血清总胆固醇的测定实验报告

TheexperimentprincipleThemeasurementoftheS-Cholisanimportantindicatorofthepreventionandtreatment，clinicaldiagnosisandnutritioninvestigationoftheASCD-atheroscleroticcardioasculardisease.theaverageperson'scontentsoftheS-Cholrangefrom2.6mmol/Lto6.5mmol/L(100~250mg/L)。Cholestenoneiscyclopentanoperhydrophenanthreneringderivative.notonlydoesitparticipateinthecomponentofthePlasmaprotein，butalsobetheessentialcompositionofthecellstructure，and becapableoftranslatingintoBR25gKayon，adrenocorticalhormoneandvitaminD，etc.cholestenone，whichiscalledbyajointnametotalcholesterol，existsasfreecholesterolandcholersterylester.themeasuringmethodof cholestenoneconsisitofchemicalcolorimetryandmethodsinenzymology.thisexperimentadoptthefirtone.cholesterolanditslipidderivativereactwitho-phthaldialdehyde，usingsulfuricacidasacatalyst，produceamaranthmaterialthatabsorbedlightat550nm。Hence，colorimetrycanbeusedforquantitativedetermination.whenthecontentoftheCholestenonewithinthelimitsof10..3mmol/L(400mg/1dL)，itwillpersentapositiverelationwithlightabsorption.Itisnotnecessarytocentrifugationinthismethod.Experimentmaterialandtheexperimentation1>Experimentmateria:o-phthaldialdehydereagent:Using50mgofto-phthaldialdehydeanddissolvingin50mlabsoluteethylalcohol,afterthatkeepincoldstoragefornolongerthanoneandahalfmonths.mixedacid:100mlofglacialaceticacidmixedwith100mlof concentratedsulfuricacidStandardstocksolutionof Cholestenone(1mg/ml):Using100mgoftheCholestenoneanddissolvingin100mlglacialaceticacid.Standardtreatmentfluidof Cholestenone(0.1mg/ml):dilutethestocksolution100timeswithglacialaceticacid  2>testsampleDilute0.1mlhumanserumwith glacialaceticacidinto4.00ml.3>experimentationTesttube1.5cm*15cm(*12);suctionpipet0.5ml(*5),10ml(*1),0.1ml(*1);SpectrophotometerOperationmethods1>makestandardcurveNumber9testtubes,afterthataddingreagentrefertothelistfollowed:numberStandardtreatmentfluidglacialaceticacido-phthaldialdehydereagentmixedacidamounttothecontentoftotalcholesterolinbloodserumunknown/mg％A550Afterthat，mixingthemuniformlyandblandly，andstewingthemfor10minutesatthetemporaryrangefrom20to37℃。Choose contentoftotalcholesterol(mg％) asx-coordinate,andA550asy-coordinatetomakethestandardcurve.2>sampletestNumber9testtubes,afterthataddingreagentrefertothelistfollowed:Number    1(Comparison)   2(sample)   3(sample)humanserumdiluted/mlo-phthaldialdehydereagent/mlglacialaceticacid/mlMixedacid /mlA550Afterthat，mixingthemuniformlyandblandly，andstewingthemfor10minutesatthetemporaryrangefrom20to37℃。Afterthatdeterminingthelightabsorbedat550nm。Then，checkthethecontentofthetotalcholesterolonthestandardcurve。ExperimentdataAccordingtothedatawegotfromthisexperiment，theaverageA550=【（样品①）A550（样品②）】/2=（0.2910.298）/2=0.295makestandardcurvewithorigin8.0withtheforthandfifthpointshieldedandfitting。Theequationofthefittingstraightlinefollowed：Y=0.00166x-0.00935 （r^2=0.99069）Substituteywitho.o295，wegotx=（0.009350.0295）/0.00166=183.343mg％Hence，thetotalcholesterolinthesampleis183.343mg％Resultanalysis1、thestandardcurveisnotperfectlinearlydependent。erroranalysisasfollowed：1>thereissomeresidualsinthepreparationofstandardsolutionwithdifferentconcentration2>thedifferentorderofaddingreagentintesttubesmakethereactantreactfordifferenttime.becauseofthat,extentofreactionhassomedifferenceswhenmeasuretheabsorbance.3>thesolutionmaybenotmixedperfectlybecausethethemixedacidisstickiness，whichcausesomeerrorintheabsorbancemeasure.4>Themostmajorerrorisfromthatthesamplesolutionisnotadequateforwashingthecuvettes.hencetheabsorbancewillbeinfluencedbylastsamplesolution.5>Wedidnotblendthesolutionagainafterwaterbath2、thesampletesthasalittlebitdiscrepancy,itmeansthatthesamplemixedhomogeneouslyReflectionQuestions1、Whatshouldbetakencareofintheoperationofthisexperiment?why?Answer:aftertheconcentrationgradientsolutionmadeup,itshouldbemixedhomogeneousandstewingthemfor10minutesinthetemperationrangefrom20to37℃。Becauseonlyinthatwaythereactionwillreactadequately，leadingtheaccurate of the absorbancewegot。2、lipidishardtobedissolvedinwater。Dispersingthemingwater，emulsionwillbeget。Whynormalpersonbloodplasmaandbloodserumarelimpidinsightwithsomuchlipid？Answer：thelipidinbloodplasmaandbloodserumisnotfree。mostofthemcombinewiththeproteininbloodplasmaandbloodserum，andtransportandexistinformoflipoprotein，andpartofthecholesterolhasthefunctionof“Sweeper”inbloodinordertokeepthebloodclean。