1. Antibiotics Antimicrobial agent
chemical substances that are produced by microorganisms which kill or inhibit the growth of other microorganisms
(cf. an antibiotic is a chemotherapeutic agent with activity against microorganisms)
Penicillin의 놀라운 능력에 감명을 받아 다른 미생물의 성장을 억제하는 유용한 미생물을 흙으로부터 찾기 위해 일생을 바친 Selman A. Waksman과 그의 연구원들은 10,000번 이상의 흙을 배양하는 실
험을 수없이 반복하던 중, 1943년 병든 닭의 목에서 배양된 특이한 미생물이 결핵균의 성장을 억제하는 것을 관찰할 수 있었고, 이 물질을 streptomycin 이라고 명명하였다. Waksman은 흙 속의 곰팡이나 미생물로부터 생성되어 다른 미생물의 성장을 억제하거나 파괴시키는 화학물질을 총칭하여 “antibiotics”라는 용어를 처음으로 사용하였다
Antimicrobial agent
Chemical that kills or inhibits the growth of microorganisms

2. Antibiotics products of secondary metabolism
commercially useful antibiotics are produced primarily by filamentous fungi (Penicillium) and by bacteria of the actinomycete group (Streptomyces) and Bacillus
most antibiotics employed medically are used to treat bacterial diseases, although a few are known that are effective against fungal diseases

3. Some antibiotics produced commercially Antibiotic
Producing microorganism
Bacitracin
Bacillus licheniformis (EFB)
Cephalosporin
cephalosporiumsp (F)
Chloramphenicol
chemical synthesis (formerly by Streptomyces venezuelae)
Cycloheximide
Streptomyces griseus
Cycloserine
Streptomyces orchidaceus
Erythromycin
Streptomyces erythreus
Griseofulvin
Penicillium griseofulvin (F)
Kanamycin
Streptomyces kanamyceticus
Lincomycin
Streptomyces lincolnensis
Neomycin
Streptomyces fradiae
Nystatin
Streptomyces noursei
Penicillin
Penicillium chrusogenum (F)
Polymyxin B
Bacillus polymyxa (EFB)
Streptomycin
Tetracycline
Streptomyces rimosus
EFB: endospore-forming bacterium; F: fungus; others: actinomycete

4. 우리나라 의약품 산업 동향 분석

6. 높은 개발 비용
내성균으로 인한 짧은 항생제 수명

7. Search for New Antibiotics
screening of antibiotic producers
testing new microbial isolates for antibiotic
production
cross-streak method
comparison with known antibiotics
animal tests for toxicity and therapeutic activity in infected animals
clinical test
commercial production
therapeutic index = toxic dose/therapeutic dose(치료 지수=독성 용량/유효 용량)
toxic dose: the drug level at which the agent becomes too toxic for the host
therapeutic dose: the drug level required for clinical treatment of a particular infection

8. Isolation using selective media
for streptomyces and identification
of antibiotic producers using
an indicator organism (지표생물)
(eg. Staphylococcus aureus)

9. cross-streak method; Testing an organism for its antibiotic spectrum
of activity

11. Postmarketing studies

12. commercial production
strain improvement: high-yielding strains which do not produce undesirable side products
     mutation/selection, genetic engineering
development of fermentation process
development of efficient purification system

13. Overall process of extraction and purification of an antibiotic

14. Penicillins
Alexander Fleming in 1929 noted that a fungal contaminant (Penicillium notatum) had lysed the bacterial cells on an agar plate of Staphylococcus aureus.

15. Penicillins
Fleming found that culture filtrate of P. notatum was effective against many
Gram positive bacteria in laboratory conditions, and he even used it to control eye infections.
However, he could not purify this compound because of its instability,
and it was not until the period of the Second World War ( )
that Florey and Chain, working in the USA, managed to produce the antibiotic on an industrial scale for widespread use.
All three scientists shared the Nobel Prize for this work, and rightly so –
penicillin rapidly became the "wonder drug" which saved literally millions of lives.
It is still a "front line" antibiotic, in common use for some bacterial infections although the development of penicillin-resistance in several pathogenic bacteria now limits its effectiveness.  

16. The action of penicillin is seen in Figure A
The action of penicillin is seen in Figure A. This shows an 'overlay plate',
in which a central colony of the fungus Penicillium notatum was allowed
to grow on agar for 5-6 days, then the plate was overlaid with a thin film
of molten agar containing cells of the yellow bacterium, Micrococcus luteus.
The production of penicillin by the fungus has created a zone of growth
inhibition of the bacterium.
Figure B shows the typical asexual sporing structures of a species of Penicillium

17. Penicillin has an interesting mode of action:
it prevents the cross-linking (transpeptidation) of small peptide chains
in peptidoglycan, the main wall polymer of bacteria.
Pre-existing cells are unaffected,
but all newly produced cells grow abnormally,
unable to maintain their wall rigidity,
and they are susceptible to osmotic lysis.

18. Bacteria in penicillin solution, 30 min

19. the first β lactam antibiotic discovered, penicillin G, is active primarily
against G+ bacteria.
basic structure of penicillins is 6-aminopenicillanic acid (6-APA).
variable side chains in position 6

21. 2) biosynthetic penicillins
1) natural penicillins
   fermentation is carried out without addition of side-chain precusors.
   only benzylpenicillin (penicillin G) is therapeutically useful.
2) biosynthetic penicillins
   fermentation is carried out with a side-chain precusor.   
   penicillin G (phenyl acetic acid), penicillin V (penoxy acetic acid),
   penicillin O (allyl mercapto acetic acid)
3) semisynthetic penicillins
   another side-chain is coupled chemically to 6-APA (produced from
penicillin G chemically or by penicillin acylase). 
eg. Ampicillin, have significantly improved medical properties:
resistance to stomach acids so that they can be taken orally
extended range of activity against some Gram-negative bacteria.

22. Kinetics of the penicillin fermentation with Penicillium chrysogenum
Penicillin G is produced in 40,000
-200,000 liter fermentors.
Highly aerobic process.
A major ingredient is corn steep liquor.
When the carbon source has been
nearly exhausted,
the penicillin production phase begins.
By feeding with various components,
the production phase can be extended
for several days.
Kinetics of the penicillin fermentation with Penicillium chrysogenum

23. Other β lactam antibiotics cephalosporins
produced by Cephalosporium acremonium,
low toxicity (less allergenicity) , broad-spectrum antibiotic
norcadicin,
clavulanic acid,
thienamycin
Ceftriaxone, a cephalosporin

24. Tetracyclines produced by Streptomyces aureofaciens
broad-spectrum antibiotics
interfere with 30S ribosomal subunit function

25. Production scheme for chlortetracycline
with S. aureofaciens
more than 300 genes are involved in
the production of tetracycline
regulation of biosynthesis is complex
glucose and phosphate repress
the chlortetracycline synthesis

26. Vitamins Vitamin B12 as supplements for human food and animal feeds
more than $700 million per year
most vitamins are made commercially by chemical synthesis.
Vitamin B12 (cobalamin), B2 (riboflavin): by microbial fermentation
Vitamin B12
synthesized in nature exclusively
by microorganisms
humans must obtain vitamin B12
from food or as vitamin supplement
human deficiency:
pernicious anemia (악성빈혈):
low production of RBCs
and nervous system disorder
Pseudomonas denitrificans:
60 mg of vitamin B12/liter
(molasses as C source)

27. Riboflavin ---> coenzyme FAD and FMN fungus Ashbya gossypii
7 g/liter
great economical competition
between microbiological process
and chemical synthesis

28. Amino acids extensive uses
in food industry (as food and feed additives)
in medicine
in chemical industry (as starting materials)
Glutamic acid as flavor enhancer (monosodium glutamate, MSG)

29. Aspartic acid, phenylalanine ---> aspartame (artificial sweetener)
Aspartame is a nutritive sweetener made by joining two amino acids:
L-phenylalanine and L-aspartic acid, with a third component called
a methyl ester group.
Very little is needed for a sweet taste, making aspartame virtually non-caloric.
Relative Sweetness: 180 to 200 times sweeter than sucrose
Lysin as food and feed additive
Amino acids can be produced microbiologically
1) by direct fermentation
2) by enzymatic synthesis (MO: source of enzyme)

30. Feedback inhibition of lysine synthesis in Brevibacterium flavum

31. How to obtain a mutant cell in which aspartokinase is no longer
(AEC, a lysin analog)
How to obtain a mutant cell
in which aspartokinase is no longer
subject to feedback inhibition
positive selection of AEC-resistant
mutants:
mutagenesis of B. flavum
plating on media containing AEC
test colonies on plate with AEC
---> 60 g lysine/liter

32. Bioconversion (Biotransformation)
the use of microorganisms to carry out a chemical reaction that is more costly or not feasible nonbiologically
1) culture of cells
2) addition of chemicals to be converted
3) further incubation: chemical reaction
4) purification of desired product
major use has been in the production of certain steroid hormones.

33. *Bioconversion has another meaning in Biology
Bioconversion is the conversion of organic materials, such as plant or animal waste, into usable products or energy sources by biological processes or agents, such as certain microorganisms or enzymes.
paper, tires, fabric, construction materials, municipal solid waste, sludge, sewage
e.g. conversion (by enzymes or microbes) of cellulosic material into sugars which can then be fermented into ethanol

34. Enzyme Production by Microorganisms enzyme
highly specific catalysts:
most of the enzymes are used by food industry
enzyme produced in the largest amounts are the bacterial proteases, used as additives in laundry detergents

35. High-fructose corn syrup (HFCS)
isoglucose in Europe
glucose-fructose in Canada
corn syrups that has undergone enzymatic processing to increase its fructose content
widely used in processed foods and beverages, including soft drinks, yogurt, cookies, salad dressing and tomato soup
is somewhat cheaper than sugar in the United States
HFCS 90 (mostly for making HFCS 55), 90% fructose and 10% glucose;
HFCS 55 (mostly used in soft drinks), 55% fructose
HFCS 42 (used in many foods and baked goods), 42% fructose

37. production of high-fructose corn syrup
1. thinning reaction by alpha-amylase :
starch polysaccharide ---> shortened chain (reduced viscosity)
2. saccharification by glucoamylase:
shortened polysaccharide ---> glucose (corn syrup, 100% glucose)
3. isomerization by glucose isomerase:
glucose ---> fructose (42%)
4. Enrichment of fructose to 90% by chromatography step

38. Vinegar used as a flavoring ingredients in salads and other foods
ethanol --- acetic acid bacteria ---> acetic acid
acetic acid bacteria
Acetobacter, Gluconobacter
fairly acid tolerant
high oxygen demand
during growth
it is important to ensure
sufficient aeration
of the medium
starting material: wine (wine vinegar in France)
beer (malt vinegar in Germany)
alcoholic apple juice (cider vinegar in USA)
pure alcohol in water (distilled vinegar)

40. Yeasts in industry
eukaryotic micro-organisms classified in the kingdom Fungi.
unicellular
reproduce asexually by budding
yeasts are the most important and the most extensively used microorganisms in industry: Saccharomyces cerevisiae

41. Yeasts in industry Yeast cells Baker's yeast (bread making)
Dried food yeast (food supplements, rich in B vitamins and protein)
Dried feed yeast (animal feed)
Yeast products
Yeast extract (culture media, food additives)
B vitamins, Vitamin D
Invertase, galactosidase (enzymes for food industry)
ATP, NAD, RNA (research biochemicals)
Fermentation products
Ethanol (for industrial alcohol and as a gasoline extender)
Beer, Wine, Whiskey, Brandy, Vodka, Rum

42. Industrial production of yeast cells
(70% moisture)
Industrial production of yeast cells