550 likes | 1.14k Views
PENICILLINS. Chapter 19. INTRODUCTION TO PENICILLINS. Antibacterial agents which inhibit bacterial cell wall synthesis Discovered by Fleming from a fungal colony (1928) Shown to be non toxic and antibacterial Isolated and purified by Florey and Chain (1938)
E N D
PENICILLINS Chapter 19
INTRODUCTION TO PENICILLINS • Antibacterial agents which inhibit bacterial cell wall synthesis • Discovered by Fleming from a fungal colony (1928) • Shown to be non toxic and antibacterial • Isolated and purified by Florey and Chain (1938) • First successful clinical trial (1941) • Produced by large scale fermentation (1944) • Structure established by X-ray crystallography (1945) • Full synthesis developed by Sheehan (1957) • Isolation of 6-APA by Beechams (1958-60) • - development of semi-synthetic penicillins • Discovery of clavulanic acid and b-lactamase inhibitors
R = Benzyl penicillin (Pen G) 6-Aminopenicillanic acid (6-APA) 6-Aminopenicillanic acid (6-APA) 6-Aminopenicillanic acid (6-APA) R = Acyl side chain Acyl side chain Acyl side chain Acyl side chain Thiazolidine ring Thiazolidine ring Phenoxymethyl penicillin (Pen V) b-Lactam ring Penicillin G present in corn steep liquor Penicillin V (first orally active penicillin) STRUCTURE Side chain varies depending on carboxylic acid present in fermentation medium
Shape of Penicillin G Folded ‘envelope’ shape
CYS CYS VAL Biosynthesis of Penicillins
Properties of Penicillin G • Active vs. Gram +ve bacilli and some Gram -vecocci • Non toxic • Limited range of activity • Not orally active - must be injected • Sensitive to b-lactamases • (enzymes which hydrolyse the b-lactam ring) • Some patients are allergic • Inactive vs. Staphylococci Drug Development • Aims • To increase chemical stability for oral administration • To increase resistance to b-lactamases • To increase the range of activity
SAR • Conclusions • Amide and carboxylic acid are involved in binding • Carboxylic acid binds as the carboxylate ion • Mechanism of action involves the b-lactam ring • Activity related to b-lactam ring strain (subject to stability factors) • Bicyclic system increases b-lactam ring strain • Not much variation in structure is possible • Variations are limited to the side chain (R)
Covalent bond formed to transpeptidase enzyme • Irreversible inhibition Mechanism of action • Penicillins inhibit a bacterial enzyme called the transpeptidase enzyme which is involved in the synthesis of the bacterial cell wall • The b-lactam ring is involved in the mechanism of inhibition • Penicillin becomes covalently linked to the enzyme’s active site leading to irreversible inhibition
NAM NAM NAM NAM NAM NAM NAM NAM NAM NAG NAG NAG NAG NAG NAG L-Ala L-Ala L-Ala L-Ala L-Ala L-Ala L-Ala L-Ala L-Ala D-Glu D-Glu D-Glu D-Glu D-Glu D-Glu D-Glu D-Glu D-Glu L-Lys L-Lys L-Lys L-Lys L-Lys L-Lys L-Lys L-Lys L-Lys Bond formation inhibited by penicillin Mechanism of action - bacterial cell wall synthesis
Cross linking Mechanism of action - bacterial cell wall synthesis
Mechanism of action - bacterial cell wall synthesis • Penicillin inhibits final crosslinking stage of cell wall synthesis • It reacts with the transpeptidase enzyme to form an irreversible covalent bond • Inhibition of transpeptidase leads to a weakened cell wall • Cells swell due to water entering the cell, then burst (lysis) • Penicillin possibly acts as an analogue of the L-Ala-g-D-Glu portion of the pentapeptide chain. However, the carboxylate group that is essential to penicillin activity is not present in this portion
Normal Mechanism Mechanism of action - bacterial cell wall synthesis Alternative theory- Pencillin mimics D-Ala-D-Ala.
Mechanism inhibited by penicillin Mechanism of action - bacterial cell wall synthesis Alternative theory- Pencillin mimics D-Ala-D-Ala.
Penicillin Acyl-D-Ala-D-Ala But 6-methylpenicillin is inactive despite being a closer analogue Mechanism of action - bacterial cell wall synthesis Penicillin can be seen to mimic acyl-D-Ala-D-Ala
D - A l a - D - A l a Blocked H O Acylation O O H Active Site O H Mechanism of action - bacterial cell wall synthesis Penicillin may act as an ‘umbrella’ inhibitor
Gram +ve and Gram -ve Cell Walls • Penicillins have to cross the bacterial cell wall in order to reach their target enzyme • Cell walls are porous and are not a barrier • The cell walls of Gram +ve bacteria are thicker than Gram -ve cell walls, but the former are more susceptible to penicillins
Thick porous cell wall • Thick cell wall • No outer membrane • More susceptible to penicillins Cell membrane Cell Gram +ve and Gram -ve Cell Walls Gram +ve bacteria
Hydrophobic barrier Outer membrane Porin Lactamase L L L enzymes Periplasmic space Thin cell wall L Cell membrane Cell • Thin cell wall • Hydrophobic outer membrane • More resistant to penicillins Gram +ve and Gram -ve Cell Walls Gram -ve bacteria
Resistance to Penicillins • Factors • Gram -ve bacteria have a lipopolysaccharide outer membrane preventing access to the cell wall • Penicillins can only cross via porins in the outer membrane • Porins allow small hydrophilic molecules such as zwitterions to cross • High levels of transpeptidase enzyme may be present • The transpeptidase enzyme may have a low affinity for penicillins • (e.g. PBP 2a for S. aureus) • Presence of b-lactamases • Concentration of b-lactamases in periplasmic space • Mutations • Transfer of b-lactamases between strains • Efflux mechanisms pumping penicillin out of periplasmic space
Penicillin Analogues - Preparation • 1) By fermentation • Vary the carboxylic acid in the fermentation medium • Limited to unbranched acids at the a-position i.e. RCH2CO2H • Tedious and slow • 2) By total synthesis • Only 1% overall yield • Impractical • 3) By semi-synthetic procedures • Use a naturally occurring structure as the starting material for analogue synthesis
Penicillin acylase or chemical hydrolysis Fermentation Semi-synthetic penicillins Penicillin Analogues - Preparation
Penicillin Analogues - Preparation Problem - How does one hydrolyse the side chain by chemical means in presence of a labileb-lactam ring? Answer - Activate the side chain first to make it more reactive Note - Reaction with PCl5 requires the involvement of a lone pair of electrons from nitrogen. Not possible for the b-lactam nitrogen.
Problems with Penicillin G • It is sensitive to stomach acids • It is sensitive to b-lactamases - enzymes which hydrolyse the b-lactam ring • It has a limited range of activity
Acid or enzyme Relieves ring strain Problem 1 - Acid Sensitivity Reasons for sensitivity 1) Ring strain
Tertiary amide Unreactive b-Lactam Folded ring system Impossibly strained Problem 1 - Sensitivity Reasons for sensitivity 2) Reactive b-lactam carbonyl group Does not behave like a tertiary amide X • Interaction of nitrogen’s lone pair with the carbonyl group is not possible • Results in a reactive carbonyl group
Further reactions Problem 1 - Sensitivity Reasons for sensitivity Acyl side chain Neighboring group participation in the hydrolysis mechanism
Decreases nucleophilicity Problem 1 - Sensitivity Conclusions • The b-lactam ring is essential for activity and must be retained • Cannot tackle factors 1 and 2 • Can only tackle factor 3 Strategy Vary the acyl side group (R) to make it electron-withdrawing to decrease the nucleophilicity of the carbonyl oxygen
electronegative oxygen Penicillin V (orally active) • Better acid stability and orally active • But sensitive to b-lactamases • Slightly less active than penicillin G • Allergy problems with some patients Very successful semi-synthetic penicillins e.g. ampicillin, oxacillin Problem 1 - Sensitivity Examples
b-Lactamase Problem 2 - Sensitivity to b-Lactamases b-Lactamases • Enzymes that inactivate penicillins by opening b-lactam rings • Allow bacteria to be resistant to penicillin • Transferable between bacterial strains (i.e. bacteria can acquire resistance) • Important with respect to Staphylococcus aureus infections in hospitals • 80% Staph. infections in hospitals were resistant to penicillin and other antibacterial agents by 1960 • Mechanism of action for lactamases is identical to the mechanism of inhibition for the target enzyme • But product is removed efficiently from the lactamase active site
Bulky group Enzyme Problem 2 - Sensitivity to b-Lactamases Strategy • Use of steric shields • Block access of penicillin to the active site of the enzyme by introducing bulky groups to the side chain • Size of shield is crucial to inhibit reaction of penicillins with b-lactamases, but not with the target transpeptidase enzyme
ortho groups important Problem 2 - Sensitivity to b-Lactamases Examples - Methicillin (Beechams - 1960) • Methoxy groups block access to b-lactamases but not to transpeptidases • Binds less readily to transpeptidases compared to penicillin G • Lower activity compared to Pen G against Pen G sensitive bacteria • Poor activity vs. some streptococci • Inactive vs. Gram -ve bacteria • Poor range of activity • Active against some penicillin G resistant strains (e.g. Staphylococcus) • Acid sensitive since there is no electron-withdrawing group • Orally inactive and must be injected
Problem 2 - Sensitivity to b-Lactamases Examples - Oxacillin Oxacillin R = R' = H Cloxacillin R = Cl, R' = H Flucloxacillin R = Cl, R' = F • Orally active and acid resistant • Resistant to b-lactamases • Active vs. Staphylococcus aureus • Less active than other penicillins • Inactive vs. Gram -ve bacteria • Nature of R & R’ influences absorption and plasma protein binding • Cloxacillin better absorbed than oxacillin • Flucloxacillin less bound to plasma protein, leading to higher levels of free drug
Problem 3 - Range of Activity Factors 1) Cell wall may have a coat preventing access to the cell 2) Excess transpeptidase enzyme may be present 3) Resistant transpeptidase enzyme (modified structure) 4) Presence of b-lactamases 5) Transfer of b-lactamases between strains 6) Efflux mechanisms • Strategy • The number of factors involved make a single strategy impossible • Use trial and error by varying R groups on the side chain • Successful in producing broad spectrum antibiotics • Results demonstrate general rules for broad spectrum activity.
Problem 3 - Range of Activity Results of varying R in Pen G 1) Hydrophobic side chains result in high activity vs. Gram +ve bacteria and poor activity vs. Gram -ve bacteria 2) Increasing hydrophobicity has little effect on Gram +ve activity but lowers Gram -ve activity 3) Increasing hydrophilic character has little effect on Gram +ve activity but increases Gram -ve activity 4) Hydrophilic groups at the a-position (e.g. NH2, OH, CO2H) increase activity vs Gram -ve bacteria
Problem 3 - Range of Activity Examples of Broad Spectrum Penicillins Class 1 - NH2 at the a-position Ampicillin and amoxicillin (Beechams, 1964) Ampicillin (Penbritin) 2nd most used penicillin Amoxicillin (Amoxil)
Problem 3 - Range of Activity Examples of Broad Spectrum Penicillins • Active vs Gram +ve bacteria and Gram -ve bacteria which do not produce b-lactamases • Acid resistant and orally active • Non toxic • Sensitive to b-lactamases • Increased polarity due to extra amino group • Poor absorption through the gut wall • Disruption of gut flora leading to diarrhea • Inactive vs. Pseudomonas aeruginosa Properties
Problem 3 - Range of Activity Prodrugs of Ampicillin (Leo Pharmaceuticals - 1969) • Properties • Increased cell membrane permeability • Polar carboxylic acid group is masked by the ester • Ester is metabolised in the body by esterases to give the free drug
PEN PEN H C OH O C O O O O PEN C C O O C C H H C M e 2 2 3 O Formaldehyde Problem 3 - Range of Activity Mechanism of prodrug activation • Extended ester is less shielded by the penicillin nucleus • Hydrolysed product is chemically unstable and degrades • Methyl ester of ampicillin is not hydrolysed in the body • Bulky penicillin nucleus acts as a steric shield for methyl ester
Problem 3 - Range of Activity Examples of broad spectrum penicillins Class 2 - CO2H at the a-position (carboxypenicillins) Examples R = H Carbenicillin R = Ph Carfecillin • Carfecillin = prodrug for carbenicillin • Active over a wider range of Gram -ve bacteria than ampicillin • Active vs. Pseudomonas aeruginosa • Resistant to most b-lactamases • Less active vs Gram +ve bacteria (note the hydrophilic group) • Acid sensitive and must be injected • Stereochemistry at the a-position is important • CO2H at the a-position is ionised at blood pH
TICARCILLIN Problem 3 - Range of Activity Examples of broad spectrum penicillins Class 2 - CO2H at the a-position (carboxypenicillins) Examples • Administered by injection • Identical antibacterial spectrum to carbenicillin • Smaller doses required compared to carbenicillin • More effective against P. aeruginosa • Fewer side effects • Can be administered with clavulanic acid
Class 3 - Urea group at the a-position (ureidopenicillins) Examples Azlocillin Mezlocillin Piperacillin Problem 3 - Range of Activity Examples of broad spectrum penicillins • Administered by injection • Generally more active than carboxypenicillins vs. streptococci and Haemophilus species • Generally have similar activity vs Gram -ve aerobic rods • Generally more active vs other Gram -ve bacteria • Azlocillin is effective vs P. aeruginosa • Piperacillin can be administered alongside tazobactam
玻璃钢生产厂家恩施玻璃钢雕塑设计方案抚州校园玻璃钢雕塑安装玻璃钢壁画雕塑厂家像玻璃钢雕塑的超市摆台周口玻璃钢座椅雕塑温州玻璃钢卡通雕塑销售玻璃钢麒麟雕塑价格湘西玻璃钢雕塑价格多少钱漳州玻璃钢花盆价格沧州校园玻璃钢雕塑安装玻璃钢雕塑行业标准宣威市玻璃钢雕塑订制南阳景观园林校园玻璃钢雕塑阜康玻璃钢头像雕塑贵港商场美陈雕塑厂家中山雕塑玻璃钢吉安佛像玻璃钢雕塑多少钱杭州小区玻璃钢雕塑厂家信阳附近校园玻璃钢景观雕塑上海户外商场美陈制造房山玻璃钢马雕塑温州玻璃钢雕塑价格行情福建商场主题创意商业美陈传播扬州玻璃钢人物雕塑商家邯郸玻璃钢景观雕塑制作厂家曲阳玻璃钢丹顶鹤雕塑漯河玻璃钢雕塑加工厂入户花园玻璃钢花盆装饰商场市集特卖美陈广安玻璃钢商场美陈香港通过《维护国家安全条例》两大学生合买彩票中奖一人不认账让美丽中国“从细节出发”19岁小伙救下5人后溺亡 多方发声单亲妈妈陷入热恋 14岁儿子报警汪小菲曝离婚始末遭遇山火的松茸之乡雅江山火三名扑火人员牺牲系谣言何赛飞追着代拍打萧美琴窜访捷克 外交部回应卫健委通报少年有偿捐血浆16次猝死手机成瘾是影响睡眠质量重要因素高校汽车撞人致3死16伤 司机系学生315晚会后胖东来又人满为患了小米汽车超级工厂正式揭幕中国拥有亿元资产的家庭达13.3万户周杰伦一审败诉网易男孩8年未见母亲被告知被遗忘许家印被限制高消费饲养员用铁锨驱打大熊猫被辞退男子被猫抓伤后确诊“猫抓病”特朗普无法缴纳4.54亿美元罚金倪萍分享减重40斤方法联合利华开始重组张家界的山上“长”满了韩国人?张立群任西安交通大学校长杨倩无缘巴黎奥运“重生之我在北大当嫡校长”黑马情侣提车了专访95后高颜值猪保姆考生莫言也上北大硕士复试名单了网友洛杉矶偶遇贾玲专家建议不必谈骨泥色变沉迷短剧的人就像掉进了杀猪盘奥巴马现身唐宁街 黑色着装引猜测七年后宇文玥被薅头发捞上岸事业单位女子向同事水杯投不明物质凯特王妃现身!外出购物视频曝光河南驻马店通报西平中学跳楼事件王树国卸任西安交大校长 师生送别恒大被罚41.75亿到底怎么缴男子被流浪猫绊倒 投喂者赔24万房客欠租失踪 房东直发愁西双版纳热带植物园回应蜉蝣大爆发钱人豪晒法院裁定实锤抄袭外国人感慨凌晨的中国很安全胖东来员工每周单休无小长假白宫:哈马斯三号人物被杀测试车高速逃费 小米:已补缴老人退休金被冒领16年 金额超20万