抗生素發(fā)酵生產(chǎn)工藝學

抗生素發(fā)酵生產(chǎn)技術(shù)AntibioticsFermentationProduction1整理課件抗生素發(fā)酵生產(chǎn)技術(shù)1整理課件概述生產(chǎn)前期研究生產(chǎn)菌改良生產(chǎn)工藝過程發(fā)酵條件的控制提取和精制IntroductionResearchinLabStrainimprovementProductionprocessingFermentationcontrolIsolation&purification抗生素發(fā)酵生產(chǎn)技術(shù)AntibioticsFermentationProduction2整理課件概述Introduction抗生素發(fā)酵生產(chǎn)技抗生素是生物在其生產(chǎn)活動過程中所產(chǎn)生，并能在低微濃度下有選擇性地抑制或殺滅其他微生物或腫瘤細胞的有機物。

Antibiotics

Anyorganicsubstancesproducedbyvariousorganismsthathavethepowerofarrestingthe

growthofothermicroorganismsorofdestroyingthem.1.概述Introduction3整理課件抗生素Antibiotics1.概述Introd歷史1929年，英國弗萊明發(fā)現(xiàn)青霉素二戰(zhàn)期間，大規(guī)模發(fā)酵生產(chǎn)世界上最早用于臨床的抗菌素History

PenicillinwasdiscoveredbyFlemingin1929.WorldWarII,fermentedinlarge-scale

Thefirstmajorantibiotictobecommercialized1.概述Introduction4整理課件歷史History1.概述Introducti根據(jù)抗生素的作用機制分類(ModesofActionofAntibiotics)

抑制細胞壁合成(CellWallSynthesis)

：青霉素抑制細胞膜功能(CellMembraneDamage)

：多烯類抗生素抑制蛋白質(zhì)合成(ProteinSynthesis)

：四環(huán)素抑制核酸合成(NucleicAcidSynthesis)

：絲裂霉素

分類Classification1.概述Introduction5整理課件根據(jù)抗生素的作用機制分類(ModesofAction復制翻譯轉(zhuǎn)錄6整理課件復制翻譯轉(zhuǎn)錄6整理課件根據(jù)抗生素的化學結(jié)構(gòu)分類β-內(nèi)酰胺類(β-lactam)：青霉素類(penicillins)

頭孢菌素類(cephalosporins)氨基糖苷類(aminoglycosides)：鏈霉素、慶大霉素大環(huán)內(nèi)酯類(macrolides)：紅霉素、麥迪加霉素四環(huán)類(tetracyclines)：四環(huán)素、土霉素多肽類(polymyxin)：多粘菌素

分類Classification1.概述Introduction7整理課件根據(jù)抗生素的化學結(jié)構(gòu)分類分類Classific

HNONSCH3CH3HCOO-H

-lactamringβ-內(nèi)酰胺環(huán)RnaturalpenicillinR=CH2-CO-8整理課件HNONSCH3CH3HCOO-H-lactamrinβ-內(nèi)酰胺環(huán)9整理課件β-內(nèi)酰胺環(huán)9整理課件兩性霉素Antibioticsare

socomplextheycanonlybesynthesized

inalivingsystem10整理課件兩性霉素Antibioticsaresocomplex產(chǎn)生菌的篩選抗菌性試驗提取、精制、鑒定毒性試驗藥理和臨床試驗ScreeningofastrainAntimicrobialtrialIsolation,purificationandidentificationToxicologicaltrial

Medicinal&clinicaltrial

2．生產(chǎn)前期研究ResearchinLab11整理課件產(chǎn)生菌的篩選Screeningofastrain2．

Agardiffusionassayagarplate瓊脂培養(yǎng)基lawnoftestbacteria測試菌苔filterpaperssoakedwithtestcompounds含藥物濾紙zonesofinhibition(nogrowth)

抑菌圈12整理課件Agardiffusionassayagarplat分析儀器Analyticalinstruments現(xiàn)代分析儀器：核磁共振毛細管電泳氣相色譜高效液相色譜質(zhì)譜Modernanalyticalinstruments:Nuclearmagneticresonance(NMR)Capillaryelectrophoresis

(CE)Gaschromatography

(GC)Highperformanceliquidchromatography

(HPLC)Massspectroscopy

(MS)

Automation

Sensitivity

Accuracy

自動靈敏準確13整理課件分析儀器現(xiàn)代分析儀器：Modernanalytica人工誘變法：紫外線、X-線、r-線照射亞硝基胍、亞硝酸、秋水仙素、氮芥等誘變劑處理ConventionalstrainselectionPhysicalmutageneg.UltravioletRadiationChemicalmutageneg.Nitrosoguanidine(NTG)Improvedstraincanbeselectedempirically3.抗生素生產(chǎn)菌改良StrainImprovement14整理課件人工誘變法：Conventionalstrainsele基因工程GeneticEngineering

3.抗生素生產(chǎn)菌改良StrainImprovementRecombinantDNATechnology（DNA重組技術(shù)）克隆15整理課件基因工程3.抗生素生產(chǎn)菌改良RecombinanStrainImprovementScale-upStrain出發(fā)菌株Agarplate平板技術(shù)PrimaryshakeflaskEvaluation搖瓶培養(yǎng)Secondaryshakeflaskevaluationdifferentmedia二級搖瓶培養(yǎng)各種培養(yǎng)基

Labfermentorevaluation

ProductionCondition小型發(fā)酵罐模擬生產(chǎn)條件MastercellbankWorkingcellbankInoculumDevelopment菌種擴大培養(yǎng)PilotplantEvaluation中試生產(chǎn)ProductionEvaluation生產(chǎn)60003003031Possiblerecycle16整理課件StrainImprovementScale-upStr17整理課件17整理課件Scale-up（比擬放大）modelsELGaddenJr<10dm3GoodproductGoodproductivity>200m3Sameproductivity?Limitations?模擬18整理課件Scale-up（比擬放大）modelsELGadden搖瓶發(fā)酵Shake-flaskfermentation10-50mlin125-500mlflaskBatchonlyLimitedcontrol:temperatureSlowmetabolizingcarbohydrateAmbientpressureBuffersneededtocontrolpHIn-processsamplingdifficultVolumedecreasebyevaporationSolidgrowthonsidewallsAntifoamnotneededAgitationlimited:shakerspeedNocontrolofdissolvedoxygen裝液量：10%(~250ml)間歇培養(yǎng)溫控受限制代謝碳水化合物慢常壓緩沖液控制pH在線取樣難容量減少瓶壁上結(jié)垢無需消泡攪拌受限制無法控制溶解氧19整理課件搖瓶發(fā)酵10-50mlin125-500m攪拌罐發(fā)酵Stirredtankfermentation10-100,000升容量間歇和補料培養(yǎng)連續(xù)控制：pH，溫度需要銨鹽或氨氣2大氣壓無需緩沖液在線取樣易補料,容量增加生長均勻一致需消泡多種攪拌控制溶解氧:通氣,加壓10-100,000litervesselsBatchandfeedpossibleContinuouscontrol:pH,temperatureAmmoniumsaltsorammonianeededPressureattwoatmospherespossibleNobuffersnecessaryIn-processsamplingeasyVolumeincreasebysugarfeedVeryuniformgrowthAntifoamoftenrequiredWidevarietyofimpellersandbafflesDissolvedoxygencontrolledbyaeration,pressure20整理課件攪拌罐發(fā)酵10-100,000升容量10-搖瓶發(fā)酵Shake-flask裝液量：10%(~250ml)間歇培養(yǎng)溫控受限制代謝碳水化合物慢常壓緩沖液控制pH在線取樣難容量減少瓶壁上結(jié)垢無需消泡攪拌受限制無法控制溶解氧10-100,000升容量間歇和補料培養(yǎng)連續(xù)控制：pH，溫度需要銨鹽或氨氣2大氣壓無需緩沖液在線取樣易補料,容量增加生長均勻一致需消泡多種攪拌控制溶解氧:通氣,加壓攪拌罐發(fā)酵Stirredtank21整理課件搖瓶發(fā)酵裝液量：10%(~250ml)10-10菌種

→種子制備

→發(fā)酵

→發(fā)酵液預處理→提取及精制→成品包裝Strain

→Preparationofseed

→Fermentation

→Treatmentofbroth→Isolation&purification→Packageofproduct4.抗生素生產(chǎn)工藝過程AntibioticsProductionProcessing22整理課件菌種→種子制備Strain→PreparatAschemeofthematerialflowinanantibioticproduction23整理課件AschemeofthematerialflowAschemeofthematerialflowinanantibioticproduction連消發(fā)酵罐緩沖罐種子罐24整理課件Aschemeofthematerialflow發(fā)酵生產(chǎn)原料

MaterialsforFermentationCarbonsource:Beet&canemolassesGlucose,lactose,starchEthanol,methanolGlycerolMaltosesyrup,cornsyrupCottonseedoil,peanutoil,soyoilLardoilMethyloleateTallow碳源:甜菜和甘蔗糖蜜葡萄糖，乳糖，淀粉乙醇，甲醇甘油麥芽糖漿，玉米糖漿綿籽油，花生油，豆油豬油油酸甲酯油脂25整理課件發(fā)酵生產(chǎn)原料MaterialsforF氮源:綿籽粉玉米粉和玉米浸出液大豆粉和大豆面花生粉菜籽粉牛血，酪蛋白水解液乳清魚粉酵母酵母膏肉粉和骨粉

Nitrogensource:CottonseedmealCorn

meal&steepliquorSoybeanmeal&flourPeanutmealRapeseedmealBeefblood,CaseinhydrolysateWheyFishmealWholeyeastYeastextractMeat&bonemeal

發(fā)酵生產(chǎn)原料

MaterialsforFermentation26整理課件氮源:Nitrogensource:發(fā)酵生產(chǎn)原料

培養(yǎng)基MediaforCellGrowth&FinalproductionComponentsRange%組成Glucose/sucrose3.0-5.0葡萄糖/蔗糖Starch淀粉3.0-5.0Cornsteepliquor3.0-5.0Calciumcarbonate0.5-1.0碳酸鈣Phosphate磷酸鹽0.1-0.5Ammoniumsulphate0.1-1.0

硫酸胺Urea尿素0.1-0.5Oil油0.1-0.5Fermentation發(fā)酵培養(yǎng)基

ComponentsRange%組成Glucose0.5-1.0Starch

0.5-5.0Cornsteepliquor5.0-8.0玉米浸提液Soyflour1.0-5.0Phosphate0.1-1.0Ammoniumsulphate0.5-1.0andfedperiodicallyOil1.0andfedcontinuouslyCornsyrupfedcontinuously

玉米糖漿連續(xù)加料Tankseed種子培養(yǎng)基27整理課件培養(yǎng)基MediaforCellGrowthStirredtankFermentation攪拌發(fā)酵28整理課件StirredtankFermentation5.發(fā)酵控制FermentationControl

SampleAnalysispHDOSugarAmmoniaPhosphateSulphateProductsPrecursorsContaminationPressureprobeLevelprobepHprobeTemp.probeDOprobeAntifoamAcid/BaseCoolingAir/agitationSugar/Oilfeed29整理課件5.發(fā)酵控制FermentationControScaleWeightSignalPumpSpeedAmplifierControllerS