomics 1854, 1167–1174. 298. Brunton L, Chabner BA, Knollman B (2011) Goodman and Gilman’s the pharmacological basis of therapeutics, 12 ed. New York: McGraw-Hill. Biological properties of vitamins of the B-complex 25 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press 577. Balamurugan K, Ortiz A & Said HM (2003) Biotin uptake by human intestinal and liver epithelial cells: role of the SMVT system. Am J Physiol Gastrointest Liver Physiol 285, G73–G77. 578. McCormick DB & Wright LD (1970) Section E –The metabolism of biotin and analogues. In M Florkin & EH Stotz (eds), Comprehensive biochemistry,21, Amsterdam: Elsevier, pp 81–110. 579. Zempleni J, McCormick DB & Mock DM (1997) Identification of biotin sulfone, bisnorbiotin methyl ketone, and tetranorbiotin-l-sulfoxide in human urine. Am J Clin Nutr 65, 508–511. 580. Lee H-M, Wright LD & McCormick DB (1972) Metabolism of carbonyllabeled 14C-biotin in the rat. J Nutr 102, 1453–1463. 581. Subramanian VS, Marchant JS, Boulware MJ, Ma TY & Said HM (2009) Membrane targeting and intracellular trafficking of the human sodiumdependent multivitamin transporter in polarized epithelial cells. Am J Physiol Cell Physiol 296, C663–C671. 582. Subramanian VS, Subramanya SB & Said HM (2011) Chronic alcohol exposure negatively impacts the physiological and molecular parameters of the renal biotin reabsorption process. Am J Physiol Renal Physiol 300, F611–F617. 583. Hassan YI, Moriyama H, Olsen LJ, Bi X & Zempleni J (2009) Nand Cterminal domains in human holocarboxylase synthetase participate in substrate recognition. Mol Genet Metab 96, 183–188. 584. Crisp SE, Griffin JB, White BR, et al. (2004) Biotin supply affects rates of cell proliferation, biotinylation of carboxylases and histones, and expression of the gene encoding the sodium-dependent multivitamin transporter in JAr choriocarcinoma cells. Eur J Nutr 43,23–31. 585. Manthey KC, Griffin JB & Zempleni J (2002) Biotin supply affects expression of biotin transporters, biotinylation of carboxylases and metabolism of interleukin-2 in Jurkat cells. J Nutr 132, 887–892. 586. Wolf B, Adam MP, Pagon RA, et al. (eds) (2000) Biotinidase deficiency. Seattle, WA: University of Washington. 587. Dakshinamurti K, Chalifour L & Bhullar RP (1985) Requirement for biotin and the function of biotin in cells in culture. Ann N Y Acad Sci 447, 38–55. 588. Wood HG & Barden RE (1977) Biotin enzymes. Ann Rev Biochem 46, 385–413. 589. Moss J & Lane MD (1971) The biotin-dependent enzymes. Adv Enzymol Relat Areas Mol Biol 35, 321–442. 590. Mari˜no-Ramírez L, Kann MG, Shoemaker BA & Landsman D (2005) Histone structure and nucleosome stability. Exp Rev Prot 2, 719–729. 591. Smith EM, Hoi JT, Eissenberg JC, et al. (2007) Feeding Drosophila a biotin-deficient diet for multiple generations increases stress resistance and lifespan and alters gene expression and histone biotinylation patterns. J Nutr 137, 2006–2012. 592. Gralla M, Camporeale G & Zempleni J (2008)Holocarboxylase synthetase regulates expression of biotin transporters by chromatin remodeling events at the SMVT locus. J Nutr Biochem 19, 400–408. 593. Hymes J, Fleischhauer K & Wolf B (1995) Biotinylation of histones by human serum biotinidase: assessment of biotinyl-transferase activity in sera from normal individuals and children with biotinidase deficiency. Biochem Mol Med 56,76–83. 594. Hymes J & Wolf B (1999) Human biotinidase isn’t just for recycling biotin. J Nutr 129, 485s–489s. 595. Camporeale G, Giordano E, Rendina R, Zempleni J & Eissenberg JC (2006) Drosophila melanogaster holocarboxylase synthetase is a chromosomal protein required for normal histone biotinylation, gene transcription patterns, lifespan, and heat tolerance. J Nutr 136, 2735–2742. 596. Narang MA, Dumas R, Ayer LM & Gravel RA (2004) Reduced histone biotinylation in multiple carboxylase deficiency patients: a nuclear role for holocarboxylase synthetase. Hum Mol Genet 13,15–23. 597. Ballard TD, Wolff J, Griffin JB, Stanley JS, van Calcar S & Zempleni J (2002) Biotinidase catalyzes debiotinylation of histones. Eur J Nutr 41, 78–84. 598. Peters DM, Griffin JB, Stanley JS, Beck MM & Zempleni J (2002) Exposure to UV light causes increased biotinylation of histones in Jurkat cells. Am J Physiol Cell Physiol 283, C878–C884. 599. Zempleni J (2005) Uptake, localization, and noncarboxylase roles of biotin. Annu Rev Nutr 25, 175–196. 600. Romero-Navarro G, Cabrera-Valladares G, German MS, et al. (1999) Biotin regulation of pancreatic glucokinase and insulin in primary cultured rat islets and in biotin-deficient rats. Endocrinology 140, 4595– 4600. 601. Spence JT & Koudelka AP (1984) Effects of biotin upon the intracellular level of cGMP and the activity of glucokinase in cultured rat hepatocytes. J Biol Chem 259, 6393–6396. 602. Vesely DL (1982) Biotin enhances guanylate cyclase activity. Science 216, 1329–1330. 603. Sol´orzano-Vargas RS, Pacheco-Alvarez D & Le´on-Del-Río A (2002) Holocarboxylase synthetase is an obligate participant in biotin-mediated regulation of its own expression and of biotin-dependent carboxylases mRNA levels in human cells. Proc Natl Acad Sci 99, 5325–5330. 604. Rodríguez-Meléndez R, Cano S, Méndez ST & Velázquez A (2001) Biotin regulates the genetic expression of holocarboxylase synthetase and mitochondrial carboxylases in rats. J Nutr 131, 1909–1913. 605. Collins JC, Paietta E, Green R, Morell AG & Stockert RJ (1988) Biotindependent expression of the asialoglycoprotein receptor in HepG2. J Biol Chem 263, 11280–11283. 606. Scheerger SB & Zempleni J (2003) Expression of oncogenes depends on biotin in human small cell lung cancer cells NCI-H69. Int J Vitam Nutr Res 73, 461–467. 607. Wiedmann S, Rodriguez-Melendez R, Ortega-Cuellar D & Zempleni J (2004) Clusters of biotin-responsive genes in human peripheral blood mononuclear cells. J Nutr Biochem 15, 433–439. 608. Ghosal A, Lambrecht N, Subramanya SB, Kapadia R & Said HM (2013) Conditional knockout of the Slc5a6 gene in mouse intestine impairs biotin absorption. Am J Physiol Gastrointest Liver Physiol 304, G64–G71. 609. Agrawal S, Agrawal A & Said HM (2016) Biotin deficiency enhances the inflammatory response of human dendritic cells. Am J Physiol Cell Physiol 311, C386–C391. 610. Kiykim E, Kiykim A, Cansever MS, Aktuglu Zeybek CA (2015) Biotinidase deficiency mimicking primary immune deficiencies. BMJ Case Rep 2015, bcr2014209275. 611. Barthelemy H, Chouvet B & Cambazard F (1986) Skin and mucosal manifestations in vitamin deficiency. J Am Acad Dermatol 15, 1263–1274. 612. Tone M, Powell MJ, Tone Y, Thompson SA & Waldmann H (2000) IL-10 gene expression is controlled by the transcription factors Sp1 and Sp3. J Immunol 165, 286–291. 613. Stratton SL, Bogusiewicz A, Mock MM, Mock NI, Wells AM & Mock DM (2006) Lymphocyte propionyl-CoA carboxylase and its activation by biotin are sensitive indicators of marginal biotin deficiency in humans. Am J Clin Nutr 84, 384–388. 614. Stratton SL, Horvath TD, Bogusiewicz A, et al. (2010) Plasma concentration of 3-hydroxyisovaleryl carnitine is an early and sensitive indicator of marginal biotin deficiency in humans. Am J Clin Nutr 92, 1399–1405. 615. Stratton SL, Horvath TD, Bogusiewicz A, et al. (2011) Urinary excretion of 3-hydroxyisovaleryl carnitine is an early and sensitive indicator of marginal biotin deficiency in humans. J Nutr 141, 353–358. 616. Baugh CM, Malone JH & Butterworth Jr CE (1968) Human biotin deficiency. A case history of biotin deficiency induced by raw egg consumption in a cirrhotic patient. Am J Clin Nutr 21, 173–182. 617. Jones P, Patel K & Rakheja D (2020) Chapter 33 –disorder: Biotin: biotinidase deficiency and holocarboxylase synthetase deficiency. In P Jones, K Patel & D Rakheja (eds), A quick guide to metabolic disease testing interpretation, 2nd ed: London: Academic Press, pp 171–174. 618. Xiong Z, Zhang G, Luo X, Zhang N & Zheng J (2020) Case report of holocarboxylase synthetase deficiency (late-onset) in 2 Chinese patients. Medicine 99, e19964. 619. Ozand PT, Gascon GG, Al Essa M, et al. (1998) Biotin-responsive basal ganglia disease: a novel entity. Brain 121, 1267–1279. 32 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press 620. Alfadhel M, Almuntashri M, Jadah RH, et al. (2013) Biotin-responsive basal ganglia disease should be renamed biotin-thiamine-responsive basal ganglia disease: a retrospective review of the clinical, radiological and molecular findings of 18 new cases. Orphanet J Rare Dis 8, 83. 621. Mock DM, Mock NI, Stewart CW, LaBorde JB & Hansen DK (2003) Marginal biotin deficiency is teratogenic in ICR mice. J Nutr 133, 2519– 2525. 622. Czeizel AE & Dudás I (1992) Prevention of the first occurrence of neuraltube defects by periconceptional vitamin supplementation. N Engl J Med 327, 1832–1835. 623. Mock DM & Stadler DD (1997) Conflicting indicators of biotin status from a cross-sectional study of normal pregnancy. J Am College Nutr 16, 252–257. 624. Mock DM, Stadler DD, Stratton SL & Mock NI (1997) Biotin status assessed longitudinally in pregnant women. J Nutr 127, 710–716. 625. Sedel F, Papeix C, Bellanger A, et al. (2015) High doses of biotin in chronic progressive multiple sclerosis: a pilot study. Mult Scler Relat Disord 4, 159–169. 626. Tourbah A, Lebrun-Frenay C, Edan G, et al. (2016) MD1003 (high-dose biotin) for the treatment of progressive multiple sclerosis: a randomised, double-blind, placebo-controlled study. Mult Scler 22, 1719–1731. 627. Couloume L, Barbin L, Leray E, et al. (2020) High-dose biotin in progressive multiple sclerosis: a prospective study of 178 patients in routine clinical practice. Mult Scler 26, 1898–1906. 628. Patel DP, Swink SM & Castelo-Soccio L (2017) A review of the use of biotin for hair loss. Skin Appendage Disord 3, 166–169. 629. Salaemae W, Booker GW & Polyak SW (2016) The role of biotin in bacterial physiology and virulence: a novel antibiotic target for Mycobacterium tuberculosis.Microbiol Spect 4.https://pubmed.ncbi. nlm.nih.gov/27227307;https://journals.asm.org/doi/epub/10.1128/micro biolspec.vmbf-0008-2015. 630. Walter P (2001) Towards ensuring the safety of vitamins and minerals. Toxicol Lett 120,83–87. 631. National Health and Medical Research Council, Australian Government Department of Health and Ageing, New Zealand Ministry of Health (2006) Nutrient reference values for Australia and New Zealand including recommended dietary intakes. Canberra, Australia: National Health and Medical Research Council. 632. Maillart E, Mochel F, Acquaviva C, Maisonobe T & Stankoff B (2019) Severe trvansient myopathy in a patient with progressive multiple sclerosis and high-dose biotin. Neurology 92, 1060–1062. 633. Pastén-Hidalgo K, River´on-Negrete L, Sicilia-Argumedo G, et al. (2020) Dietary biotin supplementation impairs testis morphology and sperm quality. J Med Food 23, 535–544. 634. Sawamura H, Fukuwatari T & Shibata K (2007) Effects of excess biotin administration on the growth and urinary excretion of water-soluble vitamins in young rats. Biosci Biotechnol Biochem 71, 2977–2984. 635. Salian SR, Nayak G, Kumari S, et al. (2019) Supplementation of biotin to sperm preparation medium enhances fertilizing ability of spermatozoa and improves preimplantation embryo development. J Assist Reprod Genet 36, 255–266. 636. Mock DM (2017) Biotin: from nutrition to therapeutics. J Nutr 147, 1487–1492. 637. Piketty ML, Prie D, Sedel F, et al. (2017) High-dose biotin therapy leading to false biochemical endocrine profiles: validation of a simple method to overcome biotin interference. Clin Chem Lab Med 55, 817–825. 638. Trambas CM, Sikaris KA & Lu ZX (2016) More on biotin treatment mimicking Graves’disease. N Engl J Med 375, 1698. 639. Piketty ML, Polak M, Flechtner I, Gonzales-Brice˜no L & Souberbielle JC (2017) False biochemical diagnosis of hyperthyroidism in streptavidinbiotin-based immunoassays: the problem of biotin intake and related interferences. Clin Chem Lab Med 55, 780–788. 640. Barbesino G (2016) Misdiagnosis of Graves’disease with apparent severe hyperthyroidism in a patient taking biotin megadoses. Thyroid 26, 860–863. 641. Elston MS, Sehgal S, Du Toit S, Yarndley T & Conaglen JV (2016) Factitious Graves’disease due to biotin immunoassay interference-a case and review of the literature. J Clin Endocrinol Metab 101, 3251–3255. 642. Batista MC, Ferreira CES, Faulhaber ACL, Hidal JT, Lottenberg SA & Mangueira CLP (2017) Biotin interference in immunoassays mimicking subclinical Graves’disease and hyperestrogenism: a case series. Clin Chem Lab Med 55, e99–e103. 643. Holmøy T & Torkildsen Ø (2016) Can vitamin D reduce inflammation in relapsing-remitting multiple sclerosis? Exp Rev Neurother 16, 233–235. 644. Sundström P & Salzer J (2015) Vitamin D and multiple sclerosis—from epidemiology to prevention. Acta Neurol Scand 132,56–61. 645. Saenger AK, Jaffe AS, Body R, et al. (2019) Cardiac troponin and natriuretic peptide analytical interferences from hemolysis and biotin: educational aids from the IFCC Committee on Cardiac Biomarkers (IFCC C-CB). Clin Chem Lab Med 57, 633–640. 646. FDA. Biotin Interference with Troponin Lab Tests –Assays Subject to Biotin Interference. Avaialble at https://www.fda.gov/medical-devices/invitro-diagnostics/biotin-interference-troponin-lab-tests-assays-subjectbiotin-interference. 647. Herman DS, Kavsak PA & Greene DN (2017) Variability and error in cardiac troponin testing: an ACLPS critical review. Am J Clin Pathol 148, 281–295. 648. Bayart J-L, Favresse J, Stoefs A, et al. (2020) Biotin interferences: have we neglected the impact on serological markers? Clin Chim Acta 503, 107–112. 649. Mock D (2012) Biochemical, Physiological, and Molecular Aspects of Human Nutrition, 3rd ed. Amsterdam, Netherlands: Elsevier. 650.. Capozzi V, Russo P, Duenas MT, Lopez P & Spano G (2012) Lactic acid bacteria producing B-group vitamins: a great potential for functional cereals products. Appl Microbiol Biotechnol 96, 1383–1394. 651. Hübner F & Arendt EK (2013) Germination of cereal grains as a way to improve the nutritional value: a review. Crit Rev Food Sci Nutr 53, 853–861. 652. Shewry PR, Hawkesford MJ, Piironen V, et al. (2013) Natural variation in grain composition of wheat and related cereals. J Agric Food Chem 61, 8295–8303. 653. Adebo OA (2020) African sorghum-based fermented foods: past, current and future prospects. Nutrients 12, 1111. 654. Bonku R & Yu J (2020) Health aspects of peanuts as an outcome of its chemical composition. Food Sci Hum Wellness 9,21–30. 655. Nti CA, Plahar WA & Annan NT (2016) Development and quality characteristics of shelf-stable soy-agushie: a residual by-product of soymilk production. Food Sci Nutr 4, 315–321. 656. King JC, Blumberg J, Ingwersen L, Jenab M & Tucker KL (2008) Tree nuts and peanuts as components of a healthy diet. J Nutr 138, 1736S–1740S. 657. Arya SS, Salve AR & Chauhan S (2016) Peanuts as functional food: a review. J Food Sci Technol 53,31–41. 658. Sathe SK, Monaghan EK, Kshirsagar HH & Venkatachalam M (2008) Chemical composition of edible nut seeds and its implications in human health. In C Alasalvar & F Shahidi (eds), Tree nuts: Composition, phytochemicals, and health effects. Boca Raton, USA: CRC Press, pp 11–36. 659. Chen CY, Lapsley K & Blumberg J (2006) A nutrition and health perspective on almonds. J Sci Food Agric 86, 2245–2250. 660. Barreca D, Nabavi SM, Sureda A, et al. (2020) Almonds (Prunus dulcis Mill. DA webb): a source of nutrients and health-promoting compounds. Nutrients 12, 672. 661. Roncero JM, Álvarez-Ortí M, Pardo-Giménez A, Rabadán A & Pardo JE (2020) Review about non-lipid components and minor fat-soluble bioactive compounds of almond kernel. Foods 9, 1646. 662. Dreher ML & Davenport AJ (2013) Hass avocado composition and potential health effects. Crit Rev Food Sci Nutr 53, 738–750. 663. Lockyer S & Spiro A (2020) The role of bread in the UK diet: an update. Nutr Bull 45, 133–164. 664. O’Connor A (2012) An overview of the role of bread in the UK diet. Nutr Bull 37, 193–212. 665. Esteve M, Farré R, Frıgola A & Pilamunga C (2002) Contents of vitamins B1, B2, B6, and B12 in pork and meat products. Meat Sci 62,73–78. 666. Probst Y (2009) Nutrient compostion of chicken meat. Publication No. 08/ 210, Project No. PRJ-001502; Rural Industries Research and Development Corporation, Australia, p 28. https://agrifutures.com.au/wp-content/ uploads/publications/08-210.pdf. Biological properties of vitamins of the B-complex 33 https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press 667. ANG CY (1980) Vitamin B6, and niacin contents of broiler meat of different strains, sexes, and production regions. J Food Sci 45, 898– 900. 668. Roe M, Church S, Pinchen H & Finglas P (2013) Nutrient analysis of fish and fish products: analytical report. Norwich: Institute of Food Research. 669. Feeney MJ, Dwyer J, Hasler-Lewis CM, et al. (2014) Mushrooms and health summit proceedings. J Nutr 144, 1128S–1136S. 670. Gahruie HH, Eskandari MH, Mesbahi G & Hanifpour MA (2015) Scientific and technical aspects of yogurt fortification: a review. Food Sci Hum Wellness 4,1–8. 671. Hampel D, York ER & Allen LH (2012) Ultra-performance liquid chromatography tandem mass-spectrometry (UPLC-MS/MS) for the rapid, simultaneous analysis of thiamin, riboflavin, flavin adenine dinucleotide, nicotinamide and pyridoxal in human milk. J Chromatogr B Analyt Technol Biomed Life Sci 903,7–13. 672. JenčoJ,Krčmová LK, Sobotka L, Bláha V, Solich P & Švec F (2020) Development of novel liquid chromatography method for clinical monitoring of vitamin B(1) metabolites and B(6) status in the whole blood. Talanta 211, 120702. 673. Heydari R & Elyasi NS (2014) Ion-pair cloud-point extraction: a new method for the determination of water-soluble vitamins in plasma and urine. J Sep Sci 37, 2724–2731. 674. Goberna-Bravo MÁ, Albiol-Chiva J, Peris-Vicente J, Carda-Broch S & Esteve-Romero J (2021) Determination of isoniazid and pyridoxine in plasma sample of tuberculosis patients by micellar liquid chromatography. Microchem J 167, 106317. 675. Amini R & Asadpour-Zeynali K (2020) Cauliflower-like NiCo2O4−Zn/Al layered double hydroxide nanocomposite as an efficient electrochemical sensing platform for selective pyridoxine detection. Electroanalysis 32, 1160–1169. 676. Immundiagnostik. ID-Vit B6. Vitamin B6. Avaialble at http://www.immu ndiagnostik.com/fileadmin/pdf/ID-Vit_VitaminB6_KIF006.pdf. 677. Recipe. ClinRep HPLC Complet Kit.Vitamin B1, B2 and B6. Available at https://recipe.de/products/combikit-vitamin-b1-b2-b6-whole-blood/. 678. Recipe. Clinmass LC-MS/MSComplet kit. Vitamin B1, B2and B6. Available at https://recipe.de/products/vitamins-b1-b2-b6-in-whole-blood/. 679. LSBio. Biotin Elisa Kit. Vitamin B7. Available at https://www.lsbio.com/eli sakits/manualpdf/ls-f10160.pdf. 680. Ministry of Education S, Science and Technology, Japan. (2015) Standard Tables of Food Composition in Japan, 7th Revised ed. https://www.mext. go.jp/en/policy/science_technology/policy/title01/detail01/sdetail01/sde tail01/1385122.htm. 681. DGE (2018) Deutsche Gesellschaft für Ernährung 2018. Available at http://www.nutrisurvey.de/info/interaktives/search.htm. 682. Buenrostro JL & Kratzer F (1984) Use of plasma and egg yolk biotin of White Leghorn hens to assess biotin availability from feedstuffs. Poult Sci 63, 1563–1570. 683. Kwon J, Cheon W, Lee S-H, Choi Y & Kim Y (2020) Monitoring of biotin content in frequently consumed foods in Korea. J Korean Soci Food Cult 35, 278–284. 684. Woollard DC & Indyk HE (2013) Biotin analysis in dairy products. London: RSC Publishing. 685. Shetty SA, Young MF, Taneja S & Rangiah K (2020) Quantification of Bvitamins from different fresh milk samples using ultra-high performance liquid chromatography mass spectrometry/selected reaction monitoring methods. J Chromatogr A 1609, 460452. 686. Abu-Elheiga L, Almarza-Ortega DB, Baldini A & Wakil SJ (1997) Human acetyl-CoA carboxylase 2. Molecular cloning, characterization, chromosomal mapping, and evidence for two isoforms. J Biol Chem 272, 10669–10677. 687. McKean AL, Ke J, Song J, et al. (2000) Molecular characterization of the non-biotin-containing subunit of 3-methylcrotonyl-CoA carboxylase. J Biol Chem 275, 5582–5590. 688. Wallace JC, Jitrapakdee S & Chapman-Smith A (1998) Pyruvate carboxylase. Int J Biochem Cell Biol 30,1–5. 689. Browner MF, Taroni F, Sztul E & Rosenberg LE (1989) Sequence analysis, biogenesis, and mitochondrial import of the alpha-subunit of rat liver propionyl-CoA carboxylase. J Biol Chem 264, 12680–12685. 34 P. Dias et al. https://doi.org/10.1017/S0954422425100097 Published online by Cambridge University Press