ANTIBIOTIK

ANTIBIOTIK

History

Paul Ehrlich:

Trypanred against trypanosomes (1904), Compound 606 (Salvarsan) (yellow), the first antibiotic (1910) against syphilis. Coined terms "magic bullet”, "chemotherapy”

Alexander Fleming: 1928

Penicillin (Penicillium notatum)

Gerhard Domagk: 1935

sulfa drugs, prontosil,sulfanilamide, isoniazid

Selman Waksman and Albert Schatz: 1943

streptomycin–first aminoglycoside(Streptomyces) against TB, coined the term “antibiotics”

Chloramphenicol, 1947, from Streptomycesvenezuelae

Tetracycline: 1948, from Streptomyces

Principle

Antibiotic should betoxic to the (pathogen) microbe and possibly harmless to the host (human being)

(Paul Erlinch)

Industrial Fermentors

125-250m3

Conditions in the fermenter are carefully monitored to regulate cell growth.

Fermenter and all pipe work must be sterile before fermentation begins

This is usually achieved by flushing the whole system with superheated steam before the production begins.

Process if frequently aerobic so fermentor has to be well aerated.

The aeration will be sufficient to mix many cultures

If the culture is thick or sticky, additional stirring is required by a motor driven paddle called an impeller.

While initially the culture may need warming to start of the process –once it has started a cooling systemis vital.

Cooling is achieved by either a water jacket or cooling coils inside the fermenter.

INDUSTRIAL PRODUCTION OF ANTIBIOTIC-PENICILLIN

The industrial production of penicillin was broadly classified in to two processes namely,

1.       UPSTREAM PROCESSING

INOCULUM PREPARATION

The medium is designed to provide the organism with all the nutrients that it requires.

Inoculation method-submerged technique

Spores -major source of inoculum

RAW MATERIALS

•CARBON SOURCES:Lactose acts as a very satisfactory carbon compound, provided that is used in a concentration of 6%. Others such as glucose & sucrose may be used.NITROGEN SOURCES:

•Corn steep liquor (CSL)

•Ammonium sulphate and ammonium acetate can be used as nitrogenous sources.MINERAL SOURCES:Elements namely potassium, phosphorus, magnesium, sulphur, zinc and copper are essential for penicillin production. Some of these are applied by corn steep liquor.

•Calcium can be added in the form of chalk to counter the natural acidity of CSL

•PAA-precursor

2.       DOWNSTREAM PROCESSING

The extraction and purification of a biotechnological product from fermentation is referred to as downstream processing.

FERMENTATION PROCESS

The medium is inoculated with a suspension of conidia of Penicillium chrysogenum.

The medium is constantly aerated and agitated, and the mould grows throughout as pellets.

After about seven days, growth is complete, the pH rises to 8.0 or above, and penicillin production ceases

STAGES IN DOWNSTREAM PROCESSING

1.       Removal of cells

The first step in product recovery is the separation of whole cells and other insoluble ingredients from the culture broth by technique such as filtration and centrifugation.

ISOLATION OF BENZYL PENICILLIN

The PH is adjusted to 2-2.5 with the help of phosphoric or sulphuric acids.

In aqueous solution at low PH values there is a partition coefficient in favor of certain organic solvents such as butyl acetate.

This step has to be carried out quickly for penicillin is very unstable at low PH values.

Antibiotic is then extracted back into an aqueous buffer at a PH of 7.5, the partition coefficient now being strongly in favor of the aqueous phase. The resulting aqueous solution is again acidified & re-extracted with an organic solvent.

These shifts between the water and solvent help in the purification of penicillin.

The treatment of the crude penicillin extract varies according to the objective, but involves the formation of an appropriate penicillin salt.

The solvent extract recovered in the previous stage is carefully extracted back with aqueous sodium hydroxide.

This is followed by charcoal treatment to eliminate pyrogens and by sterilization.

Pure metal salts of penicillin can be safely sterilized by dry heat, if desired. Thereafter, the aqueous solution of penicillin is subjected to crystallization.

FURTHER PROCESSING

For parental use, the antibiotic is packed in sterile vials as a powder or suspension.

For oral use, it is tabletted usually now with a film coating.

Searching tests (ex: for purity, potency) are performed on the appreciable number of random samples of the finished product.

It must satisfy fully all the strict government standards before being marketed

The yield of penicillin can be increased by:

Improvement in composition of the medium

Isolation of better penicillin producing mold sp. Penicillium chrysogenum which grow better in huge deep fermentation tank

Development of submerged culture technique for cultivation of mold in large volume of liquid medium through which sterile air is forced.

Mode of Antimicrobial Action

Bacteria have their own enzymes for

1.       Cell wall formation

2.       Protein synthesis

3.       DNA replication

4.       RNA synthesis

5.       Synthesis of essential metabolites

Classes of Antibiotics

B LactamAB

MacrolideAB

Tetracyclines

AminoglycosideAB

Polypeptide AB

Lyncosamide

Chloramphenicol

Rifamycins

Minor AB

Beta LactamAntibiotics

B-lactams inhibit transpeptidase.

Only effective against rapidly growing organisms that synthesize peptidoglycan. (Ineffective against mycobacteria.)

Beta Lactam Antibiotics

Penicillins

Cephalosporins

Carbapenems

Monobactams

beta lactamase Inhibitor

Penicillins

Derived from the fungus Penicillium

MO produce penicillinase(beta lactamase) hydrolyze beta lactamring inactive penicilloicacid

pH < 5 penillicacid

pH > 8 penicilloicacid

Inactivated by metal ion ( Zn, Cu), oxidizing agent

Penicillin G (Natural Penicillin)

Destroy by Gastric acid IM/IV

1stchoice gram (+) Strep., peptostreptococcus,B anthracis, Actinomycosis, Corynebacterium, Listeria, Neisseria& Treponema.

Adverse reaction allergic responses (penicillin acts as a hapten form Ag with body proteins-allergen)

Combination :

Penicillin G + Probenecid

Penicillin G + Benzathine/prokaine

K/Na Penicillin G

Penicillin V

Phenoxymethylpenicillin

Acid stable p.o

K/Na Penicillin V soluble in water, better absorption following oral admnistration

Penicillinase-Resistant Penicillins

Semysintheticpenicillins

Cloxacillin

Dicloxacillin

MethicillinNafcillin

Oxacillin

Resistant by Penicillinase

Recommended primarily for treatment staphylococcal infection resistant to other penicillins

Extended-spectrum penicillins(Amino penicillin)

Ampicillin

Aminobezylpenicillin

Acid stable

Absorbtion: disturbing by food

Inactivated by penicillinase

Broad spectrum

Haemophyllusnfluenza, Salmonella sp, Shigellasp, Proteus mrabilis, E.coli

Extended-spectrum penicillins(Amino penicillin)

Amoxicillin

Para hydroxyderivates of ampicillin

Acid stable

Absorption > ampicillin

Less GIT disturb than ampicillin

Inactivated by penicillinase

Broad spectrum

Haemophyllusnfluenza, Salmonella sp, Shigellasp, Proteus mrabilis, E.coli

Extended-spectrum penicillins(Active against P. aeruginosa)

Dinatrium ticalcillin

Analog tienylcarbenisillin

Spectrum = carbenicillin

Activity : P aeruginosa> carbenicillin

IM

Extended-spectrum penicillins(Active against P. aeruginosa)

Carbenicillindisodium

Carboxybenzylpenecillin

Gram (+) non beta lactamase(P aeruginosa)

RasematD and L

IM

ISK

Extended-spectrum penicillins(Active against P. aeruginosa)

Natriumpiperacillin

Aminobenzylpenecllinderivate

Spectrum = mezlocillin

Activity : P aeruginosa> carbenicillin

IM

NatriumMezlocillin

Spectrum = Cabenicillinand Ticalcillin

Gram + cocci, non beta lactamase, anaerob

IM/IV

ANTIBIOTIK B-LAKTAM CHEFALOSPORIN

Sejarah

Th1945 Brotzu mengisolasi Cephalosporium acremonium dari air laut disekitar P. Sardinia sifat antagonisme thd g+ & g-

Th1955-1956 Abraham dkk di Oxford mengisolasi 3 senyawa yg punya aktivitas antibiotik dari mikroorganisme tsb: sefalosporinP, penisilinN & sefalosporinC

Cephalosporin P, (a steroid antibiotic that resembles fusidicacid) with minimal antibacterial activity.

Cephalosporin N, later discovered to be identical with synnematinN (a penicillin derivative now called penicillin N)

Cephalosporin C kemiripan dgn penisilin:

–Sifatbiologi

–Sifatkimia

–Daya antibiotik<<, tapi merupakan bahan baku sintesis sefalosporin lainnya

• Cephalosporin C can be hydrolyzed by acid to 7-aminocephalosporanic acid.

Compounds containing 7-aminocephalosporanic acid are: - Relatively stable in dilute acid. - Highly resistant to penicillinase, regardless of the nature of their side chains and their affinity for the enzyme.

Cephalosporin(Structure)

This compound has been modified by the addition of different side chains to create a whole family of cephalosporin antibiotics.

Berdasarkan aktivitas antibakteri in vitro :

Sefalosporin generasi pertama

Sefalosporin generasi kedua

Sefalosporin generasi ketiga

Sefalosporin generasi pertama

Aktivitas antibakteri = ampisilin

Tidak diinaktivasi o/ penisilinase, tapi o/ sefalosporinase

Aktifthdg+ Staphylococcus aureus(kecuali Enterococcusfaecalis) & basil g- enterik termasuk E.coli, Klebsiella pneumoniae & Proteus mirabilis

Resisten: Enterobacter, Pseudomonas & Proteusprodusenindol. H. influenzae< ampisilin

Oral : sefaleksin, sefadroksil & sefadrin (parenteral juga)  aktif thd g+, basil g-<< parenteral

Parenteral: sefalotin, sefazolin& sefapirin  aktif thd basil g-

Sefalosporin generasi kedua

Spektrum= generasi I, diperluas Haemophillus influenzae, gonococcus, basil g- enterik

Parenteral: sefamandol, sefuroksim & sefonisidaktif thd H. influenzae

Sefoksitin, sefmetazol, sefotetanaktif thd basil g-anaerob: Bacteroides fragilis  generasiI I lain tdk aktif  infeksi abdominal

Peroral: sefaklor, sefprozil, lorakarbef sefuroksim aksetil & sefpodoksim  abs meningkat dgn makanan

Use: primarily for upper & lower respiratory tract infections

Sefalosporin generasi ketiga

Spektrum diperluas thd bbrp basil g-enterik termasuk strain yg memproduksi B-laktamase  per tidak sama dgn generasi I & II

Use: Meningitis, highly resistant & multi drug resistant Streptopneumoalong with vancomycin

Sefotaksimaktivitaspaling besarthdE.coli, Klebsiella, Enterobacter, Proteus, H. influenzae& Neisseria;utkmeningitis,parenteral

Parenteral: sefoperazonaktivitas thd: basil g-enterik < sefotaksim; Pseudomonas aeruginosa> sefotaksim

Seftazidim aktivitas paling besar thd Pseudomonas aeruginosa, kurang aktif thd Staphylococcus & Bacteroides fragilis

Seftriakson sangat aktif thd Meningococcus, Gonococcus (utamanya Neisseria gonorrhoeae) & H. influenzae,termasuk strain yg resisten thd ampisilin

Peroral : sefiksim aktivitas thd : koki g+ & H. influenzae produsen B-laktamase

Production

It is possible to convert penicillin V or Benzyl penicillin to a cephalosporin by chemical ring expasion.

The first generation cephalosprin Cephalexin, for example, can be made by this way.

Most cephalosporinsused in clinical practice are semi-synthetics produced from the fermentation product cephalosporin C

Cephalosporium acrimonium

The initial defined medium contained Glucose, Ammonium chloride, Methyl oleate, Metallic salts… little cephalosporin C was obtained

Production of cephalosporin C was induced by methionine, which cause the necessary thickening of the mycelium & raised cephalosporin C production to about 4g/L

Fermentation Conditions

fermentation is advantageously carried out at a temperature preferably about 25°C.

at a pH of from 5-8 and preferably about 6,

and for from 1-20 days, preferably 4-10 days.

The product is extracted from the culture fluid by adsorption on to carbon or resins rather than by solvent

AntibiotikaB-laktam yg lain (Carbapenem)

inti carbapenem (intisulfur pd penisilin diganti dg inti karbon)

Derivat dr thienamycin yg di produksi oleh Streptomyces cattleya

Aktivitas: gram +,gram -,mikrobaanaerob: Enterococcus faecalis, Bacteroides fragilis, P.aeruginosa,Serratia sp.

Penggunaan:IV/IM

Diperdagangan dikombinasi dg silastatin utk menurunkan efek samping imipenem

Monobaktam

B-laktam monosiklik

Diperdagangan hanya Aztreonam

Aktivitas : gram -,aerob, ≠ gram+ & anaerob

Resisten thdp -laktamase

Penggunaan IM/IV