抗生素發(fā)酵生產(chǎn)工藝學(xué)

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(xì)胞的有機(jī)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(jī)制分類(ModesofActionofAntibiotics)

抑制細(xì)胞壁合成(CellWallSynthesis)

：青霉素抑制細(xì)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(jī)制分類(ModesofAction復(fù)制翻譯轉(zhuǎn)錄6整理課件復(fù)制翻譯轉(zhuǎn)錄6整理課件根據(jù)抗生素的化學(xué)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(xué)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(xì)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(zhǔn)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(kuò)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(bǔ)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(bǔ)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(bǔ)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(bǔ)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(yù)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