Working Group Chair Presentation Book

AOAC SPSFAM ALLERGENS DRAFT SMPR Ͳ COMMENTS on ALLERGENS SMPR FINAL

Line Numbers(If Applicable)

Item

Comment

ProposedChange(s)

Response

Chocolateshouldbeincludedintothelistofpriorityallergens.If chocolateisaknownproblemthantheapplicabilityshouldclearly statethatchocolateisnotpossibletomeasureusingthevalidated method.

Nochange. Chocolateisanoptionalmatrixtobetestedfor candidatemethodthatclaimtoworkinchocolate.

139(table 2)

1

Chocolateisanimportantmatrixforpeanut,hazelnutandmilk.

Describethevalidationofprecisioninamoreprecisewaye.g.include numberoflevelsandreplicates

Additionalreference toAppendixDandFareadded

Shouldtheprecisiondataobtainedoverthewholeanalyticalrange?Numberoflevels?

2 56Ͳ65

Nochangerecommended. Thecommentistruebutthereis notanyprohibitionagainsttheLOQ=MDL.

116(table 1)

3

BydefinitiontheanalyticalrangecanonlystartwithanLoQ.MDLonlygivesayesorno.

Aftervalidation,LCͲMS/MSmethodswillbeusedforcomparisonwithELISAresults.Ancommercial ELISAis(often)calibratedtothewholeallergenicfoodwhileLCͲMS/MSiscalibratedtopeptides.Is comparabilityestablishedviareferencematerials?(again:traceabilityofLCͲMS/MStotheseRMsis mandatory!)

Nochange. TheworkinggroupdidnotagreetotieLCͲMS/MS resultstoELISAresults.

DiscusstraceabilityandcomparabilitytoELISAresults(note:thisSMPR discussapossiblereferencemethodforcGMPcompliance!)

4

NISTSRM2387isnotpurepeanutbutamixtureofroastedpeanut,sugar,partiallyhydrogenated vegetablesoilsandsalt.SeeNISTcertificate:proteincontentisgivenbutnotpeanutcontent.

DiscusssuitabilityofthisSRMintheworkinggroupandgiveconversion factor

Nochange. That’swillbeleftuptothemethodedeveloper.

5 96

Agree.ReplaceNISTSRM1549wuth1549a.

6 92

NISTSRM1549issupersededbyNIST1549a

DeleteNISTSRM1549

WorkingGroupagreedthatallresultstobe"reportedasppm ofthetargetallergeninfoodcommodity".

Discusstraceabilityintheworkinggroupanddiscussaconversion factor

NIST8445isawholeeggpowderwithagivenproteincontent.Howshouldamethoddeveloper traceittowholeeggwithoutconversionfactor?

7 85

AddareferencetoAppendixM: ValidationProceduresfor QuantitativeFoodAllergenELISAMethods. AppendixMdoes mandatetheuseofincurredsamples. AOACpolicyallowsfor bothkindsofsamples. Methoddeveloperdiscretion.

Recovery:Whatkindofsamplesisrequired?Spikedorincurred?ForELISAincurredispreferred.

WeshouldfollowtheguidelineforELISAwhichpreferincurred

8 67

Recovery:Howshouldamethoddeveloperdeterminethisparameter?Byspikingwithreference materialsorpeptidesoradifferentmaterial.Oneshouldrememberthatitisnotallowedtousea referencematerialforcalibrationANDspiking!Ifpeptidesareusedforcalibration,howwas traceabilityestablished?

Nochangerecommended. Methoddevelopmentissuenot SMPRissue.

Discussintheworkinggroupandremembertosolvethetraceability problem

9 67

Sincereproducibilitydeterminationisonlypossiblebyacollaborativestudy,anintraͲlaboratory reproducibilityshouldbedefinedtoeasesingleͲlabvalidationsatthebeginning

Inlcudeanewclauseafterrepeatabilityanddescribethevalidationto bedone

Nochange. AllpreviousSMPRsusedRSDRandRSDr.

10 62

7

MDL:Howshouldamethoddeveloperestimatethisparameter?Byusingblankmatricesorblank matricesspikedwithreferencematerials/peptides?Howmanyreplicates?Wehaveveryclear guidelinesforallergendeterminationbyELISAͲwhynotfor“ReferencemethodsforcGMP compliance”? LoQ:Howshouldamethoddeveloperdetermineorevenestimatethisparameter?Byusing referencematerialsorpeptidesolutionsorblankmatricesorblankmatricesspikedwithreference materials/peptides?Howmanyreplicates?Wehaveveryclearguidelinesforallergen determinationbyELISAͲwhynotfor“ReferencemethodsforcGMPcompliance”?

ReferencetoAppendixM: ValidationProceduresfor QuantitativeFoodAllergenELISAMethodsaddedtoSMPR. SMPRwillalsorefertoFDAand/orEPAdefinitionforMDL.

11 50

DiscussintheworkinggroupmaybefollowELISAguidelines

ReferencetoAppendixM: ValidationProceduresfor QuantitativeFoodAllergenELISAMethodsaddedtoSMPR.

12 46

DiscussintheworkinggroupmaybefollowELISAguidelines

Line108ofversion revised torecommend"LOQ,MDL,recovery andprecision"dataforeveryclaimedmatrix.

includeasentenceforeachparameterthatexplainstheparameterͲ specificvalidation

LoQ,MDL,recoveryandprecisiondataneedtobedeterminedforeveryclaimedmatrix

13 46Ͳ69

BytakingthelatestpublishedVITALreferencedosesC18(FoodChem.Toxicol.63:9Ͳ17,2014)itis obviousthattheMDLs/LoQsintable1arenotsufficientwhenafoodisanalyzedthatisconsumed inaservicesizeofmorethan50g. LowerMDL/LoQappropriatelytothefollowingtable.Note:C19 Hazelnut:Referencedoseasprotein:0.1mg;Referencedoseasallergenicfood:0.64mg; Minimumconcentrationtobequantifiedwhenconsuming50gfood:12.8mg/kgandfor200g3.2 mg/kg.

Nochange. WorkingGroupdiscussedon3/3/2016.Thereare multipleVITALswithdifferentmaximumpermissiable concentrations. TheWorkingGroupconsensusisthatnoneof theVITALsareinternationalconcensusnstandards,anddeclined toresettheLOQsorMDLs basedonVITALmaximum permissiableconcentrations.

116(table 1)

Milk:Referencedoseasprotein:0.1mg;Referencedoseasallergenicfood:3.03mg;Minimum concentrationtobequantifiedwhenconsuming50gfood:60.6mg/kgandfor200g15.2mg/kg.

ChangeMDLs/LoQintable1accordingtotheVITALvaluesand calculationsgivenundercomments.Discussintheworkinggroup

14

Peanut:Referencedoseasprotein:0.2mg;Referencedoseasallergenicfood:0.8mg;Minimum concentrationtobequantifiedwhenconsuming50gfood:16mg/kgandfor200g4mg/kg.

Wholeegg:Referencedoseasprotein:0.03mg;Referencedoseasallergenicfood:0.25mg; Minimumconcentrationtobequantifiedwhenconsuming50gfood:4.8mg/kgandfor200g1.2 mg/kg.

Howshouldamethoddeveloperprovethattheselectedpeptidesarenot“too”specifice.g.a sequenceisusedthatisnotpresentineverycommerciallyavailablepeanutorhazelnutvariety.On theopposite,iftheselectedpeptidesarenotspecificenough,nearbotanicalrelativesaredetected whicharemaybenotallergenicorregulated(seeprunusmahalebexample).

Atminimumachapterdescribingtheknownspecificities/selectivities shouldbeprovided.(Note:Unknownoccurrenceofpeptidesthatare notfromaallergenicsourcewillalwaysoccurinthefuture,seealso prunusmahaleb)

Nochange. Theworkinggroupdidnotagree.

15

Nochange. Theworkinggroupdidnotagree.

Whataretheminimumperformancecriteriaforpeptideselection?

Includecriteriaforpeptideselectionorgivereference

16

Nochange. WorkingGroupdiscussedon3/3/2016.Thereare multipleVITALswithdifferentmaximumpermissiable concentrations. TheWorkingGroupconsensusisthatnoneof theVITALsareinternationalconcensusnstandards,anddeclined toresettheLOQsorMDLs basedonVITALmaximum permissiableconcentrations. ANDE25

VITALvaluesarebasedonamountofproteinperservicesize.Therefore,thedefinitionofthefood allergensas“foodcommodities”withoutmentioningtheproteincontentwillestablishanonͲ comparabilitybetweenresultsobtainedbyanLCͲMS/MSmethodandVITALvalues.

Includesomeguidancefortheuserorletthemethoddeveloper describehiswayofestablishingtraceabilitytoVITALvalues

17

collaborativetest:ItshouldbecriticallycheckedifAppendixDissufficientinthecasewhereLESS than8participants(and/orLCͲMS/MSmachines)areavailable.Isthisstillcollaborativeor forbiddenatall?

Nochange. AOACpolicynotaworkinggroupdecision.

discussintheworkinggroup

18 9

Changetitleto“…selectedfoodallergens."

19 3

Thetitleisunclear

changeto“…selectedfoodallergens”

Thismeansamethodcomparisonbetweentheoriginalmethod(checkedbyanERP)andthis methodtransferredtoanotherlab.Arethereanyguidelinesforthiscase?Whatistheminimum requirednumberofmeasurementstobesurethatbothmethodsarecomparable?

Nochange. Methodcomparisionisnotaverification requirement.

20 9

Includeminimumrequirementsforverification

8