Probiotic fermented almond milk as an alternative to cow-milk yoghurt

In e na ional Jou nal o Food S udies IJFS Oc obe 2015 Volume 4 pages 201–211 P obio ic e men ed almond “milk” as an al e na i e o cow-milk yoghu Neus Be na a*, Mai e Ch´ a e aa, Ampa o Chi al a, and Chelo Gonz´ alez-Ma ´ ıneza aIns i u o de Ingenie ´ıa de Alimen os pa a el Desa ollo, Uni e si a Poli `ecnica de Val`encia, Camino de Ve a s/n, 46022, Valencia, Spain. *Co esponding au ho neube pe@up ne .up .es Tel: +34 96 387 70 00 Recei ed: 17 July 2014; Published online: 18 Oc obe 2015 In i ed pape om he 3 d In e na ional ISEKI Food Con e ence - ISEKI Food 2014 - B idging T aining and Resea ch o Indus y and he Wide Communi y - Food Science and Technology Excellence o a Sus ainable Bioeconomy Abs ac P obio ics in almond-based ma ices we e conside ed as a means o ob aining e men ed p oduc s which would co e bo h he cu en demand o heal h-p omo ing oods and o al e na i es o s anda d yoghu s. Fi s ly, he combined e ec o high p essu e homogenisa ion (HPH) and hea ea men on he physical s abili y o almond “milk” was s udied. The be e age was homogenised by applying 62, 103 and 172 MPa (MF1, MF2 and MF3 espec i ely); MF3 was also combined wi h wo di e en hea ea men s (85 ° C-30 min (LH) and 121 ° C-15 min (HH)). Bo h mic os uc u e and colloidal s abili y we e analysed in all he p ocessed samples o selec he mos sui able ea men wi h which o ob ain a s able p oduc . The selec ed almond milk was hen e men ed wi h p obio ic Lac obacillus eu e i and S ep ococcus he mophilus and he inal p oduc was cha ac e ised h oughou cold s o age ime (28 days) as o pH, acidi y, se um e en ion and s a e iabili y. A senso y e alua ion and p obio ic su i al o in i o diges ion was also conduc ed. The esul s showed ha he physical and s uc u al almond-milk p ope ies we e a ec ed by bo h HPH and hea ea men s, ob aining he g ea es s abili y in MF3-LH samples. The e men ed milk pe mi ed p obio ic su i als abo e he le el sugges ed as minimum o ensu ing heal h bene i s du ing he en i e con olled ime and, hence, can be conside ed as a unc ional ood. No di e ences in he senso y accep abili y o he p oduc we e ound be ween 1 and 28 s o age days. The e o e, a new, unc ional, e men ed p oduc was de eloped, which was sui able o a ge ed g oups, such as he lac ose-in ole an and cow-milk-p o ein alle gic popula ions. Keywo ds: Almond “milk”; Con ocal; Physical s abili y; P obio ic; Su i als; Fe men a ion 1 In oduc ion Vege able “milks” a e cu en ly a he o e on due o ei he he g owing p oblems ela ed wi h in ole ance (Fiocchi e al., 2010) o o changes in ood p e e ences. Vege able milks a e emulsi- ied p oduc s ( he lipid con en is dispe sed in an aqueous phase), which means hey a e he mo- dynamically uns able and he selec ion o p ope p ocessing condi ions plays a key ole in i s i- nal s abilisa ion. Indeed, di e en p ocess s eps, such as homogenisa ion and hea ea men s, usually p oduce changes in he a angemen o componen s, hus leading o modi ica ions in he physical s abili y o he p oduc . Copy igh © 2015 ISEKI-Food Associa ion (IFA) 10.7455/ij s/4.2.2015.a8 202 Be na e al. The mos commonly-used homogenisa ion p es- su es in he ood indus y ange be ween 20 and 50 MPa. The use o hese p essu es does no always ensu e he physical s abili y o he emulsions and he e o mula ion o he p od- uc is usually needed, by means o he addi ion o di e en emulsi ie s and/o s abilise s. The use o eme ging echnologies, such as high p es- su e homogenisa ion (HPH), may enhance he physical s abili y o he p oduc , since i p o- okes he de loccula ion o clus e s o p ima y a globules and he uni o m dispe sion o agglom- e a es, changes he p o ein con o ma ion and inc eases he emulsion’s iscosi y, among o he hings (Flou y, Des umaux, & La di`e es, 2000; Pe eda, Fe agu , Que edo, Guamis, & T ujillo, 2009; Des umaux & Ma cand, 2002). Despi e he ad an ages, cu en esea ch, based on he ap- plica ion o HPH in he p ocessing o ege able milks, p ocessing, has only ocused on soy milk (C uz e al., 2007; Li, Chen, Liu, & Chen, 2008). The e is a wide a ie y o ege able milks a ail- able on he ma ke bu he same canno be said o hei yoghu -like de i a i es. I p o- bio ic bac e ia a e used as s a e s, hese e - men ed p oduc s would mee he cu en demand o heal h-p omo ing ege able-based p oduc s. P obio ics a e li e mic oo ganisms ha , when adminis e ed in adequa e amoun s in edible ma- ices (i.e. ood p oduc s), con e heal h bene- i s on he hos (FAO/WHO, 2001), such as he educ ion o hype choles e olemia, he hos im- mune modula ion, he alle ia ion o cons ipa ion, he p o ec ion agains a elle ’s dia hoea, he p o ec ion agains colon and bladde cance , he p e en ion o os eopo osis o he modula ion o ood alle gies (Saad, Dela e, U daci, Schmi e , & B essollie , 2013). In pa icula , he so-called almond “milk” has long been used as an al e na i e o cow-milk o lac o-in ole an people, p egnan women and celiacs mainly due o hei high le els o calcium, phospho ous and po assium (Luengo, 2009). The e o e, i could be used as a base p oduc o de elop new non-dai y e men ed p oduc s wi h unc ional ea u es, in which he nu i ional and heal h bene i s o almonds and p obio ic bac e ia a e included. Almond nu s a e ich in mono- and polyunsa u a ed a y acids (mainly oleic and linoleic acids), ege able p o eins, die a y ib e, phy os e ols, polyphenols, i amins and mine als (Yada, Lapsley, & Huang, 2011); mos o which compounds ha e an ioxidan p ope ies and a p o en bene icial e ec on he plasma lipid p o- ile, low-densi y lipop o ein oxida ion and in lam- ma o y p ocesses, among o he hings (Liu, 2012; Ege , K a z, Kannenbe g, Fobke , & Wah bu g, 2011; Jones e al., 2011). The aim o his s udy was, on he one hand, o analyse he combined e ec o hea ea men s and high homogenisa ion p essu es on he phys- ical p ope ies and s abili y o almond milk in o de o de ine p ocessing condi ions which en- su e he p oduc quali y and s abili y and, on he o he hand, o de elop a non-dai y, e men ed, p obio ic p oduc wi h high quali y p ope ies by using L. eu e i ATCC 55730, a well-es ablished p obio ic (Casas & Molls am, 1997), mixed wi h S. he mophilus CECT 986. 2 Ma e ials and Me hods 2.1 P epa a ion o almond milk The be e age was p oduced by soaking and g inding almonds (P unus amygdalus L. a . dulcis) supplied by F u os Secos 3G S.L. (Va- lencia, Spain). The ex ac ion was ca ied ou in Sojama ic 1.5 (Sojama ic ® ; Ba celona, Spain), equipmen speci ically designed o he p oduc- ion o ege able be e ages, wi h a nu -wa e a- io o 8:100. The manu ac u ing p ocess akes 30 minu es a oom empe a u e. The milky liq- uid ob ained was used as a con ol sample (un- ea ed). 2.2 High p essu e homogenisa ion and hea ea men s High p essu e homogenisa ion (HPH) ea men s we e ca ied ou in a high p essu e homogenise (M-110P model; Mic o luidics In e na ional Co - po a ion, USA) by applying 62, 103 and 172 MPa (samples MF1, MF2 and MF3 espec i ely). The homogenise used uns in con inuous mode and he e en ion ime is e y sho (2-4 seconds). Raw almond milk and MF3 samples we e sub- mi ed o a low empe a u e hea ea men a 85 IJFS Oc obe 2015 Volume 4 pages 201–211 P obio ic e men ed almond “milk” 203 ° C o 30 min (known as LH and MF3LH, espec- i ely) and o a high empe a u e hea ea men , 121 ° C o 15 min (HH and MF3HH). The hea ea men condi ions chosen we e hose in which he des uc ion o all ege a i e cells and en- zymes a e ensu ed (Wals a, Wals a, Wou e s, & Geu s, 2014). 2.3 Fe men ed almond milk p ocessing Inoculum p epa a ion Lac obacillus eu e i ATCC 55730 (Biogaia, S ockholm, Sweden) and S ep ococcus he - mophilus CECT 986 (CECT, Valencia, Spain) we e ac i a ed om hei ozen o ms (s o ed in 40g/100 mL glyce ol a - 80 ° C), by ans- e ing hem o hei selec i e b o hs un il op- imal bac e ial g ow h is ob ained. The se- lec i e b o hs we e MRS (Scha lab, Ba celona, Spain) o he p obio ic Lac obacillus and M17 (Bioka Diagnos ics, Beau ais, F ance) o S. he mophilus. Incuba ion condi ions we e 37 ° C/24h/anae obically o L. eu e i, in which anae obiosis was c ea ed by using anae obic ja s and a CO2-gene a o sys em (Ana e oGenTM, Oxoid L d, Basings oke, England) and 42 ° C/24h/ae obically o S. he mophilus. Fe men a ion p ocess Almond milks submi ed o he e men a ion p ocess we e en iched wi h 1.5 % (w/w) glucose- uc ose (Sosa Ing edien s SL., Ba celona, Spain) p io o he inocula ion p ocess in o de o im- p o e he g ow h and acidi ica ion o he mixed cul u e used, based on p e ious s udies (Be na , Ch´a e , Gonz´alez-Ma ´ınez, Rod ´ıguez-Ga c´ıa, & Chi al , 2014). S ains in he exponen ial phase we e cen i uged a 8,600 xg/10 min/4 ° C o elimina e he selec- i e b o hs. Immedia ely a e wa ds, each ype o bac e ia was esuspended in PBS-1x bu e (10 mmol/L phospha e, 137 mmol/L NaCl, 2.7 mmol/L KCl, pH 7.4) sepa a ely un il eaching concen a ions o 108 colony o ming uni s (c u) pe mL. 3 mL o each suspension pe 100 mL o p ocessed almond milks we e inocula ed. The inocula ed milk samples we e hen incuba ed a he op imal empe a u e o he mixed cul u e (40 ° C). When samples eached a pH o ≈4.6, hey we e cooled o 4 ° C (end o e men a ion p ocess) and s o ed a his empe a u e un il he analyses we e pe o med. 2.4 Cha ac e iza ion o s uc u al p ope ies and s abili y o almond milk Con ocal lase scanning mic oscopy (CLSM) The mic os uc u e o bo h aw and ea ed al- mond milks was analysed by means o CLSM, by using a Nikon con ocal mic oscope C1 uni i - ed on a Nikon Eclipse E800 mic oscope (Nikon, Tokyo, Japan). Fluo escen Rhodamine B dye (Fluka, Sigma-Ald ich, Missou i, USA) was used o s ain p o eins and ca bohyd a es, while Nile Red (Fluka, Sigma-Ald ich, Missou i, USA) was used o s ain lipids, ollowing he me hodology desc ibed by Be na e al. (2014). Colloidal s abili y o almond “milks” The colloidal s abili y o bo h aw and ea ed almond milks was de e mined ia he phase sep- a a ion analysis h oughou he s o age ime (28 days) a 4 ° C. To his end, abou 15 g o sam- ples we e pou ed in o glass ubes o 16 mm in diame e and he heigh o he sepa a e phases was quan i ied. 0.04 g/ 100mL o sodium azide (Pan eac Qu´ımica S.L.U., Ba celona, Spain) was added o he samples, o a oid any mic obial g ow h du ing s o age. 2.5 Cha ac e iza ion o e men ed almond-based p oduc s pH and i a able acidi y (TA) The pH o non- e men ed and e men ed al- mond samples was measu ed a 25 ° C using a pH-me e (GLP 21+, C ison Ins umen s S.A.; Spain). The AOAC s anda d me hod was cho- sen o de e mine he TA in samples (AOAC 947.05), which consis ed o a i a ion wi h 0.1 IJFS Oc obe 2015 Volume 4 pages 201–211 204 Be na e al. mol/L NaOH solu ion (Pan eac Qu´ımica S.L.U., Ba celona, Spain), exp essing he esul s as g ams o lac ic acid pe L (Ho wi z, 2000). Simula ed gas oin es inal diges ion (SGID) Fe men ed almond milk was submi ed o a SGID. This in i o diges ion, done in iplica e, was pe o med as Glahn, Lee, Yeung, Goldman, and Mille (1998) desc ibed, bu no demine aliza- ion was ca ied ou . Po cine pepsin (800-2500 uni s/mg p o ein), panc ea in (ac i i y, 4 1 USP speci ica ions) and bile ex ac we e pu chased om Sigma-Ald ich ® (S . Louis, MO, USA). Viabili y o s a e s The su i al o bo h L. eu e i and S. he - mophilus in e men ed almond milks was quan- i ied ollowing he me hod desc ibed by he In- e na ional Dai y Fede a ion (In e na ional IDF S anda d, 1997). The selec i e media used we e MRS aga (Scha lab; Ba celona, Spain) o L. eu e i and M17 aga (Bioka Diagnos ics, Beau- ais, F ance) o S. he mophilus. The incuba- ion condi ions we e 37 ° C /48 h/ae obically o S. he mophilus and 37 ° C/24 h/anae obically o L. eu e i. Anae obiosis was c ea ed by us- ing anae obic ja s and a CO2-gene a o sys em (Ana e oGenTM; Oxoid L d, Basings oke, Eng- land) and coun s we e epo ed as log c u/mL. These analyses we e done in e men ed samples kep in chilled s o age a di e en imes and also, in he e men ed samples submi ed o a SGID. Se um e en ion capaci y (SR) The SR o bo h non- e men ed and e men ed almond milks was analysed by sample cen i u- ga ion (Medi ige -BL, JP-Selec a; Spain). The condi ions we e 2,500 xg/45 min/20 ° C and he amoun o se um sepa a ion was used o quan i y he sample s abili y. Senso y analysis A 16 membe -s ong ained panel e alua ed almond-based e men ed p oduc s a e di e en s o age imes (1, 14, and 28 days) a 4 ° C. The membe s we e selec ed on he basis o hei in e - es , a ailabili y, lack o ood alle gies and hei h eshold o basic la ou s. They we e ained o sco e a ibu es o swee ness, acidi y, almond la ou , consis ency and mou h eel and o e all accep abili y using in e al scales ha a ied om 1 (sligh ly) o 5 (ex emely). The e e ence samples we e used o se he in- e al scales o panel aining. Fo he acidi y e e ence, 1 and 2% o suc ose was added o com- me cial milk yoghu , co esponding o 3 and 1 on he scale, espec i ely, and wi h 0.2% o ci - ic acid co esponding o 5. Comme cial milk yoghu wi h 2, 5 and 14% o added suc ose le - els was used o he swee ness e alua ion, co e- sponding o 1, 3 and 5 on he scale, espec i ely. Fo consis ency and mou h eel, liquid yoghu , comme cial soy desse and Danone o iginal ® yoghu we e used as e e ences, co esponding o 1, 3 and 5 on he scale, espec i ely. Fo he al- mond la ou , he e e ence was he almond milk used in he s udy, which co esponded o 5 on he scale. Fo he senso y analysis, each panellis es ed 3 samples (cold s o ed o 0, 14 and 28 days, e- spec i ely) con aining 6% o suc ose, in o de o quan i y he a ibu es in which each one was ained. The samples we e andomly p esen ed wi h a h ee digi code. The e alua ion was con- duc ed in a no malised as ing oom a oom empe a u e. 3 Resul s and Discussion 3.1 Op imiza ion o almond “milk” p ocessing Figu e 1shows bo h he mic os uc u e and ini- ial mac os uc u e o almond milk, bo h un- ea ed and submi ed o di e en ea men s (HPH and/o hea ea men s). The oil d ople s and p o ein bodies dispe sed in he se um phase a e clea ly dis inguished in he mic os uc u e o un ea ed milk (Fig. 1A). A ce ain deg ee o p o ein loccula ion was obse ed due o hei hy- d ophobic cha ac e , since mos almond p o eins belong o he oleosin amily, wi h low-molecula weigh and poo wa e solubili y (Beisson, Fe e, Voul ou y, & A ondel, 2001). This low wa e - IJFS Oc obe 2015 Volume 4 pages 201–211 P obio ic e men ed almond “milk” 205 a ini y o p o eins con ibu es o he poo s a- bili y o he ob ained emulsions. In LH samples (Fig. 1B), he majo i y o he al- mond p o eins we e agg ega ed. In many cases, p o ein agg ega es included oil d ople s which in- duce he o ma ion o a weak gel, as was obse ed in he mac os uc u e. The MF3 ea men g ea ly educed he size o he a globules (Fig. 1C). Ne e heless, mos o he small pa icles we e loccula ed h ough p o- ein b idges, which explains he poo s abili y o he emulsion despi e he ac ha pa icles a e small in size. The poo s abilizing p ope ies o he p o ein, associa ed wi h i s high deg ee o hyd ophobici y, is he cause o he loccula ion p ocess and subsequen phase sepa a ion, as was obse ed in he mic og aphs. The MF3LH ea men caused he o ma ion o big oil d ople -p o ein agg ega es which appea embedded in a con inuous p o ein ma ix, hus enhancing he physical s abili y o he milk (Fig. 1D). This new s uc u e is he esul o he com- bined e ec o bo h ea men s. MF educes d ople size and p omo es pa ial p o ein solu- bilisa ion (C uz e al., 2007; Zhang, Li, Ta sumi, & Isobe, 2005) and he hea ea men s p o oke p o ein dena u a ion and agg ega ion, gi ing ise o he o ma ion o a 3D ne wo k which en aps big agg ega es o he small p o ein-lipid pa icles (Wals a e al., 2014). Wi h ega ds o he mac os uc u e, all he sam- ples showed phase sepa a ion a e 1 s o age day, excep hose submi ed o MF3 and MF3LH ea men s (Fig. 1). A e 2 s o age days, MF3 samples also exhibi ed phase sepa a ion. Once he sepa a ion p ocess occu ed in he di e en samples, no no able di e ences in he heigh o he sepa a e phases we e obse ed h oughou s o age pe iod (da a no shown). Ne e heless, samples submi ed o MF3LH ea men we e s able h oughou he ime s udied. The e o e, he combined e ec o homogenisa ion and he - mal ea men seems o p omo e a weak gel o - ma ion ( ela ed o he o ma ion o clus e s), mainly associa ed wi h he p o ein solubilisa ion and subsequen dena u a ion du ing he he mal ea men , which con ibu ed o he s abilisa ion o he pa icle dispe sion, hus a oiding phase sepa a ion du ing he p oduc s o age. These clus e o ma ions ha e also been obse ed in hea ed and homogenised cow milk (Wals a e al., 2014). The obse ed beha iou indica es ha , in al- mond samples submi ed o HPH, he emulsi y- ing p ope ies o he p o eins we e no sui able o s abilising a globules by in e acial p o ein adso p ion, as obse ed in mic og aphs. Only when homogenised samples we e submi ed o he mal ea men and he p o eins we e dena- u ed, did hese help o s abilise he emulsions, mainly due o a iscous e ec . 3.2 Cha ac e iza ion s. s o age ime o e men ed p ocessed almond “milk” P obio ic su i al As Budding on (2009) e iewed, heal h bene- i s ha p obio ics may exe a e di ec ly linked wi h, on he one hand, hei abili y o su i e in he medium in which hey a e p esen and, on he o he hand, hei abili y o su i e in he gas- oin es inal ac . Since a p obio ic educ ion is expec ed bo h in ood ma ices and once hey a e diges ed, a minimum numbe o iable p obio ic bac e ia o 107 c u/mL has been s ongly ec- ommended by he ime o consump ion in o de o e ec i ely p o ide such heal h unc ionali ies (Sanz & Dalmau, 2008). Figu e 2shows he iabili y o bo h L. eu e i and S. he mophilus in almond-based ma ices a e di e en cold s o age imes, in which s a- is ical di e ences be ween s o age imes we e also included by ep esen ing LSD in e als (95% con idence le el). As shown in Figu e 2, he ia- bili y o bo h s ains dec eased h oughou s o - age ime (p<0.05), especially in he case o S. he mophilus. This las s ain was used o ech- nological pu poses, since i is epo ed o im- p o e he g ow h o lac obacilli (i.e. L. eu e i) (Tamime & Robinson, 1999); hence i s iabil- i y was no as ele an as he p obio ic s ain. L. eu e i coun s we e abo e he minimum le el ecommended (107c u/mL) h oughou s o age ime, which ensu ed he po en ial p obio ic unc- ionali ies. The e o e, he e men ed p oduc de- eloped, owing o he p esence o he p obio ic L. eu e i, could be used o he educ ion o in- IJFS Oc obe 2015 Volume 4 pages 201–211 206 Be na e al. Figu e 1: Mic os uc u e (CLSM mic og aphs) and mac os uc u e (Canon images) o almond “milk” un ea ed (A), low hea ea ed (LH) (B), homogenised a 172 MPa (MF) (C) and ea ed by combined MF-LH (D). : a an ile colic and imp o emen s eeding ole ance (Sa ino, Pelle, Palume i, Ogge o, & Minie o, 2007; Ind io e al., 2008), he educ ion o cons i- pa ion (Cocco ullo e al., 2010) and modula ion o ce ain cy okines in ol ed in a opic diseases (Miniello e al., 2010). As ega ds he in i o diges ion assays ca ied ou , he iabili y o he p obio ic bac e ia a e SGID was ema kably high, as mo e han 50 % o he L. eu e i p esen in he e men ed almond milk was able o su i e agains gas o-in es inal enzymes (p<0.05), which ein o ces he unc ion- ali y o he p oduc . An ioxidan Capaci y Table 1shows he pH, i a able acidi y (TA) and se um e en ion capaci ies (SR) a e sam- ple cen i uga ion o bo h non- e men ed and e - men ed almond milks. As can be seen, he pH alues we e kep cons an h oughou he s o - age ime, which a e aged 4.65. Al hough he TA inc eased o e s o age ime, om he 7 h day onwa ds he di e ences be ween he alues we e non-signi ican (p<0.05), being a ound 2.2 g/L. This low TA alue migh ha e a posi i e e - ec on he o e all senso y accep ance o he inal p oduc , since i is a c i ical ea u e in yoghu s (Tamime & Robinson, 1999). As a as he SR alues (Table 1) a e conce ned, a g ea e se um sepa a ion was quan i ied in non- e men ed samples, while e y ew di e ences we e obse ed in he case o e men ed samples s o ed o di e en leng hs o ime. This new, weak gel s uc u e was o med in he e men ed p oduc as a esul o he loccula ion o dis- pe sed pa icles caused by o he ac ion o p o- eins and hese esul s sugges ha i was able o be e e ain pa o he se um p esen in he almond milk. IJFS Oc obe 2015 Volume 4 pages 201–211 P obio ic e men ed almond “milk” 207 Figu e 2: Viabili y o bo h L. eu e i (g ey line) and S. he mophilus (dashed black line) in e men ed almond milk a e di e en cold s o age imes (0, 1, 7, 14, 21 and 28 days). S a is ical di e ences be ween s o age imes we e also included by ep esen ing LSD in e als (95% con idence le el) Table 1: Values o pH, i a able acidi y (TA) and se um e en ion capaci y (SR) o bo h p ocessed almond “milk” (MF3LH) and e men ed de i ed p oduc (FP) h oughou s o age ime (days) a 4 ° C Sample pH TA (g/L o lac ic acid) SR (% ( / ) o p ecipi a e) MF3LH 6.567 (0.006) 0.39 (0.03) 36 (2) FP 1 d 4.657 (0.012) a1.90 (0.12) a43 (2) abc FP 7 d 4.63 (0.02) b2.23 (0.09) b42 (3) bc FP 14 d 4.657 (0.006) a2.23 (0.0) b39 (0.7) c FP 21 d 4.633 (0.012) b2.19 (0.07) b45 (3) ab FP 28 d 4.650 (0.019) ab 2.26 (1.0) b48 (3) a a,b,c Di e en le e s in same column indica es signi ican di e ences be ween samples analysed a 95% con idence le els IJFS Oc obe 2015 Volume 4 pages 201–211 208 Be na e al. Figu e 3: Panellis s’ sco es o swee ness, acidi y, consis ency and o e all accep abili y o he e men ed almond samples s o ed o 1 and 28 days a 4 ° C.a,b Di e en le e s in same a ibu e indica es signi ican di e ences be ween s o age imes (p<0.05) Senso y analysis Figu e 3shows he sco es o he a ibu es o appea ance, swee ness, acidi y, consis ency and o e all accep abili y in he e men ed almond samples es ed by he membe s o he panel (a e 1 and 28 s o age days a 4 ° C). S a is ical di e - ences be ween s o age imes we e also included, showing he homogenous g oups acco ding o a LSD es (95% con idence le el). Be o e as ing he e men ed p oduc s, he panel- lis s conside ed hei appea ance and consis ency o be good, hese no being a ibu es a ec ed by he s o age ime (p<0.05). Once he samples we e as ed, he panellis s ap- p ecia ed he samples s o ed o 1 day as be- ing signi ican ly mo e acidic and less swee han hose s o ed o 28 days; his is in spi e o he ac ha he longe samples a e s o ed, he mo e acidic hey gene ally become. This con o e sial esul could be explained by conside ing he syn- hesis o he ola ile ace ic acid b ough abou by he L. eu e i s ain (A skold e al., 2008), which is seen o ans e de ec able inega y, pungen and acidic odou s in o e men ed p od- uc s (Cheng, 2010). This acid is mainly o med a he beginning o e men a ion and, a e 28 s o age days, he nega i e e ec o ace ic acid migh ha e disappea ed due o i s ola ilisa ion. The accep abili y o he p oduc was only a ed as ai , and hus, some s a egies could be de- signed o imp o e i . The judges ound no sig- ni ican di e ences in he o e all accep abili y o samples as a ec ed by s o age ime (p<0.05). Fu he s udies a e needed o modi y in some ex en he mou h eel and/o la ou o he e - men ed p oduc , in o de o ensu e he de eloped IJFS Oc obe 2015 Volume 4 pages 201–211 P obio ic e men ed almond “milk” 209 p oduc enjoys a wide consume accep ance. 4 Conclusions Raw almond milk is an uns able dispe sion, which could be physically s abilised by he com- bina ion o low hea ea men wi h high ho- mogenisa ion p essu es. Almond milk is a p ope ma ix wi h which o ob ain e men ed de i a i e p oduc s, whe e s a e bac e ia a e able o g ow and su i e. The e men a ion p ocess modi ied he inne s uc u e o almond milk, con e ing i o a weak gel ha was able o e ain he wa e con en o he almond milk. Mo eo e , he use o p obio ic L. eu e i combined wi h S. he - mophilus as s a e inoculum pe mi ed a high p obio ic su i al a e, bo h h oughou he yp- ical shel li e o yoghu -like p oduc s (28 days) and a e an in i o diges ion p ocess, enhanc- ing he possible heal h bene i s ha he de el- oped p oduc may impa o consume s due o he p esence o L. eu e i. This new, non-dai y e men ed unc ional ood can be consumed by a ge ed g oups such as ege a ians, he lac ose- in ole an and hose alle gic o cow-milk p o ein. Ne e heless, despi e he senso y esul s, some modi ica ions in mou h eel and/o la ou should be s udied in o de o imp o e i s senso y accep - abili y and ensu e ha i enjoys a wide ma ke accep ance. Acknowledgemen s This esea ch has been ca ied ou hanks o a p ojec unded by he Uni e si a Poli `ecnica de Val`encia (PAID-05-11-2740). This wo k was also suppo ed by he “Conselle ia d’Educaci´o” o Va- lencian go e nmen , which awa ded a g an o he au ho N. Be na (ACIF/2011). 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