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PROCESSING CONTAMINANTSIN EDIBLEOILS

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PROCESSING CONTAMINANTSIN EDIBLEOILS

MCPDandGlycidylEsters

SECONDEDITION

Editedby SHAUNMACMAHON

U.S.FoodandDrugAdministration, CenterforFoodSafetyandAppliedNutrition(CFSAN), MD,UnitedStates

JESSICAK.BEEKMAN

U.S.FoodandDrugAdministration, CenterforFoodSafetyandAppliedNutrition(CFSAN), MD,UnitedStates

AcademicPressandAOCSPress

AcademicPressisanimprintofElsevier 125LondonWall,LondonEC2Y5AS,UnitedKingdom 525BStreet,Suite1650,SanDiego,CA92101,UnitedStates 50HampshireStreet,5thFloor,Cambridge,MA02139,UnitedStates TheBoulevard,LangfordLane,Kidlington,OxfordOX51GB,UnitedKingdom

PublishedincooperationwithAmericanOilChemists’ Society www.aocs.org Director, ContentDevelopment:JanetBrown

Nopartofthispublicationmaybereproducedortransmittedinanyformorbyanymeans, electronicormechanical,includingphotocopying,recording,oranyinformationstorage andretrievalsystem,withoutpermissioninwritingfromthepublisher.Detailsonhowto seekpermission,furtherinformationaboutthePublisher’spermissionspoliciesandourarrangementswithorganizationssuchastheCopyrightClearanceCenterandtheCopyright LicensingAgency,canbefoundatourwebsite: www.elsevier.com/permissions.

Thisbookandtheindividualcontributionscontainedinitareprotectedunder copyrightbythePublisher(otherthanasmaybenotedherein).

Notices

Knowledgeandbestpracticeinthis ﬁeldareconstantlychanging.Asnewresearch andexperiencebroadenourunderstanding,changesinresearchmethods,professional practices,ormedicaltreatmentmaybecomenecessary.

Practitionersandresearchersmustalwaysrelyontheirownexperienceandknowledgein evaluatingandusinganyinformation,methods,compounds,orexperimentsdescribed herein.Inusingsuchinformationormethodstheyshouldbemindfuloftheirownsafety andthesafetyofothers,includingpartiesforwhomtheyhaveaprofessionalresponsibility.

Tothefullestextentofthelaw,neitherthePublishernortheauthors,contributors,or editors,assumeanyliabilityforanyinjuryand/ordamagetopersonsorpropertyasa matterofproductsliability,negligenceorotherwise,orfromanyuseoroperationof anymethods,products,instructions,orideascontainedinthematerialherein.

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AcataloguerecordforthisbookisavailablefromtheBritishLibrary

ISBN:978-0-12-820067-4

ForinformationonallAcademicPressandAOCSPresspublications visitourwebsiteat https://www.elsevier.com/books-and-journals

Publisher: NikkiP.Levy

AcquisitionsEditor: NancyMaragioglio

EditorialProjectManager: SamW.Young

ProductionProjectManager: BharatwajVaratharajan

CoverDesigner: MarkRogers

1.Introduction1

ShaunMacMahon

Formation1

Mitigation1

Analysis2

Toxicology3

Regulations4

References5

2.Formationmechanisms7

BrianD.CraftandFrédéricDestaillats

Introduction7 MCPDesters8 Glycidylesters12

Conclusionsandperspectives20 References20

3.MitigationofMCPDandglycidylestersinedibleoils23

BertrandMatthäusandFrankPudel

Introduction23

Inﬂuenceofprecursorsonesterformation26

Mitigationin ﬁeld37

Mitigationoftherawmaterial39

Mitigationinreﬁning42

Additives52

Removalfromfullyreﬁnedoils(post-reﬁning)56 References59

4.Indirectdetectiontechniques65 KarelHrncirík

Introduction65

Mainstepsintheanalysisof2-and3-MCPDesters66 Mainapproachestowardtheanalysisofglycidylesters75

Ofﬁciallyadoptedmethods83 Methodcomparison89 Methodautomation96 Analysisofoil-basedfoodstuffs97 Challengesinfoodstuffanalysis98 Conclusions103 References104

5.Directanalyticaldetectionmethodsforthefood-borne toxicantsglycidylesters109 AliceEwertandMichaelGranvogl Introduction109 Directmethods111 References142

6.DirectdetectiontechniquesforMCPDesters145

ShaunMacMahon Background145 Directmethods146 TheInstituteofChemicalTechnology162 Conclusion191 References191

7.MethodstodetectMCPDandglycidylestersincomplexfood matrices195

ShaunMacMahon,JessicaBeekmanandMichaelGranvogl Background195 Discussion196 Conclusions217 References217

8.Toxicologicalpropertiesofglycidylesters221

GabrieleScholzandBenoîtSchilter Introduction221 Biologicalpropertiesofglycidylesters222 Biologicalpropertiesoffreeglycidol225 Conclusions231 References232

9.ToxicologicalpropertiesofMCPDfattyacidesters235

AlfonsoLampen

Abbreviations235

Introduction235

Toxicologyof3-MCPD236

Hazardpotentialof3-MCPDfattyacidesters242

Toxicologyof2-MCPDesters247

Riskcharacterization248 Conclusions249 References250

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Contributors

JessicaBeekman

FDACenterforFoodSafetyandAppliedNutrition,CollegePark,MD,UnitedStates

BrianD.Craft

NestléResearchCenter,FoodScienceandTechnologyDepartment,Lausanne, Switzerland

FrédéricDestaillats

NestléResearchCenter,FoodScienceandTechnologyDepartment,Lausanne, Switzerland

AliceEwert

GermanResearchCenterforFoodChemistry,Freising,Germany

MichaelGranvogl

UniversityofHohenheim,InstituteofFoodChemistry,DepartmentofFoodChemistry andAnalyticalChemistry(170a),Stuttgart,Germany;TechnicalUniversityofMunich, DepartmentofChemistry,ChairforFoodChemistry,Freising,Germany

KarelHrncirík

Upﬁeld,Rotterdam,theNetherlands

AlfonsoLampen

FederalInstituteforRiskAssessment(BfR),Berlin,Germany

ShaunMacMahon

FDACenterforFoodSafetyandAppliedNutrition,CollegePark,MD,UnitedStates

BertrandMatthäus

MaxRubner-Institut,DepartmentforSafetyandQualityofCereals,WorkingGroupfor LipidResearch,Detmold,Germany

FrankPudel

PilotPﬂanzenöltechnologieMagdeburge.V.,Magdeburg,Germany

BenoîtSchilter

SociétédesProduitsNestléS.A.,NestléResearch,Lausanne,Switzerland

GabrieleScholz

SociétédesProduitsNestléS.A.,NestléResearch,Lausanne,Switzerland

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CHAPTER1

Introduction

ShaunMacMahon FDACenterforFoodSafetyandAppliedNutrition,CollegePark,MD,UnitedStates

Formation

Toimproveconsumeracceptance,edibleoilsareindustriallyprocessedto removeormodifycomponentsthatcannegativelyimpactappearance,taste, andshelfstability.However,undesirablechemicalchangescantakeplace duringthereﬁningprocess.Fattyacidestersof3-chloro-1,2-propanediol (3-MCPD),2-chloro-1,3-propanediol(2-MCPD),andglycidolareheatinducedcontaminantsthatarenotpresentinvirginunreﬁnedoils,but theycanbeproducedbythehightemperaturesappliedduringdeodorization [1 3].Thereisevidencethat3-MCPDestersareformedfromironchloride and/ornaturalorganochlorinespresentinnativeoils[4 7].ThepredominantprecursorsandformationpathwaysforMCPDandglycidylesterswill bethoroughlyreviewedinChapter2ofthistext.

Mitigation

ThefactthatMCPDestersbeginformingat200 Cmakesmitigation difficult,asdeodorizationsaregenerallyrunattemperaturesgreaterthan 200 C[4].ManyfactorscontributetotheformationofMCPDandglycidyl esters.Thegrowingconditionsandharvestingofthepalmfruitcanhave profoundeffectsonanoil’scapacitytoformcontaminants.Theextraction, washing,andprocessingstepsthattakeplacepriortodeodorizationcan influencetheformationofthesetoxicantsduringdeodorization,ascanthe specificsofthedeodorizationscheme.ItisalsopossibletoremoveMCPD andglycidylestersusingappropriateadsorbentsorenzymes.Therehave beenanumberofrecentadvancesinmitigationofMCPDandglycidyl esters,includingindustrialscaleproductionofmitigatedpalmoilforusein infantformulaintheUnitedStates[8]. Chapter3 ofthistextdiscussesthe optimizationofallofthesestepstoreduceandeliminatethepresenceof thesecontaminantsinrefinededibleoils.

ProcessingContaminantsinEdibleOils

ISBN978-0-12-820067-4

https://doi.org/10.1016/B978-0-12-820067-4.00007-3

Analysis

Processededibleoilsarecommonlyconsumedworldwideandusedinthe productionofmanyfoodsincludinginfantformula,highlightingtheneed foraccurateanalyticalmethodologyfortheirdetection.Indirectapproaches requiringesterhydrolysisfollowedbyderivatizationandanalysisbyGC MSwerethe firstmethodsdevelopedtodetectMCPDandglycidylesters [9 11].Whiletheseearlymethodsplayedacrucialroleinbringing attentionfromindustryandregulatorstothepresenceofthesecontaminantsinrefinedoils,theearlyconditionsforbase-catalyzedhydrolyzeswere showntobepotentiallyunreliable,raisingquestionsaboutthetrustworthinessofindirectmethodology[12,13].Morerecently,however,these issueshavebeenaddressedandthequalityandreliabilityoftheseindirect methodshasimprovedgreatly.Severalindirectapproaches(includingthose involvingautomation)haverecentlycompletedsuccessfulcollaborative studies,reachingofficialcompendialmethodstatuswithorganizationssuch astheAmericanOilChemists’ Society(AOCS)andtheInternational OrganizationforStandardization(ISO).Theapplicationoftheseindirect methodstotheanalysisofMCPDandglycidylestersinoilsandhighfat matriceslikedressings/spreadswillbecoveredin Chapter4.

Partlyinresponsetothelackofdependabilityofearlyindirectmethodology,directmethodsweredevelopedforglycidylesters(GEs)and 3-MCPDesters,withcontaminantsanalyzedintactastheyoccurinprocessedoils.Althoughdirecttechniquesrequirealargenumberofstandards (someofwhicharenotcommerciallyavailable)forquantitativeanalysis,this methodologycanprovideinformationabouttheindividualMCPDand glycidylesterspresentinareﬁnedoil indirectmethods,ontheother hand,providequantitativeinformationregardingthetotalestercontentin anoilexpressedasboundMCPDandglycidol.Detailedinformationabout thestructureoftheindividualesterscanbeusefulinanumberofapplications.Somestudieshaveindicatedpotentialtoxicologicaldifferences betweendifferent3-MCPDesters;thisinformationislostwhenapplying indirectmethodology.Inaddition,asmethodshavebeguntobeextended tocomplexfoods,theuseofadirectapproachcanensureacceptable extractionrecoveriesofallthedifferentesters,whichdohavedistinct polaritydifferencesinsomecases.Informationaboutindividualesterscan alsobeusefulinevaluatingtheeffectivenessofmitigationapproaches.The useofdirectmethodsfortheanalysisofintactestersofglycidol(Chapter5) and3-MCPD(Chapter6)inedibleoilswillbediscussed.

Sincereﬁnedoilsaretheprimaryfatsourceincommercialinfant formulas,thepresenceofMCPDandglycidylestersisasourceofpotential concernduetoinfants’ lowbodyweightandconsumptionofinfant formulaasthesolesourceofnutritioninsomechildren.Asaresult,research effortshavefocusedonthedevelopmentofanalyticalmethodsforthe analysisofMCPDandglycidylestersininfantformula,aswellasinother complexfoodscommonlyconsumedbychildren. Chapter7 willreview theanalysisofMCPDestersandglycidylestersininfantformulaaswellas inothercomplexfoods.

Toxicology

Freeglycidol,3-MCPD,and2-MCPDallposeconcernsfromafoodsafety perspective.Glycidolisagenotoxiccarcinogenthatisprobablycarcinogenictohumans[14].AccordingtotheFederalInstituteforRiskAssessmentinBerlin,Germany(BfR),concentrationsshouldbekeptaslowasare reasonablyachievableinfood[15].Duetoitsgenotoxicity,theFoodSafety CommissionofJapanhassuggestedglycidolintakeshouldbebasedonthe ALARAprinciple(AsLowAsReasonablyAchievable)[16].Negative effectsonkidneysandreproductivesystemshavebeenseenfrom3-MCPD intoxicologicalstudies[17],anditwasclassiﬁedbytheEuropeanScientiﬁc CommitteeonFoodasanongenotoxicthresholdcarcinogen[18].Free 3-MCPDhasbeenlabeledasagroup2Bcarcinogen(possiblycarcinogenic tohumans)bytheInternationalAgencyforResearchonCancer(IARC) [19].Therearetoxicologicalconcernsshowninlimitedstudiesrelatedto 2-MCPD;oneunpublishedreportshowedthathighdosesaffectedstriated musclesandtheheart,aswellasthekidneysandtheliverinrats[20].

Relativetothefreeformsofthesecontaminants,researchonthefatty acidestersthatareformedindeodorizedoilshasonlybeeninitiatedmore recently[15,20,21].Invivotoxicologicalworkhasdemonstratedthatfree 3-MCPDisliberatedfromthediesterforminrats[22]asisglycidolfrom glycidylesters[23]inrelativelyhighyields.Initialriskassessments conductedbytheBfRhaveconcludedthatusingaworst-casescenario, infantswhoarefedonlycommercialinfantformulascouldpotentially ingestamountsofglycidoland3-MCPDexceedingtheJointFoodand AgricultureOrganization/WorldHeathOrganizationExpertCommittee onFoodAdditives(JECFA)recommendedmaximumtolerabledailyintake levels[21].Exposuresto3-MCPDestersintheUnitedStatesfrom consumptionofinfantformulas,estimatedusingdataon3-MCPDester

concentrationsinformulasamplescollectedbetween2013and2015,are higherthanthoseestimatedforEuropeaninfants.However,estimated glycidylesterexposureswerecomparableforUSandEuropeaninfantsin thattimerange[8].Thefullresultsofalltoxicologicalstudiesonthese contaminantswillbediscussedin Chapter8 (glycidylesters)and Chapter9 (MCPDesters).

Regulations

Inresponsetothedetectionoffree3-MCPDinhydrolyzedvegetable protein,soysauce,andbakedgoods,manyinternationalorganizations addressedtheissueinthosematrices.TheJECFArecommendeda maximumtolerabledailyintakefor3-MCPDof2 m[mu]g/kgbodyweight perday[24].TheEuropeanCommissionestablishedamaximumlevelof 20 m[mu]g/kg(ppb)for3-MCPDinhydrolyzedvegetableproteinandsoy sauce[25],whichwasalsoadoptedbyFoodStandardsAustraliaNew Zealand(FSANZ)[26].TheCodexAlimentariusadoptedamaximumlevel of400 m[mu]g/kg(ppb)inliquidcondimentscontainingacid-hydrolyzed vegetableprotein(excludingnaturallyfermentedsoysauce)in2008[27]. TheUSFoodandDrugAdministrationCompliancePolicyGuidestates thathydrolyzedvegetableproteinthatcontains3-MCPDatlevelsgreater than1 m[mu]g/g(ppm)isnotgenerallyrecognizedassafe(GRAS),and thereforeisanunsafefoodadditive[28].HealthCanadaalsosetamaximum contaminantconcentrationof1 m[mu]g/g(ppm)inAsian-stylesauces[29].

RegulationsregardingMCPDorglycidylesterconcentrationsin processedoilshavebeguntoappearinrecentyears.TheEuropean Commission(EC)publishedaninitialmaximumlevelforglycidylesters (expressedasglycidol)inpowderedinfantformulaof75 mg/kgthatwas reducedto50 mg/kginJuly2019;andamaximumlevelinliquidformula of10 mg/kgthatwasreducedto6 mg/kginJuly2019[30].Inaddition,the maximumlevelforboundglycidolinvegetableoilsandfatsis1000 mg/kg exceptforthoseusedtoproducebabyfoodandprocessedcereal-basedfood forinfantsandyoungchildren,wherethelimitis500 mg/kg[30].TheEC hasproposedlevelsforthesumoffree3-MCPDandbound3-MCPDfor severalproducts,althoughthesehavenotbeen finalized.Theproposed limitsforthesumofboundandfree3-MCPDare1250 mg/kgforoils/fats fromindividualoroilmixesfromcoconut,corn,canola,sunflower, soybean,andpalmkernel;2500 mg/kgforoils/fatsfromindividualormixes fromothervegetableoilsand fishoil,and2500 mg/kgformixturesofoils frombothcategories.Vegetableoilsandfatsdestinedfortheproductionof

babyfoodandprocessedcereal-basedfoodforinfantsandyoungchildren haveaproposedlimitof750 mg/kg.Infantformula,follow-onformula,and foodsforspecialmedicalpurposesintendedforinfantsandyoungchildren haveproposedlimitsof125 mg/kgforpowdersand15 mg/kgforliquids. TheEChasalsodiscussedbringing ﬁsh/marineoilsintotheexisting publishedlimitsforboundglycidol,althoughno ﬁnaldecisionshavebeen madeatpresstime.

References

[1]HrncirikK,vanDuijnG.Aninitialstudyontheformationof3-MCPDestersduring oilreﬁning.EurJLipidSciTechnol2011;113:374 9.

[2]MatthäusB,PudelF,FehlingP,VosmannKL,FreudensteinA.Strategiesforthe reductionof3-MCPDestersandrelatedcompoundsinvegetableoils.EurJLipidSci Technol2011;113:380 6.

[3]PudelF,BeneckeP,FehlingP,FreudensteinA,MatthäusB,SchwafA.Onthe necessityofedibleoilreﬁningandpossiblesourcesof3-MCPDandglycidylesters.Eur JLipidSciTechnol2011;113:368 73.

[4]DestaillatsF,CraftBD,SandozL,NagyK.Formationmechanismsofmonochloropropanediol(MCPD)fattyaciddiestersinreﬁnedpalm(Elaeisguineensis)oiland relatedfractions.FoodAdditContamA2012;29:29 37.

[5]DestaillatsF,CraftBD,DuboisML,NagyK.Glycidylestersinreﬁnedpalm(Elaeis guineensis)oilandrelatedfractions.PartI:formationmechanism.FoodChem 2012;131:1391 8.

[6]NagyK,SandozL,CraftB,DestaillatsF.Mass-defect ﬁlteringofisotopesignaturesto revealthesourceofchlorinatedpalmoilcontaminants.FoodAdditContam 2011;28:1492 500.

[7] SmidrkalJ,TesarováM,HrádkováI,BercíkováM,AdamcíkováA,FilipV.Mechanismofformationof3-chloropropan-1,2-diol(3-MCPD)estersunderconditionsof thevegetableoilre ﬁning.FoodChem2016:124 9.

[8]SpungenJH,MacMahonS,LeighJ,FlanneryB,KimG,ChirtelS,SmegalD. EstimatedUSinfantexposuresto3-MCPDandglycidylestersfromconsumptionof infantformula.FoodAdditContamA2018:1085 92.

[9]DivinováV,SvejkovskáB,DolezalM,VelísekJ.Determinationoffreeandbound3chloropropane-1,2-diolbygaschromatographywithmassspectrometricdetection usingdeuterated3-chloropropane-1,2-diolasinternalstandard.CzechJFoodSci 2004;22:182 9.

[10]WeibhaarR.Determinationoftotal3-chloropropane-1,2-diol(3-MCPD)inedible oilsbycleavageofMCPDesterswithsodiummethoxide.EurJLipidSciTechnol 2008;110:183 6.

[11]ZelinkováV,SvejkovskáB,DolezalM,VelísekJ.Fattyacidestersof3-chloropropane1,2-diolinedibleoils.FoodAdditContam2006;23:1290 8.

[12]HainesTD,AdlafKJ,PierceallRM,LeeI,VenkitasubramanianP,CollisonM.Direct determinationofMCPDfattyacidestersandglycidylfattyacidestersinvegetableoils byLC-TOFMS.JAmChemSoc2011;88:1 14.

[13]KazeN,SatoH,YamamotoH,WatanabeY.Bidirectionalconversionbetween3monochloro-1,2-propanediolandglycidolincourseoftheprocedureofDGFstandardmethods.JAmOilChemSoc2011;88:1143 51.

[14]IARC(InternationalAgencyforResearchonCancer).Someindustrialchemicals.In: IARCmonographsontheevaluationofcarcinogenicriskofchemicalstohumans,vol. 77.Lyon,France:InternationalAgencyforResearchonCancer;2000.p.469 86.

[15]BakhiyaN,AbrahamK,GürtlerR,AppelKE,LampenA.Toxicologicalassessmentof 3-chloropropane-1,2-diolandglycidolfattyacidestersinfood.MolNutrFoodRes 2011;55:509 21.

[16] FoodSafetyCommissionofJapan(FSCJ).Considerationsonglycidolanditsfattyacid estersinfoods.RiskAssessRep-NovelFoods&FoodAddit2015.FS/185/2015,Japan. [URL: http://www.fsc.go.jp/english/evaluationreports/others/annex_glycidol_26-52.pdf

[17]ChoWS,HanBS,NamKT,ParkK,ChoiM,KimSH,JeongJ,JangDD.Carcinogenicitystudyof3-monochloropropane-1,2-diolinsprague-dawleyrats.Food ChemToxicol2008;46:3172 7.

[18]EuropeanCommissionHealthandConsumerProtectionDirectorate.Opinionofthe scientiﬁccommitteeonfoodon3-monochloro-propane-1,2-diol(3-MCPD).2001.

[19]IARC(InternationalAgencyforResearchonCancer).IARCmonographsonsome chemicalspresentinindustrialandconsumerproducts.Food&DrinkWkly:3Monochloro-1,2-Propanediol2016:349 74.

[20]SchilterB,ScholzG,SeefelderW.Fattyacidestersofchloropropanolsandrelated compounds:toxicologicalaspects.EurJLipidSciTechnol2011;113:309 13.

[21]BuhrkeT,WeißhaarR,LampenA.Absorptionandmetabolismofthefood contaminant3-chloro-1,2-propanediol(3-MCPD)anditsfattyacidestersbyhuman intestinalcaco-2cells.ArchToxicol2011;85:1201 8.

[22]AbrahamK,AppelKE,Berger-PreissE,ApelE,GerlingS,MielkeH, CreutzenbergO,LampenA.Relativeoralbioavailabilityof3-MCPDfrom3-MCPD fattyacidestersinrats.ArchToxicol2013;87:649 59.

[23]AppelKE,AbrahamK,Berger-PreissE,HansenT,ApelE,SchuchardtS,VogtC, BakhiyaN,CreutzenbergO,LampenA.Relativeoralbioavailabilityofglycidolfrom glycidylfattyacidestersinrats.ArchToxicol2013;87:1649 59.

[24]WHO.Safetyevaluationofcertainfoodadditivesandcontaminants,3-chloro-1,2propanediol.WHOFoodAdditSer2002;48. http://www.inchem.org/documents/ jecfa/jecmono/v48je18.htm

[25]EuropeanCommissionHealthandConsumerProtectionDirectorate.Commission regulation(EC)No1881/2006of19December2006:settingmaximumlevelsfor certaincontaminantsinfoodstuffs.2006.

[26]FSANZ.Chloropropanolsinfood,ananalysisofthepublichealthrisk;technicalreport seriesNo.15;foodstandards,Australia/NewZealand.2003. http://www.foodstandards. gov.au/publications/Documents/TechnicalReportChloropropanolReport11Sep03. doc.

[27]CodexAlimentarius.Codexgeneralstandardforcontaminantsandtoxinsinfoodand feed,codexstan193 1995;amended.2012. www.codexalimentarius.org/download/ standards/17/CXS_193e_2012.pdf

[28]U.S.FoodandDrugAdministration.Guidancelevelsfor3-MCPD(3-chloro-1,2propanediol)inacid-hydrolyzedproteinandasian-stylesauces.CompliancePolicy GuideSection500.500.March14,2008. http://www.fda.gov/ICECI/Compliance Manuals/CompliancePolicyGuidanceManual/ucm074419.htm

[29]HealthCanada.Canadianstandards(maximumlevels)forvariouschemicalcontaminantsinfoods.Modiﬁed.June28,2012. http://www.hc-sc.gc.ca/fn-an/securit/chemchim/contaminants-guidelines-directives-eng.php

[30]EuropeanCommission.Commissionregulation(EU)2018/290of26February2018 amendingregulation(EC)No1881/2006asregardsmaximumlevelsofglycidylfatty acidestersinvegetableoilsandfats,infantformula,follow-onformulaandfoodsfor specialmedicalpurposesintendedforinfantsandyoungchildren.2018.

CHAPTER2

Formationmechanisms

BrianD.CraftandFrédéricDestaillats NestléResearchCenter,FoodScienceandTechnologyDepartment,Lausanne,Switzerland

Introduction

Sincethepublicationof[1]andtheheightenedawarenessoffattyestersof monochloropropanediol(MCPD-FE)inrefinededibleoils,thecircumstancessurroundingtheirformationhavebeensubjecttolargeamountsof speculation.Forinstance,someresearchersspeculatedthatprecursorsfor MCPD-FEformation(e.g.,chlorineanddiacylglycerols)arepresentin partiallyrefinedoils[2].Otherresearcherssuspectedthattherefining processresultsintheuncontrolledintroductionofcertaincompoundsto theoils(e.g.,inorganicchloridesinthestrippingstream),soitshouldbethe firstplacetoexploremitigationstrategies[3].Further,amidanalytical developmentsinMCPD-FEquantification,anotherfamilyofcompounds wasdiscoveredinrefinededibleoils,namelythefattyestersofglycidol (G-FE).G-FEwerefoundtobepartiallyresponsibleforinflationofthe resultsofMCPD-FEquantificationsduetothegenerationofartifacts duringsamplepreparationbeforeanalysisusingindirectmethods[4].

Despitetheseearlyhurdles,somerecentbreakthroughsweremadeby Refs.[5 7]ontheformationmechanismsofbothMCPD-FEandG-FE duringpalmoilrefining.Withinthischapterwewilltakeafocusedlook atthestatusoftheliteraturetodateasitpertainstotheformationpathways ofMCPD-FEandG-FEinrefinededibleoils.Criticaltopicswillbe covered,includingthemostprevalentprecursorcompoundsanddetailed formationmechanismsresponsibleforthegenerationoftheseprocess contaminantsduringoilproductionandrefining.BecausebothMCPD-FE andG-FEhavebeenfoundinthehighestaverageabundanceinpalmoil, themajorityoftheresearchreviewedhereininvolvescrudepalmoil productionandrefining.

ProcessingContaminantsinEdibleOils

ISBN978-0-12-820067-4 https://doi.org/10.1016/B978-0-12-820067-4.00004-8

MCPDesters

Precursors

Chlorineisubiquitousinnature.Thus,onecanspeculateaboutawide varietyofchlorinesources,whetherorganicorinorganic,aspotential precursorcompoundstoMCPD-FEformationduringedibleoilproduction.Further,ahostoflipidtypesandcompositions(e.g.,acylglycerols, phospholipids,glycolipids)areavailableintherawmaterialsusedtoproduceedibleoils.Manyoftheselipidscouldtheoreticallyinteractwith chlorinesourcesandresultintheformationofMCPD-FEduringoil refining.Thecriticalprecursorsresponsible,however,aremostlydependent ontheoiltype,quality,and,toalesserdegree,thecircumstances ofmanufacture,aswillbedescribedbelow.Giventhatrefinedpalmoil specificallyhasbeenshowntocontainsignificantlevelsofMCPD-FE (2.7mg/kg)[8],ithasbeenexclusivelyusedasamodelmatrixinthe literature.

The firstquestionoftenraisedregardingMCPD-FEprecursorsisthe originofchlorineinvolvedintheMCPD-FEreactionduringoilrefining andwhyitispotentiallymoreabundantincrudepalmoil(CPO)in comparisontoothercrudevegetableoils[9].Recently[5],demonstrated thatmanysourcesofcovalentlyboundinorganicchlorineexistatppm (mg/kg)levelsincrudepalmoil,includingFeCl3,FeCl2,MgCl2,and CaCl2.Further,a “pool” (n ¼ 300)oforganicmonochlorinatedcompoundswasalsofoundanditappearstoundergoatransformation throughoutthestagesofoilrefiningwithcertaincompoundsbeingformed whileothersdecomposeovertime.Inordertoelucidatethecompositionof themorepredominantchlorine “donor” compounds[5],usedLC-MSn in theframeworkofmodelexperiments.Thereintheauthorsidentifieda specificfamilyofchlorinatedcompoundspresentinboththelipids extractedfromhand-pickedMalaysianpalmfruitsandcommerciallyprocuredCPOsamples. Fig.2.1,takenfromRef.[5];showstheproposed structureandchemicalformulasofthismonochlorinatedfamilyofcompounds.Theauthorssuggestthatgiventheirstructuralsimilaritiesto phytosphingosines,itisperhapsmorelikelythatthechlorinedonors identifiedareendogenousplantmetabolitesasopposedtochlorinated contaminantsintroducedtotheoilpalm’sdirectenvironmentduring growthandmaturation.Extrapolatingfromthishypothesis,onemighttend tothelogicthattherawmaterialsintendedforproductionofeachedible vegetableoilhavetheirownreactive-chlorinepoolcapableofdonating chlorineduringoilrefiningandultimatelyresultinginMCPD-FE generation.

C42H83O

4NCl700.60280

C42H85O4NCl702.61807

C42H83O5NCl716.59723

C42H85O5NCl718.61357

C42H85O6NCl734.60809

Figure2.1 Proposedstructureandchemicalformulasofanorganochlorinefamilyof compoundsfoundincrudepalmoil. (ReprintedwithpermissionfromNagyK,SandozL, CraftBD,DestaillatsF.Mass-defect ﬁlteringofisotopesignaturestorevealthesourceof chlorinatedpalmoilcontaminants.FoodAdditContam2011;28:1492 1500).

IntermsofthemostpredominantlipidprecursorsofMCPD-FEin edibleoils[1],proposedthattheremaybealinkbetweenthecontentof diacylglycerols(DAGs)inrefinededibleoilsandtheirMCPD-FElevels. ThisassertionwaslikelyduetothefactthatthehighestMCPD-FElevels wereobservedwithinthefruitpulpoilsanalyzed.Fruitpulpoils,suchas oliveandpalm,areknownforhavinghighDAGcontentscomparedto seedoils,duetothegreaterprevalenceoflipolyticreactionsduringharvest [10].Thiscorrelation,however,hasbeendisprovedinrecentliterature [11,12].AlthoughDAGscouldpotentiallyreactwithchlorinedonors duringoilrefiningandresultintheformationofMCPD-FE,theyarenot themostcriticallipidprecursorsoftheseprocesscontaminants.Further, lipidssuchasmonoacylglycerols(MAGs),phospholipids,andglycolipidsare largelyremovedduringoildegummingandarenotpresentduringthelater stagesofoilrefining[10].

BecausethebulkofMCPD-FEhavebeenshowntobegenerated duringoildeodorization[2,13],theentiretyoftheaforementionedlipid classesisnotexpectedtobegreatlyinvolvedinMCPD-FEformation reactions.Thisofcourseleavesthetriacylglycerols(TAGs)upforconsideration.TAGscanrepresentmorethan90% 95%(v/v)ofreﬁnedvegetableoils,whetherpressedfromnuts,seeds,orfruitpulps.TAGsare, therefore,themostlogicalcriticallipidprecursoravailableforMCPD-FE formationduringoildeodorization.TheresultsofinvitrothermalreactionexperimentscarriedoutbyRef.[7]appeartoconﬁrmthishypothesis [7].demonstratedincontrolledconditionsthatTAGs,notDAGs,are preferentiallyreactingwithchlorinedonorstoformMCPD-FE.

Formationpathways

Aspreviouslymentioned,MCPD-FEareformedalmostcompletelyduring thedeodorizationunitoperationofedibleoilrefining.Assuch,themajorityofscientificresearchconductedonMCPD-FEformationpathways hasbeencarriedouteitherinconditionsmimickingoildeodorizationor withinbench-top,pilotscale,orcommercialdeodorizationunits[7]. showedthroughinvitroexperimentsthat3-MCPDdiesters,whicharethe mostpredominantformofMCPD-FEinrefinedoils[14],canbegenerated attemperaturesaslowas180 200 C.Itfollowsfromthisobservationthat withineithertypeofedibleoilrefining(chemicalorphysical),thetypical deodorizationconditionsemployedstronglyfavorMCPD-FEformation. Forexample,typicalchemicalrefiningoperationsforpalmoilinvolvea deodorizationstepataround240 C,whereasphysicalrefiningoperations caninvolveadeodorizationstepatevenhighertemperatures(260 270 C) inordertoremoveexcessfreefattyacids(FFAs)[10].

InordertodeterminetheoriginofchlorineinvolvedinMCPD-FE formationduringoilrefining[2,11],examinedthecontentofchloride ionspresentinoilspre-andpost-deodorization.Theythenattemptedto correlatetheselevelswiththeultimatelevelsofMCPD-FEobservedinthe fullyrefinedoils.Unfortunately,littleornocorrelationwasobserved[3]. attemptedtodetermineifthechlorineresponsibleforMCPD-FEformation wasoriginatingfromthestrippingsteamappliedduringoildeodorization, butwithasimilarnegativeresult.Onlyrecently[5],demonstratedthatboth inorganicandorganicchlorinatedcompoundsarepresentatppm(mg/kg) levelsinpartiallyrefinededibleoils.Further,theauthorsdemonstratedthat thethermaldecompositionoforganicchloride-containingcompoundsin CPOwasfoundtocoincidestronglywiththeevolutionof3-MCPD diestersduringthethermaltreatmentofCPO(see Fig.2.2).Giventhat 3-MCPDdiestersarethepredominantclassofMCPD-FEinrefinedoils, thestudyof[5]hasprovencausalitybehindthegreatestportionof MCPD-FEformedduringthedeodorizationofedibleoils.

Agivenorganic(orinorganic)chlorinatedcompoundmaydecompose atacertaintemperature,abovewhichthereleasedreactivechlorinecan theninteractwithTAGandresultintheformationofMCPD-FE.Thefact thatthegraphicin Fig.2.2 crossesat180 C,whereastheformationtemperatureofMCPD-FEinthestandardizedinvitroexperimentsof[7]was 180 200 C,isquiteacoincidence.Inordertouncoverwhetheracertain reactivechlorineintermediatewasmostresponsibleforMCPD-FE

Sumoforganochlorines SumofMCPDs

Figure2.2 Thesimultaneousdecompositionofsomekeyorganochorines(n ¼ 8) monitoredandtheformationof3-MCPDdiestersduringthethermaltreatmentof crudepalmoil. (ReprintedwithpermissionfromNagyK,SandozL,CraftBD,DestaillatsF. Mass-defect ﬁlteringofisotopesignaturestorevealthesourceofchlorinatedpalmoil contaminants.FoodAdditContam2011;28:1492 1500).

formation[5],monitoredthedecompositionoftheorganochlorinesfound incrudepalmoilsviaLC-MSn experiments.Theauthorsreportedthat hydrogenchloride(HCl)isatypicalthermaldecompositionproductofthe organochlorinepoolmonitored.Thus,HClcouldprovetobethepredominantformofreactivechlorineresponsibleforMCPD-FEformation duringoildeodorization.

SeveralpotentialformationmechanismsofMCPD-FEhavebeen recentlyreviewedintheliterature[15,16].Giventhatmanyofthepast mechanisticstudiesofMCPD-FEformationwerecarriedoutinhydrophilicmedia,ithaslongbeensuggestedthatTAGunderwenthydrolysisto DAGasa ﬁrststepintheMCPD-FEformationreaction.DAGsthen reactedwithchlorinedonorcompoundsresultingintheformationof acyloxoniumionintermediatesandeventualnucleophilicsubstitutionof chlorideionontheglycerolbackbone[17].Morerecently,however,itwas proventhatTAGscanactdirectlyasasubstrateforMCPD-FEformation [7].Further[18],provedwithinfrared(IR)spectroscopythatheatingTAG inthepresenceofLewisacidscanleadtocyclicacyloxoniumionformation.Thus,isittheoreticallyplausibletoassumethatthetwopathways couldbefavoredinthecaseofhydrophobicsystemslikeoildeodorization.

OnepathwayinvolvesthereactionofTAGdirectlywithHClformedby thermaldegradationofchlorinedonors,nucleophilicsubstitutionofchlorideionontheglycerolbackbonetoformanMCPD-FE,and ﬁnallythe releaseofafattyacid.Thesecondmechanism ﬁrstinvolvestheformationof anacyloxoniumionintermediatecompound.ThesetwoMCPD-FEformationmechanismsweresummarizedbyRef.[7]andappearin Fig.2.3 [7]. alsoprovedthroughinvitroexperimentsthattheMCPD-FEformation reactionisregioselectiveandpreferentialtothesn-1(3)positionsonthe glycerolbackbone.

Becausethecriticalprecursorsandpredominantformationmechanisms havebeenelucidatedinthecaseofpalmoilproductionandrefining,itwas thenpossibletospeculateaboutthepotentialpredominantrootcauses responsibleforthemanifestationofMCPD-FEtherein[5].demonstrated thatchlorinatedcompounds(n ¼ 300found)canbemonitoredincrude palmoilsutilizingmass-defect filteringofisotopesignatures.Further,the authorsthendiscoveredthatthesesamecompoundscanbeutilizedto segregatecommercialpalmoilsamples(n ¼ 26)basedontheirprocessing stageusingmultivariatestatisticalanalysis(see Fig.2.4).Thegroupingof commercialsamplesfromcrudetorefined-bleachedtorefined-bleacheddeodorized,asshownin Fig.2.4,suggeststhatchlorinatedcompounds undergoatransformationthroughoutpalmoilproductionandrefining. Thistruthmightlendtothelogicthatchlorineissimilarlytransformed throughouttheagriculturalprocessinvolvedinpalmoilgrowth,maturation,andharvest. Fig.2.5 isaschematictakenfromRef.[19]andservesasa root-causeanalysisofMCPD-FEformationduringrefinedpalmoilproduction.Itsummarizespotentiallocations/sourcesfortheinfluxofchlorine fromtheenvironment,accumulationofthechlorineinthepalmplantand fruits,andtransformationofchlorineintomoreliposolubleformsduring CPOproduction,followedbytheresultantformationofMCPD-FE duringoilrefining.Asimilartypeofroot-causeanalysiscouldprove beneficialinthecaseofotherrefinedvegetableoilcropsandtheassessment oftheirpotentialforproductionofMCPD-FEafterharvestandrefining.

Glycidylesters

Precursors

GiventhehistorybehindthediscoveryofG-FEinedibleoils,researchers oftenpooledG-FEwithMCPD-FE(oftentermedMCPDestersand relatedcompounds)[3].Thisfactledtotheassumptionthatthese

Figure2.3 ProposedmechanismsfortheformationofMCPD-FEfromTAGathightemperaturesinthepresenceofHClevolvedfromthe thermaldecompositionoftraceorganochlorines.Twoputativepathwaysincludingtheformationofreactivecycloacyloxoniumionintermediate(upperpanel)oradirectnucleophilicsubstitutionreaction(lowerpanel)aredisplayed.Bothpathwaysresultintheformationof anMCPD-FEmoleculeandthereleaseofafattyacid. (ReprintedwithpermissionfromDestaillatsF,CraftBD,SandozL,NagyK.Formation mechanismofmonochloropropanediol(MCPD)fattyaciddiestersinreﬁnedpalm(Elaeisguineensis)oilandrelatedfractions.FoodAdditContam 2012;29:29 37).

Figure2.4 Principalcomponentanalysisofchlorine-containingcompounds(n ¼ 300) presentincrude,partially,andfullyrefinedpalmoilsamples(n ¼ 26).Thegroupingof oilsamplesbasedonrefiningstage(i.e.,fromcrudetorefined-bleached[RB],to refined-bleached-deodorized[RBD])suggeststhatthesecompoundsundergoa transformationduringoilprocessing. (ReprintedwithpermissionfromNagyK,SandozL, CraftBD,DestaillatsF.Mass-defect filteringofisotopesignaturestorevealthesourceof chlorinatedpalmoilcontaminants.FoodAdditContam2011;28:1492 1500).

compoundfamilieswereverycloselyrelatedandpotentiallysubjectto interconversion.Assuch,bothMCPD-FEandG-FEwerethoughttoshare thesameprecursors.Thisassumptionhasbeendisprovedrecently[20]. reportedastrongpositivecorrelationbetweenDAGlevelsaloneandthe amountofG-FEcontainedinreﬁnedpalmoils.Becausefruitpulpoilslike palmoilnaturallycontainhigherlevelsofDAG( 3% 4%)[21],thismay suggestpossiblecausalityastowhytheirdeodorizedcounterpartsalso containhigherlevelsofG-FEincomparisontoseedoilcrops,as demonstratedbyRef.[4].

ThestrongcorrelationbetweenDAGcontentsandG-FEformationhas essentiallybeenvalidatedonanindustrialscale[22].recentlyreportedthat high-DAGoilsmarketedforhealthandwellnesshadtobetakenoffthe marketpurportedlydueto “highlevels” ofG-FE.Further[23],demonstratedthatcommerciallyreﬁnedoilsrichinDAG(87%)cancontainmore than10-foldgreaterG-FElevelsrelativetooilswithlowerDAG (3.9% 6.8%)contents.TheonlyotherprovenlipidprecursortoG-FEis MAG[6].showedthatbothMAGandDAGcanresultinformationofGFEuponthermaltreatment,althoughformationfromDAGismost

Oil Palm Growth & Maturation

Biosynthesis of hydrophilic organochlorines in palm fruits

Palm Oil Production

Formation of lipophilic organochlorines during fruit bunch sterilization

Palm Oil Refining

Reaction of liposoluble organochlorines with palm oil triacylglycerols at high temperatures

Accumulation of inorganic chloride in the oil palm plant

Influx of inorganic chloride from the environment (e.g., KCl, NH4Cl, MgCl2, FeCl3, FeCl2) (5)

MCPD diester formation

Soil, Fertilizer, & Irrigation

Figure2.5 Root-causeanalysisofthefactorsinvolvedintheformationofMCPD-FE duringreﬁnedpalmoilproduction,including(1)chlorineinﬂuxfromtheenvironment,(2)accumulationofinorganicchlorideintheplant,(3)bioconversionofinorganicchloridestoorganochlorinesinpalmfruits,(4)formationofliposoluble organochlorinesduringfruitbunchsterilization,and(5)reactionofliposolubleorganochlorineswithTAGinpalmoilduringoildeodorization. (Reprintedwithpermission fromCraftBD,NagyK,SandozL,DestaillatsF.Factorsimpactingtheformationof monochloropropanediol(MCPD)fattyaciddiestersduringpalm(Elaeisguineensis)oil production.FoodAdditContam2012;29:354 361).

favored.ThefactthatMAGlevelsinreﬁned-bleachedoilsareoftenquite low(mean < 0.1%)[24],however,rendersthisreactionroutelesssigniﬁcantinthecaseofediblevegetableoils.

Formationpathways

TheformationofG-FEfromDAGduringoildeodorizationwasshownto besignificantattemperaturesatorabove230 C[11].Ref.[20]confirmed thistemperatureofG-FEformationbydeodorizingrefined-bleachedpalm oilfrom180to240 Catincrementsof20 C.G-FEvalueswerereportedas thesumoftheglycidyl-palmitate(16:0G-FE),stearate(18:0G-FE),oleate (18:1G-FE),linoleate(18:2G-FE),andlinolenate(18:3G-FE)species