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KrishnaPrasadRai*,ShivSagar Chaudhary,Sovita Shrestha,Som Kanta Rijal,Bal Kumari
SharmaandJiwan P.Lama
DepartmentofFoodTechnologyandQualityControl,CentralFoodLaboratory,Kathmandu,
Nepal
Introduction Researchdesignincludingsampling Results&Discussionwithkey
findings Conclusions Recommendations
PresentationLayout
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ATotalDietStudy(TDS)hasbeenusedasatoolforestimatingthelevelofdietaryexposuretocertainchemicalcontaminantscomingfromdifferentfoodsourcesamonggeneralpopulationsince1960s.
ItisrecognizedasacosteffectiveapproachtoestimatingdietaryexposureofcontaminantsbytheWorldHealthOrganization(WHO)in1978.
WhatisTotalDietStudy(TDS)?
AflatoxinsB1,B2,G1andG2hasbeendefinedastotalaflatoxins(TA)byCodexAllementarius Commission(CAC),whicharenaturallyoccurringtoxincompoundsproducedasmetabolitesbythefungiAspergillus flavusandAspergillus parasiticus growingonavarietyoffoodandfeedproductsunderfavorabletemperatureandhumidity.
Likewise,aflatoxinsM1andM2arethesecondarymetabolitesofG1andG2,whicharefoundinanimalsmilkandmeat.
Similarly,theothermostcommonmycotoxins areochratoxins,tricothecenes,zearalenone andfumonisins.
WhatisTotalaflatoxins?
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Aflatoxinsarepotentiallyhazardoustohumanandanimalshavingstrongimmunosuppressive,mutagenic,teratogenicandcarcinogeniceffects(HusseinandJeffrey,2001).
ChemicalStructuresofAflatoxins
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Agency Food MRL ReferenceGoN, Nepal Peanuts, beans and other
cereals products20g/kg Food Standard,
(2067)CAC Peanuts 15g/kg Codex Standard 193-
1995CAC Brazil nut, Almond,
Pistachio10g/kg Codex Standard 193-
1995EU Peanut/Groundnut 15g/kg for
unprocessed(EC) No 1881/2006
4g/kg for proceesed
EU Almond 10g/kg for directhuman consumpton
(EC) No 1881/2006
PFA , India Beans and cereals 30g/kg PFA, (2004)
FDA All products except milkdesigned for human
20g/kg Cornell University(2013)
Country Commodities Mycotoxin Level of presence Reference
Syria Different milks AFM1 >20 to 765 ng/l. Ghanem & Orfi,(2009)
Iran Human breast milk AFM1 0.326.7 ng/kg). Sadeghi et al.,(2009)
Tunisia Tunisian foods OTA 3.5 5.3 ng/g Ghali et al., (2008)Zearalenone 10.4 11.8 ng/g.
Athens Breakfast cereals AFB1 mean 1.42 ng/g Villa and Markaki(2009).OTA Mean 0.18 ng/g
Iran Pistachio AFB1 9- 45.7ng/g Sheibani,Ghaziaskar, (2009).AFB2 ND- 8ng/g
Nepal Ground nuts AFB1 3.78-39.69ppb Rai and Chaudhari,(2011)
Pakistan Buffaloesmilk
AFM1 0.027 g/ L Hussain et al.,(2008)
Cows milk 0.044lg/L
Mycotoxinscontaminationinsomefoods
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Themajorobjectiveofthisworkwastofindoutthecontaminationleveloftotalaflatoxinsinsomedietaryrawfoods(viz.maize,peanutsandtheirproducts)aswellastoassessthecrudedietaryexposureofthosecontaminantstoNepalesepeople.
GeneralObjective
SamplecollectionfromdifferentdistrictsofNepal
Quantitativeanalysisfortotalaflatoxinsincollectedfoodsamples
Assessmentofcrudedietaryexposurelevel
SpecificObjectives
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Thisstudydidnotincludetheeffectsofstorageconditionforrawfoodsinsamplingarea.
Theresultisintheformofcombineaflatoxins(B1,B2,G1andG2)termedastotalaflatoxinssothattheindividualaflatoxin wasnotdeterminedinfoodsamples.
Limitationsofstudy
Maize Peanuts Maizebasedproducts(e.g.cornflakes,suzi grits,Lito,
popcornetc.), Peanut/nutbasedproducts(e.g.Dalmoth,peanutbutter,
nutcracker,proteaseetc.), Legumes(e.g.soybean,beansetc.)and Others(e.g.Uwa,wheat,buckwheat,etc.).
FoodSamples
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SamplingmethodClustersampling SamplingDistricts:32 Totalno.ofsamples322 Sampleweight:500gto1kg AllthesampleswerepackedinPEbagscarefullyand
storedindrystorageatroomtemperaturetilltheanalysiswascompleted.
Samplingplan
Commoditywisesamplingplan
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RegionwiseSamplingplan
Region
Food commodities
TotalMaize Peanut
MaizeBasedproducts
Peanut/Nutsbasedproducts
Legumes
Others
Eastern 26 30 9 5 3 2 75
Central 38 36 12 0 1 1 88
Western 18 16 13 5 0 21 73Mid-western 19 10 7 1 0 8 45Farwestern 13 10 9 6 0 3 41
Sub total 114 102 50 17 4 35 322
EnzymelinkedImmunosorbentassay (ELISA)HELICATotalAflatoxinsAssay(USA),wasusedfortheQuantitativedetectionforaflatoxinsB1,B2,G1andG2.
LOD
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ELISAKitReader
Readtheopticaldensity(OD)ofeachmicrowellwithamicrotiterplatereaderusinga450nmfilter
Allowtheparticulatemattertosettle,thenfilter5 10mloftheextractthroughaWhatmanNo1filterpaperandcollecttheclearfiltratetobetested.Thesample
isnowreadyfortesting
Allthesamplesweremixedandstirredmanuallyforaminimumof2minutes.
Weight20gofsievedportionofthethenadd100mloftheExtractionSolvent(70%methanol:30%DW)ina250mlstopperedVolumetricflask
Grindsampletotheparticlesizeoffine(50%passesthrougha20meshscreen).
Sampleextractionandanalysis
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Samplepreparation
S.N. TDS foods Contamination No. % of Contamination
1 Maize 80 70.17
2 Peanuts 74 72.54
3 Maize based
products
40 80
4 Peanut based
products
14 82.35
5 Legumems 3 75
6 Others 8 22.85
PresenceofTotalaflatoxinsinfoodsamples
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Regions Maize Peanut
MaizedBased Pdts
Peanut/nuts based pdts Legumes Others
Eastern nd to 17.4nd to35.7 1 to 13.4 6.3 to 16 1 to 2.6 nd
Centralregion nd to 30
nd to24.5
1.1 to23.1 - nd nd
Westernregion nd to 30 nd to 30 nd to 30 nd to 30 -
nd to12.3
Midwestern nd to 27 nd to 5.8 nd to 9.6 10.2 - nd to 4.5Farwestern 1 to 30 1.9 to 21 nd to 25 nd to 20.5 -
nd to22.1
TotalaflatoxinsindifferentregionofNepal
Averageaflatoxinsindifferentfoodcommodities
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TAexceedingthemandatorylimitofGoN
Food Commodities
ICMR RDA (g/d)
Total average aflatoxin (g/ kg of food)
Crude aflatoxin consumption (g/ day)
Reference weight in Kg
Male (g/kg b.w./day)
Female (g/kg b.w./day)
60kg*** 50kg***Maize 183 7.89 1.44 0.024 0.029Peanut* 40 8.11 0.32 0.005 0.006Maize based products 183 6.39 1.17 0.019 0.023Peanut based products* 40 8.65 0.34 0.006 0.006Legumes 26 0.54 0.014 0.0002 0.0003Others** 183 2.53 0.463 0.008 0.0093
CrudedietaryexposureofTA
*Takenfrombalanceddietfornormalwomen(highcostdiet)(Swaminathan,1991)**Othersproductsmeansmostlycerealbasedotherthanmaize***ReferenceweighttakenfromGopalan etal.,(2004)
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Food Commodities
ICMR RDA (g/day)
Region***Eastern Central Western Midwestern Far western
Male Femal
e Male Femal
e Male Femal
e Male Femal
e Male Femal
e
60kg 50kg 60kg 50kg 60kg 50kg 60kg 50kg 60kg 50kgMaize 183 0.012 0.014 0.024 0.029 0.024 0.028 0.025 0.029 0.045 0.054Peanut* 40 0.007 0.009 0.003 0.003 0.007 0.009 0.0005 0.0006 0.007 0.009Maize based products 183 0.027 0.032 0.013 0.016 0.013 0.016 0.015 0.018 0.029 0.036Peanut based products* 40 0.007 0.009 0 0 0.005 0.006 0.007 0.008 0.004 0.004Legumes 26 0.0006 0.0008 0 0 0 0 0 0 0 0Others** 183 0 0 0 0 0.016 0.019 0.001 0.001 0.008 0.009
CrudedietaryintakeofTA(Regionwise)
*Takenfrombalanceddietfornormalwomen(highcostdiet)(Swaminathan,1991)**Othersproductsmeansmostlycerealbasedotherthanmaize***ReferenceweighttakenfromGopalanetal.,(2004)0=Nototalaflatoxinfoundinsamplesornosampletaken^Resultsarein(g/kgb.w./day)
Among322rawfoodsamples,TAwasfoundin68.01%ofsamples;however,theaverageTAleveldidnotcrossedthemandatorystandardforfood(20ppb)orforfeed(30ppb).
Over80%ofpeanut,maizeandtheirproductswerefoundtobecontaminatedwithtotalaflatoxins.
Keyfindings
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Interestingly,18.32%ofrawfoodsampleshowedhigherconcentrationthantheMRLofGoN,wheremaize,peanut,theirproductsandothercontributedas15.78%,11.76%,11.76%,2.86%respectively.
CrudedietaryexposureofTAwasfoundtobehighestinFarwesternregion(0.045g/kgb.w./dayformaleand0.054g/kgb.w./dayforfemale),whileEasternregionshowedthelowestinrawmaize.
Keyfindingscontd..
Theaveragecontentoftotalaflatoxinsindifferentfoodsamplesandtheirdietaryexposuretohumanwasfoundtobewithinthesafelimit;however,68%sampleswerefoundtobecontaminatedindifferentconcentrations.
Moreover,about18%werefoundtobeheavilycontaminatedwithTA.
Therefore,astrategicinterventionprogramisrequiredatpolicylevelofgovernmenttocontrolsuchcontaminantsinfoodcommodities.
Conclusions
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NationalReferenceRecommendedDietaryAllowance(RDA)shouldbeestablished.
thegovernmentsideaswellasUNagencysuchasWHO,FAOcouldworkjointlyinfuturetoestablishtheNationalRDAinNepal.
Since,thisisapreliminaryworkonTDSinNepal,scopesforothercontaminantsaswellasotherfoodcommodities(rawandcookedstate)shouldbecoveredinfuturestudiesplan.
Recommendations
AnalyticalcompetencyismostvitalpartofTDSwork;thereforegovernmentaswellasdevelopmentpartnersshouldprioritizeonbuildingupoffacilityandcompetencyforinstrumentalanalysisforallkindsofcontaminantsincountry.
Recommendationscontd..
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WewouldliketoacknowledgeWHO,countryofficeforNepalforsupportingthisprogramin2012.
WearealsogratefultoDFTQCforgivingthisopportunitytoourteam.
Acknowledgements
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