重組人白細胞介素-6中試純化工藝研究.pdf

1、天津大學碩士學位論文重組人白細胞介素6中試純化工藝研究姓名：章剛申請學位級別：碩士專業(yè)：生物化工指導教師：甘一如2000.11.1AbstractInthisthesisthepurificationofrecombinanthumaninterlrukin6(rhlL一6)forthepurposeofcommercializationwasstudiedRhlL一6wasexpressedinthegeneticengineerin

2、gEcolicells(providedbyBeijingMedicineUniversity)throughfermentationAfterdisruptionoftheEcoliceilsandseveralseparationandpurificationsteps，theinclusionbodywasprepared，whichWBSthematerialofthestudyDenaturationsystemusedind

3、issolvingofinclusionbodyWasstudiedandtwosystemidealismweregotAfterdissolvedofinclusionbodytherhlL一6wasrefoldedbydilutingthedenaturedsolutionandthusgotrenaturedsolufionWascomeintonextpurificationstepsForthefirststepofpuri

4、fication，severalmethodssuchasultrafiltration，ionexchangechromatographygelfiltrationandhydrophobicinteractionchromatographywerestudied，andionexchangechromatographywasfinallydecidedasthefirststepThethree—steppurificationpr

5、ocesswasfinallyadopted，whichWassequentiallycomposedofDEAEanionexchange，QSepharoseFastflowanionexchange，andPhenylHighsubstitutehydrophobicinteractionchromatographyTherenaturedsolutionWasloadedontheDEAEanionexchangecolumnw

6、hichWasdesignedtemovemostpotentialpyrogen，andtheflowthroughWascollectedAfteritspHWasregulated，theflowRhroughWasloadedontheQSepharoseFFanionexchangecolumnandthecomponentwhosepurityWaSabove85％wascollectedandcomeintothenext

7、stepInthisstep，theyieldofrhIL6Wasabout17％ThetargetsampleinstepQSepharoseFFafterprocessed，WaSloadedonthePhenylFF(HS)hydrophobicinteractionChromatographycolumn，inthisstep，impuritieswerefartherremovedandthefinalproductWaSgo

8、aen，andtheyieldofrhlL一6wasabout65％Throughthethreepurificationsteps，thepurityofthefinalproductWasabove95％，thebioactivityabove1xloSu／mg，thedeterminationofpyrogenshownnegative，andthetotalyieldofrhlL6wa3about11％Themethodincr

9、easingtheyieldofrhlL6wasstudiedTheflowthroughinthesecondchromatographyafterconcentratedbyultrafiltration，denatured，andrefoldedwasreenteredtheprocessagainTheyieldofrML一6WaSincreasedby2～3％comparedwiththenonrecycledprocessK