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Antibacterial tests
• Purpose:
– To understand chemical methods to inactivate
bacteria.
• Theory
– Bactericide (bactericidal)
• Cide: kill; bactericide: a agent which can kill bacteria
– Bacteriostatic
• Static: not active; bacteriostatic agent: a agent which
inhibit growth of bacteria
• All bactericides are bacteriostatic agents depending
on concentration. For example, high concentration
kills bacteria but low concentration only inhibits.
• Not all bacteriostatic agents are bactercides because
of antibacterial mechanism. For example,
tetracycline and chloramphenicol
Tetracycline四環黴素
• Tetracycline antibiotics inhibit protein
synthesis by inhibiting the binding of
aminoacyl-tRNA to the mRNA-ribosome
complex. They do so mainly by binding to the
30S ribosomal subunit in the mRNA
translation complex.
• Oxytetracycline is one of
most commonly used
antibiotics in fishery.
Chloramphenicol 氯黴素
• Chloramphenicol is a bacteriostatic
antimicrobial originally derived from the
bacterium Streptomyces venezuelae, isolated
by David Gottlieb, and introduced into clinical
practice in 1949.
• It was the first antibiotic to be manufactured
synthetically on a large scale. Chloramphenicol is
effective against a wide variety of microorganisms
and still widely used in low income countries
because of its low cost.
• This antibiotic is still used in aquatic industry.
• It functions by inhibiting peptidyl transferase
activity of the bacterial ribosome, binding to the
50S ribosomal subunit, preventing peptide bond
formation.
Ampicillin安比西林
• Ampicillin is a beta-lactam antibiotic that has
been used extensively to treat bacterial
infections since 1961. It is considered part of
aminopenicillin in family.
• Ampicillin acts as a competitive inhibitor of
the enzyme transpeptidase. Transpeptidase is
needed by bacteria to make their cell walls. It
inhibits the third and final stage of bacterial
cell wall synthesis, which ultimately leads to
cell lysis.
屬青黴素類。
目前人類醫學治療常用。
逐漸被 另一相似產品，amoxicillin取代。
Streptomycin 鏈黴素
Streptomycin is derived from
the actinobacterium Streptomyces griseus.
Streptomycin binds to the small 16S rRNA of the 30S
subunit of the bacterial ribosome.
Streptomycin was first isolated on October 19, 1943
and the first antibiotic used to cure the disease
tuberculosis (TB). 肺結核
對第八對顱神經有損害作用，可引起前庭功能障礙
和聽覺喪失。若發現耳有堵塞感或耳鳴，應立即停
葯。
抗生素殺菌的選擇性
• Magic bullet: only kill the bad guys (bacteria)
• 原核細胞特有組織
– Cell wall: peptidoglycan, Gram positive
– Ribosome: 50S and 30S subunit
– 真核細胞的ribosome: 60S and 40S subunit
–
台灣對水產養殖用藥之最高殘留限量
• Ampicillin 安比西林 肌肉 魚 0.05 ppm
• Chlortetracycline、Oxytetracycline及Tetracycline 氯
四環黴素、羥四環黴素及四環黴素 肌肉 魚、大
明蝦、蝦 0.2 ppm
Microbiology
• Antibiotics
– Tetracycline (30S ribosome) 30μg 四環黴素
– Streptomycin (DNA gyrase) 10μg 鏈黴素
– Chloramphenicol (protein synthesis) 30μg 氯黴素
– Ampicillin (cell wall) 10μg 安比西林
– E. coli , S. aureus
– Pre-preparation
– 105 CFU/mL (two tubes/table)
• Spread plate of 104 CFU/plate
• 4 plates/group
– Each bacterial species has two plates
– Place two discs on a plate
• 2 disc/plate
• measure the diameter (直徑) of the clear zone
• Bactericide (殺菌 ; cide: kill) vs. bacteriostatic
(靜菌或抑菌: static: stay at one place or
status, don’t change)
• Both experiments are bacteriostatic not
bactericide tests.
• Bacteria do not grow yet.