1、BRITISH STANDARD BS ISO 11023:1999 Liquorice extracts (Glycyrrhiza glabra L.) Determination of glycyrrhizic acid content Method using high-performance liquid chromatography ICS 71.100.60; 71.040.40 NO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAWBS ISO 11023:1999 This British S
2、tandard, having been prepared under the direction of the Consumer Products and Services Sector Committee, was published under the authority of the Standards Committee and comes into effect on 15 September 2001 BSI 08-2001 ISBN 0 580 38314 8 National foreword This British Standard reproduces verbatim
3、 ISO 11023:1999 and implements it as the UK national standard. The UK participation in its preparation was entrusted to Technical Committee AW/54, Essential Oils, which has the responsibility to: A list of organizations represented on this committee can be obtained on request to its secretary. Cross
4、-references The British Standards which implement international or European publications referred to in this document may be found in the BSI Standards Catalogue under the section entitled “International Standards Correspondence Index”, or by using the “Find” facility of the BSI Standards Electronic
5、 Catalogue. A British Standard does not purport to include all the necessary provisions of a contract. Users of British Standards are responsible for their correct application. Compliance with a British Standard does not of itself confer immunity from legal obligations. aid enquirers to understand t
6、he text; present to the responsible international/European committee any enquiries on the interpretation, or proposals for change, and keep the UK interests informed; monitor related international and European developments and promulgate them in the UK. Summary of pages This document comprises a fro
7、nt cover, an inside front cover, the ISO title page, the ISO foreword page, pages 1 to 8, an inside back cover and a back cover. The BSI copyright date displayed in this document indicates when the document was last issued. Amendments issued since publication Amd. No. Date CommentsA Referencenumbe r
8、 ISO11023:1999 (E) INTERNATIONAL STANDARD ISO 11023 Firstedition 19991015 Liquoriceextracts( Glycyrrhizaglabra L.) Determinationofglycyrrhizicacid contentMethodusinghighperformance liquidchromatography Extraitsderglisse( Glycyrrhizaglabra L.)Dterminationdelateneuren acideglycyrrhiziqueMthodeparchrom
9、atographieliquidehaute performanceISO11023:1999(E) OSI OSI9991 rllAightsreserved.Unelssotherwisespeceifid,noparotfhtsipubcilatoinamybereproducedorutziliednianyofrmorbyan ymsnae,lecetrinoc .rehsilbupehtmorfgnitirwninoissimreptuohtiw,mliforcimdnagniypocotohpgnidulcni,lacinahcemro noitazidradnatSrofnoi
10、tazinagrOlanoitanretnI 65elatsopesaC 02evneG1121HC dnalreztiwS tenretnI hc.osiosi dnalreztiwSnidetnirP ii Foreword ISO(theInternationalOrganizationforStandardization)isaworldwidefederationofnationalstandardsbodies(ISO memberbodies).TheworkofpreparingInternationalStandardsisnormallycarriedoutthroughI
11、SOtechnical committees.Eachmemberbodyinterestedinasubjectforwhichatechnicalcommitteehasbeenestablishedhas therighttoberepresentedonthatcommittee.Internationalorganizations,governmentalandnongovernmental,in liaisonwithISO,alsotakepartinthework.ISOcollaboratescloselywiththeInternationalElectrotechnica
12、l Commission(IEC)onallmattersofelectrotechnicalstandardization. InternationalStandardsaredraftedinaccordancewiththerulesgivenintheISO/IECDirectives,Part3. DraftInternationalStandardsadoptedbythetechnicalcommitteesarecirculatedtothememberbodiesforvoting. PublicationasanInternationalStandardrequiresap
13、provalbyatleast75%ofthememberbodiescastingavote. InternationalStandardISO11023waspreparedbyTechnicalCommitteeISO/TC54, Essentialoils . AnnexAofthisInternationalStandardisforinformationonly.INTERNATIONALSTANDARD OSI ISO11023:1999(E) 1 Liquoriceextracts( Glycyrrhizaglabra L.)Determinationof glycyrrhiz
14、icacidcontentMethodusinghighperformanceliquid chromatography 1Scope ThisInternationalStandarddescribesamethodfordeterminingtheglycyrrhizicacidcontentofliquoriceextract (Glycyrrhizaglabra L.)byhighperformanceliquidchromatography. Themethodisnotapplicabletoraworgroundliquoriceroot. 2Principle Thesampl
15、eandstandardsolutionsareprepared,thentheglycyrrhizicacidcontentisdeterminedbyhigh performanceliquidchromatographyusingthemethoddescribedinthisInternationalStandard. 3Reagents Useonlyreagentsofrecognizedanalyticalgrade,unlessotherwisespecified. 3.1 Water,HPLCgrade. 3.2 Referencesubstance ,monoammonia
16、calglycyrrhizate(GMA). Ifareferencematerialofguaranteedpurityisnotavailable,itisrecommendedthattheusersofthisInternational Standardcometoanagreementbetweentheinterestedpartiesonthepurityofthereferencesubstance. 3.3Acetonitrile ,HPLCgrade. 3.4 Aceticacid ,analyticalgrade. 3.5 Elutionsolvent(mobilepha
17、se), composedofthefollowing: 38volumesacetonitrile(3.3), 61volumeswater(3.1), 1volumeaceticacid(3.4). Usingthemeasuringcylinder(4.3),preparetheelutionsolventasfollows. Mix1volumeofaceticacidwith61volumesofwater,thenfilterthemixturethroughafilterforaqueoussolvents (4.5). Filter38volumesofacetonitrile
18、throughafilterfororganicsolvents(4.4). Addthefilteredacetonitriletothefilteredwater/aceticacidmixture.Mix,thendegastheelutionsolventintheultra soundcell(4.8.3)oranysuitablesystem. Donotkeepthissolventmorethan48hatambienttemperature.ISO11023:1999(E) OSI 2 4Apparatus Usuallaboratoryapparatusand,inpart
19、icular,thefollowing. 4.1 Pipettes,ofcapacities5mland10ml. 4.2 Volumetricflasks ,ofcapacities50mland100ml. 4.3 Measuringcylinder . 4.4 Filterfororganicsolvents ,0,5 m mporesize. 4.5 Filterforaqueoussolvents ,0,45 m mporesize. 4.6 Oven,capableofbeingmaintainedat105C+ 2C. 4.7 Analyticalbalance ,capable
20、ofweighingtothenearest0,0001g. 4.8 Separationsystem ,asfollows. 4.8.1 Chromatograph,highperformanceliquidphase. 4.8.2 Pumpingsystem ,forobtainingandmaintainingaconstantorprogrammedhighpressureflow. 4.8.3 Solventdegassingsystem ,suchasanultrasoundcelloranysuitablesystem. 4.8.4 Ultravioletdetectionsys
21、tem ,adjustabletoawavelengthof254nm. 4.9 Recorder orintegrator ,ofcompatibleperformancewithalltheapparatus. 4.10 Column,asfollows: material:stainlesssteelorglass; length:10cmto25cm; internaldiameter:0,4cmto0,5cm; stationaryphase:bondedphasesilicawithoctadecylC18derivedfunctionalgroup,maximumparticle
22、size 5 mm; columnefficiency:thenumberrecommendedoftheoreticalplatesis7000to10000. 4.11 Precolumn ,asfollows: material:stainlesssteel; length:10mmor25mm; internaldiameter:2mmor4mm; stationaryphase:bondedphasesilicawithoctadecylC18derivedfunctionalgroupOSI ISO11023:1999(E) 3 5Procedure 5.1 Preparation
23、ofstandardsolutions 5.1.1 Preparationofthe0,5mg/mlsolution Ina100mlvolumetricflask(4.2)weigh,tothenearest10 4 g,50mgofmonoammoniacalglycyrrhizate(3.2). Addtheelutionsolvent(3.5)anddissolvethemonoammoniacalglycyrrhizate(3.2).Makeuptothemarkwitha furtherquantityofelutionsolvent. Filterifnecessarythrou
24、ghtheorganicsolventfilter(4.4). Itisessentialtoprepareanewsolutioneveryday. 5.1.2 Preparationofdilutesolutionscontaining0,05mg/ml,0,075mg/mland0,1mg/ml 5.1.2.1 0,05mg/mlstandardsolution Usingapipette(4.1),transfer5mlofthe0,5mg/mlstandardsolution(5.1.1)toa50mlvolumetricflask(4.2).Make uptothemarkwith
25、theelutionsolvent(3.5). 5.1.2.2 0,075mg/mlstandardsolution Usingapipette(4.1),transfer15mlofthe0,5mg/mlstandardsolution(5.1.1)toa100mlvolumetricflask(4.2). Makeuptothemarkwiththeelutionsolvent(3.5). 5.1.2.3 0,1mg/mlstandardsolution Usingapipette(4.1),tranfer10mlofthe0,5mg/mlstandardsolution(5.1.1)to
26、a50mlvolumetricflask(4.2).Make uptothemarkwiththeelutionsolvent(3.5). Itisessentialtoprepareanewsolutioneveryday. 5.2 Preparationofthesolutionforanalysis 5.2.1 Preparethesolutionforanalysissuchthattheareaoftheglycyrrhizicacidpeakobtainedisbetweenthe peaksfortheothertwostandardsolutions,onthebasisoft
27、herecommendedmassesandvolumesspecifiedin Table1,accordingtotheassumedglycyrrhizicacidcontentofthesample. Table1Examplesofquantitiesandvolumestobeusedaccordingtotheassumedglycyrrhizicacid contentoftheliquoriceextracts Assumed glycyrrhizicacid content Massoftest portion Volumeofflask Volumeofpipetted
28、solution Volumeofflask V 1 V 2 V 3 %m gm lm lm l 3to5 1 50 5 50 5to10 1 100 5 50 10to15 0,50 100 5 50 20to25 0,25 100 5 50ISO11023:1999(E) OSI 4 5.2.2 Weigh,tothenearest10 -4 g,atestportionofmass m 0 intoavolumetricflaskofvolume V 1 ml. Addanappropriatequantityofelutionsolvent(3.5)todissolvethesampl
29、e.Thenmakeuptothemarkwithelution solvent. Usingapipette,transferavolume( V 2 )ofthissolutiontoavolumetricflaskofvolume V 3 ml. Addanappropriatequantityofelutionsolvent(3.5),mixandmakeuptothemarkwithelutionsolvent. Filterthislastsolutionthroughthefilterfororganicsolvents(4.3)priortoinjecting. 5.3 Det
30、erminationofthewaterandvolatilemattercontentofthesample Weigh,tothenearest10 -4 g,atestportion( m 0 )ofexactly2gandplaceinacalibratedwatchglass. Placeitintheoven(4.6)setat105Cfor24h,untilaconstantmassisreached.Coolthenweightheresidue( m 1 ). Thewaterandvolatilemattercontentofthesample, w,expressedas
31、apercentagebymass,isequalto: w mm m = - () % 01 0 100 where m 0 isthemassofthetestportion,ingrams; m 1 isthemassoftheresidueobtained,ingrams. 6Determination 6.1 Inject10 m lofsolution5.1.2.1. 6.2 Inject10 m lofsolution5.1.2.2. 6.3 Inject10 m lofsolution5.1.2.3. 6.4 Inject10 m lofthesolutionforanalys
32、is(5.2.2). 6.5 Carryouttheoperationsdescribedin6.1to6.4twice. NOTE Iftheinjectionanddetectionsystemsallowthis,itispossibletoinjectquantitiesfrom5 m lto10 m l. 6.6 Measuretheareaoftheglycyrrhizicacidpeakonthechromatogramsobtainedforthestandardsolutions(see annexA). 6.7 Plotacurveofglycyrrhizicacidpea
33、kareaagainsttheconcentrationofmonoammoniacalglycyrrhizateinthe standardsolutions. Acalibrationlinepassingthroughtheoriginisobtained. 6.8 Usingthechromatogramforthesamplesolution,measuretheareaoftheglycyrrhizicacidpeak. Fromthecalibrationline,readthemonoammoniacalglycyrrhizateconcentration, c,inmilli
34、gramspermillilitre.OSI ISO11023:1999(E) 5 7 Expressionofresults Theglycyrrhizicacidcontent,w G , expressedasapercentagebymassondrymatter,iscalculatedusingthe formula: w cVV p mV w G () = - 13 02 100 822 100 839 where m 0 isthemassofthetestportion,ingrams; c isthemeanvalueoftheconcentrationsobtained,
35、inmilligramspermillilitre,ofmonoammoniacal glycyrrhizatereadfromthecalibrationline; p isthepurityofthereferencesubstance,inpercent; w isthewaterandvolatilemattercontentofthesampleforanalysis,asapercentagebymass,determinedin 5.3; 822 isthemolarmassofglycyrrhizicacid; 839 isthemolarmassofmonoammoniaca
36、lglycyrrhizate; V 1 , V 2 , V 3 havethesamemeaningsasindicatedinTable1. 8Test report Thetestreportshallspecify: allinformationnecessaryforthecompleteidentificationofthesample; thesamplingmethodused,ifknown; thetestmethodused,withreferencetothisInternationalStandard; alloperatingdetailsnotspecifiedin
37、thisInternationalStandard,orregardedasoptional,togetherwithdetailsof anyincidentswhichmayhaveinfluencedthetestresult(s); thetestresult(s)obtained;or iftherepeatabilityhasbeenchecked,thefinalquotedresultobtained.ISO11023:1999(E) OSI 6 AnnexA (informative) Examplesofchromatograms FigureA.1Chromatogram
38、ofaliquoriceextractOSI ISO11023:1999(E) 7 FigureA.2ChromatogramofthereferencesubstanceISO11023:1999(E) OSI 8 Bibliography 1 ISO8432, EssentialoilsAnalysisbyhighperformanceliquidchromatographyGeneralmethod.BS ISO 11023:1999 BSI 389 Chiswick High Road London W4 4AL BSI British Standards Institution BS
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