Chapter 11.

ANTIBIOTICS AND CHEMOTHERAPEUTIC AGENTS

I.H.Siddique

 

11.1  CHAPTER REVIEW:

When a pathogenic organism attacks a susceptible host and overcomes the defense mechanisms of the body, the result is a disease process in that host. Any chemical agent which is used in the treatment of any disease is known as chemotherapeutic agent. Antimicrobial chemotherapeutic agents are used to treat infectious diseases. Antibiotic is a chemical substance produced by a microorganism which has the capacity, in dilute solutions, to inhibit the growth of or to kill other organisms. The important characteristic of these antibiotics is that they will stop the growth of specific organisms without causing major injury to the host. Thus, the antibiotics have selective toxicity. Antimicrobial chemotherapeutic agents may be antibacterial, antifungal, antiviral, or broad-spectrum antibiotics, destroying not only bacteria, but also rickettsias, chlamydias, and other pathogenic viruses.

THe historical development of antibiotics, their sources, mode of action, their specific activities, harmful side effects, and methods of testing the antibiotics are discussed in the following chapter.

 11.2 Introduction:

11.2.1 Chemotherapeutic Agents-

The chemotherapeutic agents Interfere directly with the multiplication of organisms and in concentrations not harmful to the host. Paul Ehrlich formulated the principles of selective toxicity and recognized the specific chemical relationship between parasites and drugs. He introduced arsphenamine, an organic compound of arsenic, as a cure for syphillis and other spirochetal diseases. Likewise, the organic arsenicals, and synthetic dyes, like trypan blue, were also found useful in the treatment of trypanosomiasis.

11.2.2 Chemotherapeutic Agents Active Against Bacteria

Domagk reported that the compound sulfonamide-crysoidin (Prontosil) would cure streptococcal infections in mice. It was found that the compound was only effective in the animal body and did not affect the organism in vitro. This lead to finding that the active part of this compound was sulfanilmide. Following this discovery, other potent derivatives were synthesized and found to be more effective therapeutically and were named 'sulfonamides'. Compounds in this group include sulfapyridine, sulfathiozole, sulfadiazine, and sulfaguanidine.

 

11.3 Competitive Inhibition

Certain inhibitors have been known to compete with normal substrates for binding to the active site of an enzyme. This principle of competitive inhibition explained the action of sulfanilamide: In order for the bacterium to synthesize folic acid (one of the B vitamins necessary for growth) it has to synthesize and put together several large molecules. One of these molecules, which makes up part of the folic acid molecule, is named paraminobenzoic acid. The chemical structure of PABA is very similar to that of sulfanilamide. Because of this similarity the enzyme attaches itself to the sulfanilamide (when it is present in sufficient amounts) but then cannot make it fit into the folic acid molecule. Therefore, the cell is not able to make folic acid and hence is unable to grow. It must be kept in mind that this is not a killing effect, and treatment must be continued long enough to allow the body defenses to destroy and get rid of the infecting organism. Only organisms which synthesize their own folic aice are affected by the action of the sulfonamides, and many sensitive bacteria develop a resistance to the sulfonamides so that they are no longer effective as chemotherapeutic agents. Sulfadiazine, particularly, remains in clinical use, but antibiotics have replaced the sulfanonamides for most purposes. (fig. 1) 

11.4 Antibiotics

Chemical compounds derived from or produced by living organisms, capable, in small concentrations, of inhibiting the life processes of microorganisms. About 150 or more of the antibiotics, produced by bacteria and fungi, have been identified. However, only a very few of these antibiotics have been found sufficiently non-toxic to be of use as therapeutic agents.

11.4.1 PENICILLIN.

Alexander Fleming (1929) observed cultures of Staphylococcus aureus lysed by a mold contaminant and this led to the discovery of penicillin. Chain & Florey (1940) purified the active principle and found it to be effective against gram-positive organisms both in vitro and in vivo. The penicillins are derived from certain species of Penicillium (eg., P. notatum and P. chrysogenum) and are obtained by extraction of cultures grown in special media. The various natural penicillins have been designated as F, G, K, O, and X. Penicillin G is the most satisfactory type to manufacture and use and 90% of commercially available penicillin is of this type. Semi-synthetic penicillins like Ampicillin, Cloxacillin and Methicillin have been introduced commercially. These have a antimicrobial spectrum. Cephalosporins derived from a mold, Cephalosporium resemble penicillins.

Penicillins lyse growing cells but have no effect on the viability of resting cells. Penicillin interferes in the synthesis of the cell wall of the bacterial cell, allowing the rest of the cell to grow. Without the protective cell wall, osmotic lysis of the cell results.

11.4.2 STREPTOMYCIN.

Streptomycin was discovered by Schatz, Bugie and Waksman in 1944. It is effective against Mycobacterium tuberculosis and many Gram-negative organisms. This antibiotic is obtained from fungus Streptomyces griseus. It is highly toxic and its prolonged use may cause deafness. Dihydrostreptomycin is a derivative of Streptomycin and is obtained by its dehydrogenation.

It is bactericidal in activity, and does not cause lysis of cells. A small amount of growth is required for its proper activity because it interferes with the biosynthesis of protein. Streptomycin treatment sometimes also leads to resistant mutant development against other kinds of chemotherapeutic agents. These mutants may sometimes require Streptomycin for their growth. This phenomenon is called `resistance' and 'dependence.'

11.4.3 BROAD-SPECTRUM ANTIBIOTICS.

Broad-spectrum antibiotics have a wide range of bactericidal activity. The two most common antibiotics of this class are Chloramphenicol and Tetracycline. Both of these have identical antimicrobial activity but are structurally different. Chloramphenicol is not only effective against gram-positive bacteria, but also gram-negative bacteria. The bacterial spectrum of chlortetracycline is similar to that of chloramphenicol. Chloramphenicol (Chloromycetin) was discovered by Ehrlich (1947) and is produce by Streptomyces venezuelae. It is now produced by chemical synthesis. Chlortetracycline (Aureomycin) was discovered by Duggar (1947) and is produced by Streptomyces aureofaciens. Oxytetracycline (Terramycin) was discovered by Finlay and associates (1950) and is produced by Streptomyces rimosus.

How do Chloramphenicol and Tetracycline act? Chloramphenicol binds with the ribosomes with their 50 subunits. Tetracycline interferes with the binding of charged RNA by the ribosomes, but chloramphenicol does not have this effect. Chloramphenicol inhibits amino acid incorporation which results in the incomplete polypetide formation.

11.4.4 POLYMYXINS.

These are groups of cyclic polypeptides which are produced by Bacillus polymyxa and other related bacilli. The most common polymxins are B and E. Polymyxin B is usually effective against gram-negative organisms and particularly Pseudomonas aeruginosa. It is nephrotoxic so its use is limited to local and alimentary canal infections.

How do polymyxins act? These antibiotics have surface-active properties of a cationic detergent. They damage the bacterial membrane and cause cell lysis.

11.4.5 BACITRACIN.

This antibiotic is produced by Bacillus licheniformis. It is effective against most of the gram-positive organisms like penicillin.

11.4.6 Neomycin.

This is derived from Streptomyces fradiae. It has a similar action like Streptomycin.

11.4.7 Erythromycin.

This is derived from Streptomyces erythraeus. It is effective against most of the gram-positive and some gram-negative organisms when given orally.

11.4.8 Other Antibiotics

l. CEPHALOSPORINS.

A fungus, Cephalosporium acremonium, yields several antibiotics, called cephalosporins. They resemble penicillins; and are active against both gram-postive and gram-negative bacteria, eg., spectinomycin which is used for the penicillin-resistant gonococci. Recently new B-lactam antibiotics, and particularly the third-generation cephalosporins, active against resistant organisms, have been developed. Cefotazime, Moxalactam and Cefoperazone are among the 10 to 15 members of this group. The penicillins were the firs generation of B-lactam antibiotics (isolated from P. notatum). The penicillins are now in their fourth generation each new generation showing an increased spectrum of activity. The second generation of B-lactams are the cephalosporins, first isolated from the fungus Cephalosporium acremonium. All the cephalosporins are produced by chemical modification of naturally occurring side chains--except moxalactam, which is totally synthetic. The key therapeutic feature of these agents is also the B-lactam ring. Held in a strained configuration by the fused ring system, the B-lactam and adjacent atoms have a spatial configuration similar to that of a peptidoglycan used in the synthesis of the bacterial cell wall. The antibiotics substitute for this peptidoglycan and inhibit enzymes important to cell wall synthesis. The B-lactams are effective only against actively growing bacteria. Since mammalian cells have a much different membrane, the B-lactams are highly specific for bacteia and have remarkably few side effects.

2. Antifungal Agents

a. NYSTATIN (MYCOSTATIN).

Antifungal agent derived from Streptomyces noursei. It is used as a local treatment for Candida infections.

b. AMPHOTERICIN B.

A broad-spectrum antifungal agent produced by Streptomyces nodosus. It is given intravenously and is effective against systemic fungus infections.

c. GRISEOFULVIN.

An oral antifungal antibiotic obtained from four kinds of Penicillium. It is fungustatic and not a fungicide. It has a wide range of activity against practically all of the dermatophytic fungi (eg. Microsporum, Epidermophyton, and Trichophyton). It does not act, however, on Candida albicans

d. CYCLOHEXIMIDE.

Produced by several Streptomyces species. it is active against fungi and has some antiprotozoal and anti-tumor activity.

 

11.5 Mechanism of Action of Antibiotics

At the cellular and subcellular level, most antimicrobial agents function in the following manner:

A. Antimicrobial Action through Inhibition of Cell Wall Synthesis.

Examples: Bacitracin, Cephalosporins, Cycloserine, Penicillins, Ristocetin, Vancomycin.

B. Antimicrobial Action through Inhibition of Cell Membrane Function.

Examples: Amphotericin B, Colistin, Nystatin, Polymyxins.

C. Antimicrobial Action through Inhibition of Protein Synthesis.

Examples: Chloramphenicol, Erythromycins, Lincomycins, Tetracyclines, Aminoglycosides: Amikacin, Gentamicin, Kanamycin, Neomycin, streptomycin, Tobramycin, etc.).

D. Antimicrobial Action through Inhibition of Nucleic Acid Synthesis.

Examples: Nalidixic Acid, Novobiocin, Pyrimethamine, Sulfonamides, Trimethoprim, Rifampin).

 

11.6. Resistance to Animicrobial Drugs

There are many different mechanisms by which microorganisms exhibit resistance to drugs.

A. Microorganisms produce enzymes that destroy the active drug.

Examples: Staphylococci resistant to Penicillin G produce a beta-lactamase that destroys the drug. Other beta-lactamases are produced by gram-negative rods.

B. Microorganisms change their permeability to the drug,

Example: tetracyclines accumulate in susceptible bacteria but not in resistant bacteria. Resistance to polymyxins is also associated with a change in permeability to the drugs.

C. Microorganisms develop an altered structural target for the drug,

Example: chromosomal resistance to aminoglycosides is associated with the loss or alteration of a specific protein on the 30S subunit of the bacterial ribosome that serves as a binding site in susceptible organisms.

D. Microorganisms develop an altered metabolic pathway that bypasses the reaction inhibited by the drug,

Example: some sulfonamide-resistant bacteria do not require extracellular PABA but, like mammalian cells, can utilize preformed folic acid.

E. Microorganisms develop an altered enzyme that can still perform its metabolic function but is much less affected by the drug than the enzyme in the susceptible organism,

Example: in some sulfonamide-susceptible bacteia, the tetrahydropteroic acid synthetase has a much higher affinity for sulfonamide than for PABA. In sulfonamide-resistant mutants, the opposite is the case.

 

11.7 Harmful Reactions of Antibiotics

A. Hypersensitivity or Allergy.

Penicillin is particularly notorious in this respect. Skin rashes, joint pains, and anaphyltic-like manifestations may result following penicillin therapy.

B. Induction of Bacterial Resistance.

The organisms develop resistance to a given antibiotic.

C. Effect on the Replacement Flora.

Prolonged use of the antibiotics tends to encourage multiplication of undersiralbe organisms like Proteus or Pseudomonas species which maybe harmful to the host.

D. Interactions at the Receptor Site

Many of the antibiotic drugs exert pharmacodynamic effects in the host in addition to their effects on the infecting pathogen. Several antibiotics produce neuromuscular blockade by competitive antagonism of acteylcholine at the myoneural junction. Bacitracin, colistimethate, streptomycin, dihydrostreptomycin, gentamicin, kanamycin, neomycin, paromomycin, polymyxin B, and viomycin have additive neuromuscular blocking effects among themselves and with other neuromuscular blocking agents. Tetracyclines may enhance the rate of development of cachexia because they exert an antianabolic effect. They would increase the catabolic effect of flucocorticoids. Chloramphenicol may interfere with antibody production in active immunization proceudres, eg., immune response to tetanus toxoid.

E. Toxic Interactions

Several of the antibiotcs will interact with other drugs and with each other to produce ototoxicosis. Dihydrostreptomycin, kanamycin, neomycin, ristoecetin, streptomycin, vancomycin, and other ototoxic drugs such as furosemide and ethacrynic acid may have progressive cumulative effects that can be addictive and may produce permanent deafness. Dimenhydrinate (Dramamine) may mask the appearance of ototoxic signs. Tetracycline, administered parenterally, may interact with methoxyflurane to produce an impairment of renal functions which may have fatal outcome.

11.8 Factors Affecting Antimicrobial Activity

A. pH of Environment.

Some drugs are more active at acid pH (eg., nitrofurantoin); others at alkaline pH (eg., aminoglycosides, sulfonamides).

B. Componets of medium.

Sodium polyanethol sulfonate and other anionic detergents inhibit aminoglycosides. PABA in tissue extracts antagonizes sulfonamides. Serum proteins bind penicillins in barying degrees, ranging from 40% fro methicillin to 98% for dicloxacillin.

C. Stability of drug.

At incubator temperature, several antimicrobial agents lose their activity. Chlortetracycline is inactivated rapidly and penicillins more slowly, whereas aminoglycosides, chloramphenicol, and polymyxin B are quite stable for long periods.

D. Size of incoculum.

In general, the larger the bacterial inoculum, the lower the sensitivity of the organisms.

E. Length of incubation.

The longer the incubation, the greater is the chance for resistant mutants to emerge.

F. Metabolic activity of microorganisms.

In general, actively and rapidly growing organisms are more susceptible to drug action than those in the resting phase. 

11.9 Interactions altering activity against pathogen

Some combinations of antibiotics act synergistically; others cause antagonistic interactions. Certain bacteriostatic antibotics (chloramphenicol, tetracyclines, erythromycin, sulfonamides) antagonize the bacterial antibiotics (penicillins, cephalosporins, streptomycin) because bacteriostatic drugs prevent multiplication of the organisms whereas the bactericidal drugs kill only multiplying bacteria. However, bacteriostatic drugs do not antagonize all bactericidal antibiotics, eg., polymyxins kill bacterial that are not multiplying. The degree of inhibition from these interactions depends on the relative concentrations of the antibiotics present. Corticosteriods may decrease the clinical response to bacteriostatic antibiotics by decreasing the inflammatory response and diminishing the phagocytic competency of leukocytes. The eradication of the infection, following bacteriostasis, is ultimately dependent on host defenses.

11.10 Antimicrobial activity in vitro-

Antimicrobial activity is measured in vitro in order to determine

 11.11 Avoidance of interactions

There are several measures the practitioner can take to minimize the chances of occurrence of thes interactions. Where possible, avoid multiple-drug therapy. The knowledgeable use of a single drug in the management of an infectious disease is vastly superior to the blind administration of a series of drugs with no regard as to how they may influence one another. Avoid combination products. With oral dosage forms, adjust regimen relative to feeding times. Generally, avoid simultaneous use of a bactericidal drug and a bacteriostatic drug in the same patient; for example, do not give penicillin parenterally and apply chlortetracycline ointment topically to treat staphylococcal dermatitis. Know the assets and the limitiations of each drug.

 

 Study Questions

1. What is meant by Chemotherapy and chemotherapeutic agent?

2. What are broad-spectrum antibiotics? Name several having a broad-range spectrum.

3. Briefly describe several anti-fungal agents.

4. Describe and discuss the harmful side effects of antibiotic therapy.

5. Describe several methods for testing antibiotic activity.

6. What is competitive inhibition? How does it work with sulfonamides?

7.What is the mechanism of action of the polymyxins?

8. How does streptomycin affect protein synthesis?

9. What are the tetracyclines?

l0. Define the Followings: Penicillinase, R-factors, plasmids, cephalosporins, synergism, isoniazid, chloramphenicol.

ll. Define drug resistance. How is it produced? What measures can be taken to minimize drug resistance?

l2. Define a chemotherapeutic agent. Distinguish between a synthetic chemotherapeutic agent and an antibiotic.

 

 

 

References

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Freemann, B.A. l985. Burrows Textbook of Microbiology. W.B. Saunders Company, Philadelphia, PA.

Tortora, G.J. , B. R. Funke, and C.L.Chase. l982. Microbiology. An Introduction. The Benjamin/ Cummings Publishing Co.,Inc. Menlo Park, CA.

Joklik, W.K., H.P. Willett, and D.B.Amos. Zinsser Microbiology. l984. Appleton-Century-Crofts, Norwalk, Conn.

Siddique, I.H. Antibiotic Residues in Foods. Handbook of Natural Toxins, Vol.7, Food Poisoning. l992. Marcel Dekker Publications, New York, N.Y.