YeastYeast andand TheirTheir UsesUses inin FeedFeed

Mika Alewynse, PhD Division of Animal Feeds Center for Veterinary Medicine Presented at the April 2010 AAFCO Feed Administrators Seminar, Hickory Knob State Park, South Carolina IngredientIngredient DefinitionsDefinitions

„ WhereWhere areare theythey inin thethe AAFCOAAFCO OfficialOfficial Publication?Publication? –– YeastYeast „ Section 96 –– FermentationFermentation productsproducts „ Section 36 –– VariousVarious otherother sectionssections „ Distillers and miscellaneous products, for example TentativeTentative DefinitionsDefinitions

„ If have enforcement discretion letter, are fine with CVM – Ingredient name establishes common or usual name „ AAFCO OP has Model Regulation 6(f) – Tentative definitions for ingredient shall not be used until adopted as official, unless no official definition exists or the ingredient has a common acceptable name that requires no definition, (i.e. sugar). „ Some firms try to use this clause to say no definition is required „ If no other definition exists, a tentative definition can be used „ What does your state law say? AppropriateAppropriate DefinitionsDefinitions

„ HowHow dodo II knowknow thethe appropriateappropriate definition?definition? –– YeastYeast areare describeddescribed usingusing thethe sectionsection 96,96, YeastYeast –– BacterialBacterial andand moldmold ingredientsingredients areare describeddescribed usingusing sectionsection 36,36, FermentationFermentation ProductsProducts –– AndAnd nevernever dodo thethe twotwo mixmix…….... AppropriateAppropriate DefinitionsDefinitions (cont)

„ So,So, howhow dodo II telltell whatwhat isis what?what? –– LookLook atat thethe scientificscientific namename ofof thethe organismorganism –– LookLook atat wherewhere thethe organismorganism isis listedlisted „ Types of yeast are identified in section 96, Yeast „ Bacteria and molds are identified in section 36, Fermentation Products „ Exceptions AppropriateAppropriate DefinitionsDefinitions (cont)

„ ExceptionsExceptions – Direct-fed Microorganisms, definition 36.14 „ Yeast (as defined elsewhere) „ Elsewhere is in section 96, Yeast – Enzymes „ Microbial enzyme sources should be listed using section 36, Fermentation products definitions – Alpha-galactosidase – Invertase – Lactase – Lipase AppropriateAppropriate DefinitionsDefinitions (cont)

„ II knowknow thethe section,section, whatwhat’’ss next?next? „ LikeLike allall ingredients,ingredients, readread thethe definitionsdefinitions closely!closely! „ TheThe texttext inin thethe definitiondefinition isis youryour friend!friend! YeastYeast DefinitionsDefinitions

„ IdentifyIdentify whatwhat isis inin thethe ingredientingredient andand itsits microbialmicrobial originorigin „ OnlyOnly aa fewfew organismsorganisms – Saccharomyces cerevisiae „ Saccharomyces boulardii reclassified as S. cerevisiae „ Label must use Saccharomyces cerevisiae – Kluyveromyces marxianus – Candida utilis, formerly Torula

„ TwoTwo broadbroad categoriescategories – Whole cells and solubles YeastYeast DefinitionsDefinitions (cont)

„ WholeWhole cellscells -- withwith andand withoutwithout mediamedia – Dead – “non-fermentative” „ 96.1 Primary dried yeast „ 96.3 Irradiated dried yeast „ Source of vitamin D „ 96.7 Candida (Torula) dried yeast „ 96.5 Grain distillers dried yeast „ T96.12 Hydrolyzed yeast – Viable – “direct-fed microbial” definitions „ 96.2 Active dry yeast „ 96.8 Yeast culture YeastYeast DefinitionsDefinitions (cont)

„ BrewersBrewers YeastYeast –– “non-fermentative” „ Obtained from the production of beer and ale „ Separate definition for distillers yeast, 96.5 –– 96.496.4 BrewersBrewers drieddried yeastyeast –– 96.1096.10 BrewersBrewers liquidliquid yeastyeast „ SolublesSolubles –– 96.996.9 MolassesMolasses yeastyeast condensedcondensed solublessolubles „ Liquid fermentation media –– T96.11T96.11 YeastYeast extractextract „ Cell contents of yeast YeastYeast DefinitionsDefinitions (cont)

„ 96.8 Yeast culture* is the dried product composed of yeast (Saccharomyces cerevisiae and/or Kluyveromyces marxianus) and the media on which it was grown, dried in such a manner as to preserve the fermenting activity of the yeast. The media must be stated on the label. *Note: No reference to media in main ingredient listing is required when yeast culture forms a component of a proprietary mixed feed. – Grown on ground corn, hominy feed

„ When is media listing required? „ Must be on single ingredient feed but could be listed on mixed feed label YeastYeast DefinitionsDefinitions (cont)

„ DefinitionsDefinitions areare oldold andand needneed updatingupdating – Novel yeast species should not be marketed under these definitions as there are concerns „ Particularly from ethanol production „ Pathogenicity and potential for toxin production – Botanical classification Saccharomyces is too broad – Asking for comments about proposed yeast culture definition and other yeast species with history of use in feed QuestionsQuestions

Images courtesy of The Microbe World at www.edu.pe.ca/southernkings/microclass.htm FermentationFermentation ProductsProducts inin FeedFeed

Mika Alewynse, PhD Division of Animal Feeds Center for Veterinary Medicine

Presented at the April 2010 AAFCO Feed Administrators Seminar, Hickory Knob State Park, South Carolina FermentationFermentation ProductProduct DefinitionsDefinitions

„ Section 36 in the OP „ Identify what is in the ingredient and its microbial origin „ Most are byproducts from other processes – Check that substance of interest is still there „ Three broad categories – Whole cells, solubles, mixtures

„ Terminology – Mycelium – composed of cells from a mold – Biomass – composed of cells from a bacteria – Water soluble – exactly that – Broth – growth media, generally from mold fermentation FermentationFermentation ProductProduct DefinitionsDefinitions (cont)

„ 36.236.2 ExtractedExtracted ______presscakepresscake „ 36.1336.13 ExtractedExtracted ______(_(presscake, meal or pellets)_)_ – Both composed of mycelium (whole organism) – Obtained from an organism „ Penicillium or Streptomyces – Obtained from a particular production practice „ Citric acid fermentation „ Ask about whether organism historically used for this purpose „ Firms developing novel organisms and may take advantage of lack of identity FermentationFermentation ProductProduct DefinitionsDefinitions (cont)

„ 36.936.9 UndriedUndried extractedextracted ______solidssolids andand fermentationfermentation solublessolubles –– ComposedComposed ofof myceliummycelium (whole(whole organism)organism) andand brothbroth –– ObtainedObtained fromfrom anan organismorganism „ Penicillium or Streptomyces –– ObtainedObtained fromfrom aa particularparticular productionproduction practicepractice „ Citric acid fermentation –– MixtureMixture ofof thingsthings FermentationFermentation ProductProduct DefinitionsDefinitions (cont)

„ 36.436.4 DriedDried extractedextracted ______fermentationfermentation solublessolubles –– ComposedComposed ofof extractedextracted brothbroth (substances(substances mustmust bebe solublesoluble toto bebe inin aa broth)broth) –– ObtainedObtained fromfrom anan organismorganism „ Penicillium or Streptomyces –– ObtainedObtained fromfrom aa particularparticular productionproduction practicepractice „ Citric acid fermentation FermentationFermentation ProductProduct DefinitionsDefinitions (cont)

„ 36.636.6 DriedDried ______fermentationfermentation extractextract „ 36.736.7 DriedDried ______fermentationfermentation solublessolubles – Both composed of water soluble materials, not particles – Only for production of enzymes „ Refer back to Table 30.1 in section 30 Enzymes for suitable organisms – If these ingredients are being marketed as a source of something other than an enzyme, look closely at the composition FermentationFermentation ProductProduct DefinitionsDefinitions (cont)

„ 36.1636.16 DriedDried LL--lysinelysine fermentationfermentation productproduct „ 36.1736.17 LiquidLiquid LL--lysinelysine fermentationfermentation productproduct – Both composed of biomass (bacterial cells) and L-lysine – Only acceptable organism is Corynebacterium glutamicum – Only as source of L-lysine FermentationFermentation ProductProduct DefinitionsDefinitions (cont)

„ 36.1136.11 DriedDried ______fermentationfermentation productproduct „ 36.1236.12 LiquidLiquid ______fermentationfermentation productproduct – Very general definitions – Widely used in feed for multitude of purposes „ Including forage and grain inoculants – Can be composed of whole organism and/or fermentation media/broth – Obtained from a listed organism FermentationFermentation ProductProduct DefinitionsDefinitions (cont)

„ 36.1436.14 DirectDirect--fedfed microorganismsmicroorganisms (DFM)(DFM) –– ListedListed inin ingredientsingredients listlist ofof labellabel usingusing „ 36.11 Dried ______fermentation product „ 36.12 Liquid ______fermentation product or occasionally as „ a mycelium definition – Many extraction processes kill the organism –– CannotCannot bebe aa solublessolubles definitiondefinition „ Solubles do not contain whole microbial cell WhatWhat areare DFMDFM Products?Products?

„ ContainContain livingliving (viable)(viable) microorganismsmicroorganisms „ TwoTwo mainmain categoriescategories 1)1) FeedFeed inoculantsinoculants „ IntendedIntended toto affectaffect compositioncomposition and/orand/or digestibilitydigestibility ofof feedfeed componentcomponent „ MayMay oror maymay notnot bebe viableviable whenwhen consumedconsumed byby animalanimal „ ExamplesExamples ––InoculantsInoculants forfor silagesilage andand highhigh moisturemoisture graingrain WhatWhat areare DFMDFM Products?Products? (cont)

2)2) DFMDFM productsproducts areare intendedintended toto provideprovide livingliving (viable)(viable) microorganismsmicroorganisms forfor additionaddition toto animalanimal feedfeed –– WhyWhy dodo thethe firmsfirms wantwant toto dodo that?that? „ Scientific literature suggests beneficial effects in animals „ Mirrors consumer interest in “natural” foods NewNew OrganismsOrganisms

–– ApplicableApplicable toto DFMsDFMs andand feedfeed inoculantsinoculants „ ObtainObtain AAFCOAAFCO ingredientingredient definitiondefinition –– NoNo historyhistory ofof unsafeunsafe useuse // inhabitantinhabitant ofof GIGI tracttract –– PathogenicityPathogenicity –– NoNo commercialcommercial productionproduction ofof antibioticsantibiotics –– ManufacturingManufacturing datadata –– InformationInformation onon thethe intendedintended useuse MicroorganismsMicroorganisms

„ AcceptedAccepted organismsorganisms listedlisted inin FermentationFermentation productsproducts definitiondefinition 36.1436.14 „ UniqueUnique labellabel aspectsaspects –– organismsorganisms areare listedlisted usingusing 22 definitionsdefinitions –– TheThe GenusGenus speciesspecies asas listedlisted inin 36.1436.14 –– TheThe typetype ofof fermentationfermentation productproduct InoculantsInoculants andand thethe OPOP

„ ModelModel regulationsregulations –– GuaranteesGuarantees -- regulationregulation 4(g)4(g) „ StatementStatement forfor uniformuniform interpretationinterpretation andand policypolicy #23#23 –– RegistrationRegistration andand labelinglabeling ofof silagesilage additiveadditive productsproducts „ Guarantees, directions for use, suitable ingredients DFMsDFMs andand thethe OPOP

„ ModelModel regulationsregulations –– AcceptabilityAcceptability andand contentcontent -- regulationregulation 9(b)(4)9(b)(4) –– GuaranteesGuarantees -- regulationregulation 4(g)4(g) „ PetPet && specialtyspecialty petpet foodfood regulationsregulations –– GuaranteesGuarantees –– regulationregulation PF4(h)PF4(h) DFMDFM LabelsLabels

„ DFMDFM labelslabels areare notnot uniqueunique „ EverythingEverything thatthat isis neededneeded forfor aa ““regularregular”” feedfeed labellabel isis alsoalso neededneeded forfor DFMsDFMs „ So,So, whatwhat shouldshould bebe there?there? –– IfIf marketedmarketed asas DFM,DFM, regulationregulation 9(b)4II9(b)4II requiresrequires thethe statementstatement „ ““containscontains aa sourcesource ofof livelive (viable)(viable) naturallynaturally occurringoccurring microorganismsmicroorganisms”” CompleteComplete LabelLabel

„ NameName „ PurposePurpose (content)(content) statementstatement „ GuaranteesGuarantees „ IngredientIngredient listlist „ DetailedDetailed useuse directionsdirections „ Caution/Caution/ warningwarning statementsstatements „ Manufacturer/Manufacturer/ distributordistributor identifieridentifier „ FirmFirm namename andand locationlocation „ NetNet contentcontent ProblemProblem AreasAreas

¾ WhatWhat organismsorganisms ¾ IngredientIngredient listlist ¾ UseUse directionsdirections ¾ GuaranteesGuarantees ¾ ““UseUse byby”” datesdates (not(not mandated)mandated) WhatWhat OrganismsOrganisms toto Use?Use?

„ AcceptedAccepted organismsorganisms areare listedlisted inin definitiondefinition 36.1436.14 – Includes bacteria and fungi as listed – Yeast (as defined elsewhere) „ YeastYeast areare defineddefined inin SectionSection 9696 – Two yeast organisms are acceptable „ Saccharomyces cerevisiae „ Kluyveromyces marxianus „ Candida (Torula) definition is not for viable organisms WhatWhat OrganismsOrganisms toto Use?Use? (cont)

„ WhatWhat aboutabout microbesmicrobes inin thethe EnzymeEnzyme table,table, 30.1?30.1? –– ThereThere isis somesome overlap,overlap, butbut thethe organismorganism mustmust bebe inin definitiondefinition 36.1436.14 „ WhyWhy isis thethe listlist soso short?short? –– LivingLiving microbesmicrobes growgrow andand reproducereproduce „ Concerns about toxin production and ability to cause human and animal illnesses IngredientIngredient ListList

„ DoDo notnot listlist justjust thethe namename –– AspergillusAspergillus nigerniger „ UseUse FermentationFermentation definitionsdefinitions fromfrom SectionSection 3636 –– LiquidLiquid LactobacillusLactobacillus plantarumplantarum fermentationfermentation productproduct –– DriedDried AspergillusAspergillus oryzaeoryzae fermentationfermentation productproduct PotentialPotential DefinitionsDefinitions

„ Which fermentation ingredient? „ Look at the definitions – 36.11 Dried ____ Fermentation Product is the product derived by culturing ____ on appropriate nutrient media for the production of one or more of the following: enzymes, fermentation substances, or other microbial metabolites, and dried in accordance with approved methods and good manufacturing practices. Protein, amino acids, fat, fiber, cell count, enzyme activity or nutrient metabolite level shall be guaranteed where applicable…

– 36.12 Liquid ____ Fermentation Product is the liquid product derived by culturing or fermenting ____ on appropriate liquid nutrient media for the production of one or more of the following: enzymes, fermentation substances, or other microbial metabolites, and stabilized by approved methods in accordance with good manufacturing practices. Percent solids, cell count, enzyme activity or nutrient metabolite level shall be guaranteed where applicable… PotentialPotential DefinitionsDefinitions (cont)(cont)

„ ForFor definitionsdefinitions 36.1136.11 andand 36.12,36.12, fillfill inin thethe blankblank withwith organismorganism fromfrom 36.1436.14 ––DriedDried BacillusBacillus subtilissubtilis fermentationfermentation productproduct „ IfIf thethe definitiondefinition statesstates ““extract,extract,”” ““extracted,extracted,”” oror ““solubles,solubles,”” probablyprobably notnot suitablesuitable forfor viableviable productproduct YeastYeast inin aa DFMDFM productproduct

„ YeastYeast areare different!different! „ DoDo notnot useuse FermentationFermentation definitionsdefinitions – Dried Saccharomyces cerevisiae fermentation product „ UseUse yeastyeast definitionsdefinitions inin SectionSection 9696 „ TheThe yeastyeast mustmust bebe alive,alive, soso firmsfirms shouldshould checkcheck withwith theirtheir suppliersupplier – Active dry yeast, 96.2 – Yeast culture, 96.8 „ IfIf thethe definitiondefinition statesstates ““nonnon--fermentative,fermentative,”” yeastyeast areare deaddead andand itit isis notnot aa DFMDFM UseUse DirectionsDirections

„ RememberRemember thethe endend useruser „ IncludeInclude avoirdupoisavoirdupois unitsunits –– IfIf differentdifferent speciesspecies oror animalanimal classesclasses requirerequire differentdifferent amountsamounts ofof microbes,microbes, statestate thatthat „ Use rates often differ among species GuaranteesGuarantees

„ RegulationRegulation 4(g)4(g) –– GuaranteesGuarantees forfor microorganismsmicroorganisms shallshall bebe statedstated inin colonycolony formingforming unitsunits perper gramgram (CFU/g)(CFU/g) whenwhen directionsdirections areare forfor usingusing thethe productproduct inin grams,grams, oror inin colonycolony formingforming unitsunits perper poundpound (CFU/lb)(CFU/lb) whenwhen directionsdirections areare forfor usingusing thethe productproduct inin pounds.pounds. –– AA parentheticalparenthetical statementstatement followingfollowing thethe guaranteeguarantee shallshall listlist eacheach speciesspecies inin orderorder ofof predominance.predominance. GuaranteesGuarantees (cont)(cont)

„ EitherEither avoirdupoisavoirdupois oror metric,metric, correspondcorrespond toto thethe useuse directionsdirections „ IncludeInclude microorganismsmicroorganisms –– NotNot requiredrequired thatthat individualindividual speciesspecies bebe listedlisted separatelyseparately „ IncludeInclude colonycolony formingforming unitsunits –– NotNot cellcell countcount GuaranteesGuarantees (cont)(cont)

„ WhatWhat isis inin thethe guarantee?guarantee? BacillusBacillus subtilissubtilis………….1.1 millionmillion CFU/CFU/ gg

AddAdd 11 kgkg (1000(1000 g)g) toto eacheach tonton ofof completecomplete feedfeed

–– MicroorganismMicroorganism –– B.B. subtilissubtilis –– UnitsUnits –– 11 millionmillion CFU/CFU/ gg „ in metric since use directions are metric GuaranteesGuarantees (cont)(cont)

Total lactic acid bacteria…….1 x 106 CFU/ lb (Lactobacillus plantarum, Lactobacillus acidophilus, and Enterococcus faecium) Feed 2 ounces per head per day

– Bacterial type – Total lactic acid bacteria „ Cannot include yeast in this guarantee as yeast are not bacteria – Units – 1 million CFU/ lb – Microorganisms – L. plantarum, L. acidophilus, E. faecium GuaranteesGuarantees (cont)(cont)

LactobacillusLactobacillus plantarumplantarum ……...1...1 xx 10104 CFU/CFU/ lblb LactobacillusLactobacillus acidophilusacidophilus ……..1..1 xx 10104 CFU/CFU/ lblb EnterococcusEnterococcus faeciumfaecium ……………….1.1 xx 10106 CFU/CFU/ lblb FeedFeed 22 ouncesounces perper headhead perper dayday – Microorganisms – L. plantarum, L. acidophilus, E. faecium – Units – 104 and 106 CFU/ lb „ in avoirdupois since use directions are avoirdupois

„ Guarantees can be long, especially if there are multiple organisms OtherOther IssuesIssues

„ ““UseUse byby”” datesdates notnot requiredrequired – Why include them? „ DFM must contain living microorganisms „ Everything dies even with preservatives – Use of “use by” dates prevents questions concerning guarantees with an out-of-date product „ HeatHeat – Many organisms adversely affected by heat „ Pelleting „ Storage DFMDFM ClaimsClaims

„ Must be based on scientific data in animal species of interest with the specific microbial species – “Yeast supplementation can aid in maintaining cellulolytic bacteria population in the rumen of animals fed diets containing greater than 50% concentrate”

„ CPG Sec. 689.100 Direct-Fed Microbial Products http://www.fda.gov/ICECI/ComplianceManuals/CompliancePolicyGuidan ceManual/ucm074707.htm

„ CVM Policy On Competitive Exclusion Products http://www.fda.gov/AnimalVeterinary/NewsEvents/CVMUpdates/ucm12 7971.htm DFMDFM ClaimsClaims (cont)(cont)

„ UseUse ofof ““probioticprobiotic”” oror ““prebioticprebiotic”” onon thethe labellabel nownow acceptableacceptable „ GuidanceGuidance forfor IndustryIndustry -- ComplementaryComplementary andand AlternativeAlternative MedicineMedicine ProductsProducts andand theirtheir RegulationRegulation byby thethe FoodFood andand DrugDrug AdministrationAdministration -- http://www.fda.gov/RegulatoryInformation/Guidances/ucm144657.htm – “Probiotics may be regulated as dietary supplements, foods, or drugs under the Act, depending on the product's intended use” IssuesIssues forfor MicrobesMicrobes

„ TaxonomyTaxonomy changeschanges „ ProductionProduction ofof antimicrobialsantimicrobials „ NontraditionalNontraditional genusgenus speciesspecies „ BioengineeredBioengineered organismsorganisms –– FoodFood AdditiveAdditive approvalapproval neededneeded „ PathogenicityPathogenicity andand toxintoxin testingtesting –– DifferentDifferent animals,animals, differentdifferent susceptibilitiessusceptibilities Mikrobial Good Stuff Contains a source of live (viable) naturally occurring microorganisms Guaranteed Analysis Lactobacillus plantarum (min.) 1 x 105 CFU/ lb Lactobacillus acidophilus (min.) 1 x 105 CFU/ lb Enterococcus faecium (min.) 1 x 106 CFU/ lb or Lactic acid bacteria (min.) 1.2 x 106 CFU/ lb (Enterococcus faecium, Lactobacillus plantarum, Lactobacillus acidophilus)

Ingredients: Ground limestone, dried Enterococcus faecium fermentation product, dried Lactobacillus plantarum fermentation product, dried Lactobacillus acidophilus fermentation product, sodium sorbate (preservative), mineral oil

Directions for Use Feed 2 ounces per cow per day Feed 0.5 oz per pig per day Store at room temperature and in a dry place Use within 12 months of manufacture Manufactured by Mika Industries, 7519 Stand Place, Rockville, MD 20855 Net Wt 44 lb (20 kg) Manufactured 4/12/10 Questions?Questions?

Pictures courtesy of Tree of Life (www.tolweb.org/Ascomycota) EnzymesEnzymes UsedUsed inin FeedFeed

Mika Alewynse, PhD Division of Animal Feeds Center for Veterinary Medicine

Presented at the April 2010 AAFCO Feed Administrators Seminar, Hickory Knob State Park, South Carolina EnzymesEnzymes andand thethe CFRCFR

„ ApprovedApproved ingredientsingredients listedlisted in:in: –– TitleTitle 2121 CodeCode ofof FederalFederal RegulationsRegulations „ 582.1585 Papain „ 582.1685 Rennet „ 573.130 Aminoglycoside 3’- phosphotransferase II for use in bioengineered oilseed rape, tomato, cotton „ NothingNothing uniqueunique inin CFRCFR forfor enzymesenzymes –– NovelNovel enzymesenzymes „ Food additive regulation EnzymesEnzymes andand thethe OPOP

„ AcceptedAccepted enzyme/enzyme/ sourcesource organismorganism combinationscombinations listedlisted inin TableTable 30.130.1 „ UniqueUnique aspectaspect –– EnzymeEnzyme MarketingMarketing CoordinationCoordination documentdocument „ ModelModel regulationsregulations – Acceptability - regulation 9(b)(5) – Purpose – regulation 3(a)(3)(VI) – Guarantees - regulation 4(h) „ PetPet && specialtyspecialty petpet foodfood regulationsregulations – Guarantees – regulation PF4(h) EnzymeEnzyme SpecificSpecific UsesUses

„ WhyWhy areare therethere soso manymany differentdifferent enzymesenzymes usedused inin feed?feed? –– AddedAdded toto affectaffect processingprocessing ofof ingredientingredient „ amylase –– AddedAdded toto alteralter digestibilitydigestibility ofof feedfeed ingredientingredient „ phytase –– AddedAdded toto alteralter characteristicscharacteristics ofof digestadigesta „ xylanase EnzymeEnzyme SpecificSpecific UsesUses (cont)(cont)

„ There is no such thing as a generic animal feed – Multitude of species „ Companion animals, traditional livestock, non- traditional species (fish, llamas, ostriches) – All stages of an animal’s lifespan „ Neonate, growing, mature, breeding, elderly (primarily companion animals) „ Current supported use statements are animal species specific – Reduction of digesta viscosity with poultry diets „ Functionality statements are not – Hydrolyzes phytate EnzymeEnzyme SpecificSpecific UsesUses (cont)(cont)

„ PlantsPlants andand byproductsbyproducts areare largelarge proportionproportion ofof manymany feedsfeeds „ OftenOften containcontain undesirableundesirable componentscomponents –– Phytate,Phytate, reducesreduces phosphorusphosphorus availabilityavailability –– StachyoseStachyose andand raffinose,raffinose, lessless availableavailable sourcessources ofof energyenergy EnzymeEnzyme LabelsLabels

„ WhereWhere areare thethe enzymeenzyme regulationsregulations „ WhatWhat shouldshould bebe onon thethe labellabel „ TroubleTrouble spotsspots CompleteComplete LabelLabel

„ NameName „ PurposePurpose statementstatement „ GuaranteesGuarantees „ IngredientIngredient listlist „ DetailedDetailed useuse directionsdirections „ Caution/Caution/ warningwarning statementsstatements „ Manufacturer/Manufacturer/ distributordistributor identifieridentifier „ FirmFirm namename andand locationlocation „ NetNet contentcontent ProblemProblem AreasAreas forfor LabelsLabels

„ IngredientIngredient listlist „ UseUse directionsdirections „ GuaranteesGuarantees „ ““UseUse byby”” datesdates (not(not mandated)mandated) IngredientIngredient ListList

„ NoNo – Phytase – Phytase from Aspergillus niger – Aspergillus niger fermentation extract (phytase) „ YesYes – Aspergillus niger fermentation extract „ IfIf productproduct containscontains aa fermentationfermentation ingredientingredient – Use definition from Section 36, Fermentation Products IngredientIngredient ListList (cont)(cont)

„ CanCan II useuse anan organismorganism listedlisted inin 36.14,36.14, thethe DirectDirect--fedfed Microorganisms,Microorganisms, forfor anan enzymeenzyme product?product? „ AnAn enzymeenzyme andand thethe organismorganism thatthat producesproduces itit areare linked,linked, asas indicatedindicated inin TableTable 30.130.1 –– CannotCannot mixmix andand matchmatch PotentialPotential DefinitionsDefinitions

„ WhichWhich fermentationfermentation ingredient?ingredient? „ LookLook atat thethe definitionsdefinitions – 36.6 Dried ____ Fermentation Extract is the dried product resulting from extracting and precipitating by means of non-aqueous solvents or other suitable means, the water soluble materials from a fermentation conducted for maximum production of enzymes using a non-pathogenic strain of the microorganism in accordance with good manufacturing practices.

– 36.7 Dried ____ Fermentation Solubles is the dried material resulting from drying the water soluble materials after separation of suspended solids from a fermentation conducted for maximum production of enzymes using a non-pathogenic strain of the microorganism in accordance with good manufacturing practices. PotentialPotential DefinitionsDefinitions (cont)(cont)

– 36.11 Dried ____ Fermentation Product is the product derived by culturing ____ on appropriate nutrient media for the production of one or more of the following: enzymes, fermentation substances, or other microbial metabolites, and dried in accordance with approved methods and good manufacturing practices. Protein, amino acids, fat, fiber, cell count, enzyme activity or nutrient metabolite level shall be guaranteed where applicable…

– 36.12 Liquid ____ Fermentation Product is the liquid product derived by culturing or fermenting ____ on appropriate liquid nutrient media forfor the production of one or more of the following: enzymes, fermentation substances, or other microbial metabolites, and stabilized by approved methods in accordance with good manufacturing practices. Percent solids, cell count, enzyme activity or nutrient metabolite levellevel shall be guaranteed where applicable… PotentialPotential DefinitionsDefinitions (cont)(cont)

„ WhatWhat aboutabout enzymesenzymes obtainedobtained fromfrom plantsplants oror animals?animals? –– WhatWhat isis thethe commoncommon andand usualusual namename ofof thethe ““ingredient?ingredient?”” „ Dried pineapple „ Fig extract „ Dried pancreas „ Phytase canola UseUse DirectionsDirections

„ RememberRemember thethe endend useruser –– IncludeInclude avoirdupoisavoirdupois unitsunits –– IfIf differentdifferent speciesspecies oror animalanimal classesclasses requirerequire differentdifferent amounts,amounts, labellabel shouldshould statestate thatthat „ Phytase - use rates often differ among layers, broilers, turkeys, and swine ThoseThose @*!!@*!! GuaranteesGuarantees

„ RegulationRegulation 4(h)4(h) – Guarantees for enzymes shall be stated in units of enzymatic activity per unit weight or volume, consistent with label directions. The source organism for each type of enzymatic activity shall be specified, such as: Protease (Bacillus subtilis) 5.5 mg amino acids liberated/min./milligram. If two or more sources have the same type of activity, they shall be listed in order of predominance based on the amount of enzymatic activity provided. GuaranteesGuarantees (cont)(cont)

„ EnzymeEnzyme MarketingMarketing CoordinationCoordination documentdocument –– EitherEither avoirdupoisavoirdupois oror metric,metric, correspondcorrespond toto thethe useuse directionsdirections –– IncludeInclude sourcesource organismorganism –– IncludeInclude unitsunits –– ListList sourcessources byby contributioncontribution ofof enzymaticenzymatic activityactivity GuaranteesGuarantees (cont)(cont)

„ WhatWhat isis inin thethe guarantee?guarantee? ProteaseProtease ((BacillusBacillus subtilissubtilis)) 5.55.5 mgmg aminoamino acidsacids liberated/liberated/ minute/minute/ gramgram –– ProteaseProtease –– typetype ofof enzymeenzyme activityactivity –– B.B. subtilissubtilis –– sourcesource organismorganism –– mgmg aminoamino acidsacids liberated/liberated/ minute/minute/ gg –– unitunit ofof enzymaticenzymatic activityactivity GuaranteesGuarantees (cont)(cont)

„ GuaranteesGuarantees cancan bebe longlong – Protease (Bacillus subtilis) 5.5 mg amino acids liberated/ minute/ gram „ FirmsFirms cancan splitsplit andand cancan explainexplain unitsunits elsewhereelsewhere onon labellabel – Protease (Bacillus subtilis) 5.5 units*/ g • 1 unit of protease activity liberates 1 mg amino acid from casein/ minute • Should mention assay conditions, i.e., pH, temperature GuaranteesGuarantees (cont)(cont)

„ MostMost problematicproblematic areare thethe unitsunits „ WhatWhat isis there?there? mgmg aminoamino acidsacids liberatedliberated// minuteminute// gg –– mgmg aminoamino acidsacids liberatedliberated –– whatwhat thethe enzymeenzyme doesdoes –– minuteminute –– timetime unitunit –– gg –– unitunit ofof enzymeenzyme productproduct GuaranteesGuarantees (cont)(cont)

„ UnitsUnits areare enzymeenzyme specificspecific –– whatwhat enzymeenzyme doesdoes „ UnitsUnits areare alsoalso assayassay specificspecific forfor aa particularparticular typetype ofof activityactivity GuaranteesGuarantees--EnzymeEnzyme SpecificSpecific

mgmg aminoamino acidsacids liberatedliberated// minuteminute „ LookLook atat FunctionFunction inin TableTable 30.130.1 –– PhytasePhytase –– hydrolyzeshydrolyzes phytatephytate –– ProteaseProtease –– hydrolyzeshydrolyzes proteinsproteins –– LipaseLipase –– hydrolyzeshydrolyzes triglyceridestriglycerides (fat)(fat) –– CellulaseCellulase –– breaksbreaks downdown cellulosecellulose GuaranteesGuarantees--AssayAssay SpecificSpecific

„ UnitsUnits areare alsoalso assayassay specificspecific forfor aa particularparticular typetype ofof activityactivity „ FewFew standardstandard assaysassays „ AssayAssay cancan measuremeasure 11 ofof 22 thingsthings – Change in substrate „ Decrease in protein concentration „ mg casein hydrolyzed/ minute – Change in end product „ Increase in amino acid levels „ mg amino acids liberated/ minute GuaranteesGuarantees--AssayAssay SpecificSpecific (cont)(cont)

„ WhatWhat toto choose?choose? –– PickPick easiesteasiest toto measure,measure, generally,generally, breakdownbreakdown productproduct „ Protease – increase in amino acid concentrations „ Phytase – increase in amount of free phosphorus „ Amylase – increase in sugar levels GuaranteesGuarantees (cont)(cont)

„ AreAre therethere anyany Standards?Standards? –– AOACAOAC –– FoodFood ChemicalChemical CodexCodex ““UseUse byby”” DatesDates

„ NotNot requiredrequired „ WhyWhy includeinclude them?them? –– EnzymesEnzymes areare proteinsproteins andand activityactivity dependsdepends onon proteinprotein structurestructure –– ProteinProtein structurestructure degradesdegrades withwith timetime eveneven withwith preservativespreservatives „ InclusionInclusion couldcould preventprevent questionsquestions concerningconcerning guaranteesguarantees withwith anan outout--ofof-- datedate productproduct Mikazyme Contains a source of phytase, derived from Aspergillus oryzae Phytase increases the digestibility of phytin-bound phosphorus in swine and poultry diets Guaranteed Analysis Phytase (A. oryzae) min. 2500 FTU/ g One FTU is the amount of phytase which liberates 1 micromole of phosphorus per minute from sodium phytate at pH 5.5 and 37 C (under conditions of the assay)

Ingredients: Sodium sulfate, …, …, …, hydrogenated vegetable oil, starch, dried Aspergillus oryzae fermentation extract Directions for Use (mixing directions) For swine add 200-400 g/ton complete feed to provide 500 FTU/ kg feed For broilers add 200 – 400 g/ton complete feed to provide 500 FTU/ kg feed For layers add 100-200 g/ton of complete feed to provide 250 FTU/ kg feed Store at room temperature and in a dry place Use within 12 months of manufacture Warning (human health concern) May cause eye, skin, and respiratory irritation Inhalation may cause allergic reaction in sensitized individuals Manufactured by Mika Industries, 7519 Stand Place, Rockville, MD 20855 Net Wt 44 lb (20 kg) Manufactured 4/12/10 Mikazyme’s Swine Feed with Phytase For starter pigs weighing 11 to 44 lb Phytase increases the digestibility of phytin-bound phosphorus in swine and poultry diets Guaranteed Analysis Crude protein (min.) 20% Lysine (min.) 1.2% etc Phytase (A. oryzae) (min.) 500 FTU/ kg* Ingredients: Corn, …, ……, …, dried Aspergillus oryzae fermentation extract, cobalt carbonate, calcium iodate, sodium selenite Feeding Directions Feed as the complete ration to starter pigs weighing 11 to 44 lbs

*One FTU is the amount of phytase which liberates 1 micromole of phosphorus per minute from sodium phytate at pH 5.5 and 37 C (under conditions of the assay)

Manufactured by Blue Bird Feed Mill, City, State Zip Net Wt 50 lb (22.67 kg) Questions?Questions?

Images courtesy of www-news.uchicago.edu, www.wpi.edul, www.chemlin.net OtherOther QuestionsQuestions

„ SpirulinaSpirulina -- blueblue--greengreen algaealgae – Falls under limited use ingredients for species whose nutritional requirements are not well understood – Accepted for use in ornamental (aquarium) fish diets only – Not acceptable in other animal diets „ No data and information submitted to support these other uses – Testing is necessary, generally for microbiological, heavy metal, and chemical contaminants „ Acacia petals (Acacia nilotica) – Accepted for use in cockatiel bird diets only OtherOther ClaimsClaims

„ Must be based on scientific data in animal species of interest with the specific species „ Must relate back to the nutritional content of product – Nutritive value, aroma, flavor „ These statements are probably OK – Supports the Digestive System – Supports the Immune System „ These claims are not for a microbial product – Promotes Healthy Shiny Hair Coat „ Acceptable for a product with vitamin E or essential fatty acids – Promotes Vitality in older horses