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Enhancing bio emedia ion esea ch wi h mixed-e ec s models: A s a is ical app oach
o enzyme kine ics analysis
Chike An hony Nweze 1, *, Alisa Ch is ophe Onyemezi i 1, Nwanneamaka Ri a Oze 1, Alex Ali Bila 1, S anley
Chinonso Ukane o 1 and Kel in Izuchukwu Me enini 2
1 Depa men o Chemis y, Fede al Uni e si y o Technology, PMB 1526, Owe i, Imo S a e, Nige ia.
2 Depa men o Compu e Science, Fede al Uni e si y o Technology, PMB 1526, Owe i, Imo S a e, Nige ia.
Wo ld Jou nal o Ad anced Resea ch and Re iews, 2025, 26(02), 410-415
Publica ion his o y: Recei ed on 21 Ma ch 2025; e ised on 27 Ap il 2025; accep ed on 30 Ap il 2025
A icle DOI: h ps://doi.o g/10.30574/wja .2025.26.2.1466
Abs ac
Bio emedia ion o pe oleum-con amina ed soils elies hea ily on enzyma ic ac i i ies as p oxies o mic obial unc ion
and soil heal h. This s udy e alua es he e ec i eness o a ious o ganic and ino ganic amendmen s—namely municipal
was e, calcium oxide, Aspilia a icana, and Eupa o ium odo a a—in enhancing enzyma ic ac i i ies in used engine oil-
con amina ed soils. By applying mixed-e ec s models and enzyme kine ics analysis, we in es iga e he in luence o
ea men s and subs a e concen a ion on phospha ase, u ease, dehyd ogenase, and ca alase ac i i ies. Ou indings
highligh municipal was e as he mos e ec i e ea men , consis en ly yielding he highes enzyma ic eloci ies and
ca aly ic e iciencies o e 126 days. Mixed-e ec s models p o ided obus insigh in o ixed and andom e ec s,
cap u ing a iabili y ac oss ime and ea men s. This wo k demons a es he po en ial o in eg a ing s a is ical
modeling wi h biochemical assessmen s o op imize bio emedia ion s a egies.
Keywo ds: Mixed-E ec s Models; Enzyme Kine ics; Phospha ase; Dehyd ogenase; Ca alase; Soil Remedia ion
1. In oduc ion
Enzyme-media ed bio emedia ion has eme ged as a sus ainable and cos -e ec i e app oach o pollu an deg ada ion
in con amina ed en i onmen s. The abili y o enzymes o ca alyze he b eakdown o o ganic and ino ganic pollu an s
has been well-documen ed in ecen s udies, wi h applica ions in soil, wa e , and was ewa e ea men (Singh e al.,
2019; Sha ma & Reddy, 2021; Pa k e al., 2023). Despi e hese ad an ages, bio emedia ion e iciency is in luenced by
mul iple ac o s, including enzyme ype, subs a e concen a ion, and en i onmen al condi ions such as empe a u e
and pH (Chen e al., 2020; Kuma & Pa el, 2022).
One o he p ima y challenges in enzyma ic bio emedia ion is he inhe en a iabili y in enzyme ac i i y due o
luc ua ions in hese ac o s, making i di icul o es ablish p edic i e models o emedia ion e iciency (Gup a e al.,
2018; Osei e al., 2023). T adi ional s a is ical me hods, such as linea eg ession, o en ail o cap u e his complexi y,
as hey assume homogenei y in expe imen al condi ions and do no accoun o andom a ia ions ac oss di e en
expe imen al se ups (Jones & Taylo , 2019; Bane jee e al., 2021). To o e come his limi a ion, mixed-e ec s models
ha e been p oposed as a obus s a is ical ool ha can inco po a e bo h ixed e ec s (e.g., enzyme ype and subs a e
concen a ion) and andom e ec s (e.g., expe imen al a iabili y and ba ch di e ences) (Zhang e al., 2021; Nwankwo
e al., 2023).
By in eg a ing mixed-e ec s modeling in o enzyme kine ics analysis, esea che s can imp o e he accu acy o
bio emedia ion p edic ions and op imize enzyme-subs a e in e ac ions o enhanced deg ada ion e iciency (Wilson
Wo ld Jou nal o Ad anced Resea ch and Re iews, 2025, 26(02), 410-415
411
e al., 2018; Lee e al., 2023). P e ious s udies ha e demons a ed ha mixed-e ec s models ou pe o m adi ional
eg ession app oaches in biological and en i onmen al esea ch by educing unexplained a iabili y and accoun ing o
hie a chical da a s uc u es (Ga cia & Pa el, 2020; Yamamo o e al., 2022). This s udy aims o apply mixed-e ec s
models o enzyme kine ics da a, quan i y he e ec s o ime, enzyme ype, and subs a e concen a ion on emedia ion
ou comes, and ul ima ely enhance he s a is ical obus ness o bio emedia ion esea ch.
2. Ma e ials and Me hods
2.1. Expe imen al Design
Soil samples we e amended wi h municipal was e, calcium oxide, Aspilia a icana, Eupa o ium odo a a, and un ea ed
con ol. The soil was amended wi h Municipal was e, Calcium oxide, Aspilla A icana, and Eupa o ium Oda a a a e
a i icial pollu ion. The soil samples we e collec ed using a soil auge om he op and lowe soil o analysis a e
homogeniza ion. Enzyme assays we e conduc ed a mul iple ime poin s (Day 0 o Day 126), measu ing he ac i i y o
phospha ase, u ease, dehyd ogenase, and ca alase.
2.2. Enzyme Kine ics and S a is ical Modeling
Enzyme ac i i ies we e quan i ied using Michaelis-Men en kine ics and e alua ed ia Linewea e -Bu k plo s. Mixed-
e ec s linea eg ession models we e applied o assess ixed e ec s ( ea men , subs a e concen a ion, ime) and
andom e ec s (sampling a iabili y). Key pa ame e s included Maximum Veloci y (Vmax), Michaelis-Men en cons an
(Km), and Ca aly ic e iciency (Vmax/Km).
3. Resul s
3.1. Mixed-E ec s Modeling o Enzyme Ac i i ies
3.1.1. Phospha ase
In Figu e 3.1 below, phospha ase ac i i y and ends o e ime ac oss ea men s we e illus a ed. Municipal was e and
calcium oxide exhibi ed consis en ly highe eac ion eloci ies han o he ea men s, while he con ol and Eupa o ium
odo a a showed minimal enzyma ic esponse.
Figu e 1 Linewea e -Bu k plo s e alua ing phospha ase enzyme kine ics a di e en emedia ion pe iods (Day 0 o
Day 126)
Wo ld Jou nal o Ad anced Resea ch and Re iews, 2025, 26(02), 410-415
412
S a is ical analysis showed a signi ican ea men e ec (F (4, 310) = 6.10, p < .001), bu no signi ican e ec o ime (F
(6, 308) = 0.28, p = .9479). Pos hoc compa isons con i med he supe io pe o mance o municipal was e and calcium
oxide.
3.1.2. U ease
Bo h ea men and ime we e signi ican p edic o s (p < .001). Municipal was e exhibi ed he highes u ease ac i i y,
peaking a Day 14. A signi ican in e ac ion be ween ea men and subs a e concen a ion sugges ed subs a e
inhibi ion a high le els.
Figu e 2 Linewea e -Bu k plo s e alua ing u ease enzyme kine ics a di e en emedia ion pe iods (Day 0 o Day
126)
Bo h ea men and ime e ec s we e signi ican : (F(4, 310) = 33.14, p < .001) and (F(6, 308) = 48.52, p < .001),
espec i ely.
3.1.3. Dehyd ogenase
Dehyd ogenase ac i i y co ela ed posi i ely wi h subs a e concen a ion and showed s ong ea men and ime
e ec s. Municipal was e consis en ly inc eased Vmax and educed Km o e ime.
Wo ld Jou nal o Ad anced Resea ch and Re iews, 2025, 26(02), 410-415
413
Figu e 3 Linewea e -Bu k plo s e alua ing dehyd ogenase enzyme kine ics a di e en emedia ion pe iods (Day 0 o
Day 126).
Signi ican ea men (F (4, 310) = 32.58, p < .001) and ime (F (6, 308) = 48.42, p < .001) e ec s we e obse ed.
3.1.4. Ca alase
Ca alase ac i i y ollowed a simila end, wi h municipal was e yielding he highes Vmax/Km, and calcium oxide
showed mode a e imp o emen s.
Wo ld Jou nal o Ad anced Resea ch and Re iews, 2025, 26(02), 410-415
414
Figu e 4 Linewea e -Bu k plo s e alua ing ca alase enzyme kine ics a di e en emedia ion pe iods (Day 0 o Day
126)
Ca alase ac i i y was in luenced by ea men (F (4, 310) = 20.22, p < .001) and ime (F (6, 308) = 83.82, p < .001).
4. Discussion
The in eg a ion o mixed-e ec s models enabled nuanced in e p e a ion o enzyma ic esponses unde di e en
bio emedia ion ea men s. Municipal was e eme ged as he mos e ec i e amendmen , suppo ing mic obial g ow h
and enzyme syn hesis, as shown by highe Vmax and lowe Km alues o e ime. The in e ac ion e ec s u he e ealed
how enzyme pe o mance can pla eau o decline wi h excessi e subs a e, an insigh c ucial o ield applica ion.
T adi ional models o en o e look hese complexi ies, whe eas mixed-e ec s modeling accoun s o empo al ends,
epea ed measu es, and he e ogenei y in soil condi ions. The abili y o isola e andom e ec s o e ed deepe insigh s
in o unexplained a iabili y, suppo ing mo e accu a e conclusions.
5. Conclusion
This s udy unde sco es he impo ance o s a is ical modeling in en i onmen al biochemis y. Mixed-e ec s models
p o ide a powe ul amewo k o e alua ing enzyme kine ics, o e ing cla i y in in e p e ing complex in e ac ions in
soil bio emedia ion. The esul s ad oca e o he use o municipal was e as a obus amendmen and encou age u he
explo a ion o enzyme-subs a e dynamics using ad anced s a is ical ools.
Compliance wi h e hical s anda ds
Acknowledgmen s
The au ho s wish o hank M . S anley C. Ukane o o his cons uc i e e ision o he manusc ip .
Disclosu e o con lic o in e es
No con lic o in e es o be disclosed.
Wo ld Jou nal o Ad anced Resea ch and Re iews, 2025, 26(02), 410-415
415
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