Fermentation
DESPITE 1958
the economic problems of and the lack of epidemics such as the A4sian flu outbreak of a year ago, pharmaceutical sales reached new heights. Antibiotics for h u m a n medicine alone were valued a t $380,000,000: with another $67,000,000 used in animal feeds, veterinary drugs, and other miscellaneous products. By 1965 one estimate expects h u m a n needs alone will be valued a t $440,000,000, coupled with a substantial increase in nonpharmaceutical uses. Since the war: and widespread use of antibiotics, the average death rate caused by eight infectious diseases has been reduced by 58%, with tuberculosis leading the way a t 78%.
SAMUEL C. BEESCH attended Ohio State University where he received his B.A. in 1938 and a M.S. in 1948. From 1939 to 1950 he was associated with Publicker Industries Inc., Philadelphia, Pa. Since then he has been on the staff of Chas. Pfizer and Co., where he i s head of the Biochemical Pilot Plant. He is the author of numerous patents and papers on industrial fermentation processes. He is a member of the ACS, Society of Industrial Microbiologists, Society of American Bacteriologists (New York City Branch), N. Y. Sewage and Industrial Wastes Association, and American Ordnance Association. He holds a reserve commission as a Sanitarian (Lt. Comdr.) in the U. S. Public Health Service. FRED W. TANNER, Jr., attended the University of Illinois (B.S., 1938; M.S., 1939) and Cornell University (Ph.D., 1942). After a year at the N. Y. Agricultural Experiment Station, Geneva, he joined the U.S.D.A. a t the Northern Regional Research Laboratory, Peoria, and in 1948 the Biochemical Research Department a t Pfizer. Publications and patents have dealt with various fermentation products including vitamins and antibiotics. He is a member of the Society of American Bacteriologists.
1086
A second economic anal) sis of the drug industry estimates antibiotic sales in 1958 a t $431,000,000; steroid hormones, $140,000,000 (with corticoids accounting for $95,000,000 and sex hormones, $25,000,000), and vitamins, $250,000,000. Thus, these three classes comprise $800,000,000 million of the total ethical and proprietary market of $1,900,000,000 in sales. T h e wonder drugs of the 1930’s, the sulfas, still command $42,000,000. Anticancer agents from microorganisms in the future may comprise another important group of products if the current explorations are successful. Certain other products, such as organic acids, have continued their
Table I.
growth. A domestic producer has announced plans to increase production facilities for citric acid. Gluconic and ascorbic (an indirect fermentation product) acids are finding wider uses. Isoascorbic acid (also an indirect product) is being used for its antioxidant properties. \-itamin prices, both of fermentation and synthetic origin, continue to decline but with high volume sales. Importation has influenced prices. Several dramatic new types of corticosteroid hormones have broadened the market and increased competition. Fluorosteroids were quickly followed by potentiation effects accomplished by
Expansion in the Drug and Fermentation Industry
Company .4bbott Laboratories -4bbott Laboratories
Type of Facility Research Mfg.
Location North Chicago, Ill. Sidney, Australia
Allen and Hanburys, Ltd. Bristol Laboratories (Bristol-Myers Co.) Carter Products Carling Breweries Carling Breweries Federated Pharmaceutical Co., Ltd. Fleishman Laboratories Lakeside Laboratories Lederle (American Cyanamid ) Lederle (American Cy-anamid) Merck & Co.
Mfg.
Tehran, Iran
Products ..... Fine chemicals, pharmaceuticals Pharmaceuticals
Research
Syracuse, N. Y.
Cancer research
Mf5. Mfg. Mfg. Mfg.
Cranbury, N. J. Toronto, Ont. Atlanta, Ga. Jamaica, B.TV.1.
Research
Chicago, 111.
Pharmaceuticals Beer Beer Pharmaceuticals, antibiotics Basic research, quality control
Research Mfg.
Milwaukee, \Vis. Gossport, England
Antibiotics, drugs
Mfg.
TVellborn, Ont.
.4ntibiotics
Primpi, India
Streptomycin, dihydrostreptomycin Fermentation, steroids Vitamins, hormones .4ntibiotics, hormones Citric acid
Mfp. (tech. assistance to India) Merck Sr. Co. Mfg. Merck S: Co. Mfg. Mfg. Merck & Co. Miles Laboratories Mfi. Parke-Davis Research Parke-Davis Office-mfg.-lab. Research A. H. Robbins Research Schering Corp. Schering A. G. (Berlin) Mfg. E. R . Squibb & Sons Mfg.-distribution (Olin Mathieson) Syntex Research L-pjohn Co. Mfg.-distribution Upjohn Co. Upjohn Co. Upjohn Co. \l.’arner-Lambert
Mfg. Mfg. Research Mfg.
Tl’estern \l’holesale Drugs, Ltd. TVyeth Laboratories
Mfg.-sales facilities Research
INDUSTRIAL AND ENCslNEERlNG CHEMISTRY
Buenos Aires India Cali, Colombia Elkhart, Ind. Ann .4rbor, Mich. Bogota, Colombia Richmond, Va. Union, N. J. Japan France Mexico City France BoTota, Colombia Johannesburg, S. A. Kalamazoo, Mich. Villawood, Australia \.’ancouver! B. C. Radnor, Pa
.....
.....
Pharmaceuticals
.....
Biochem. pilot plant Pharmaceuticals Pharmaceuticalsdistribution
.....
Pharmaceuticalsdistribution Upjohn Products Upjohn Products I
.
.
.
.
Cosmetics, pharmaceuticals Pharmaceuticals
velopment in the antibiotics industry of Europe, as well as Russia and its satellites. There is reason to believe that Soviet-produced antibiotics will be weapons of economic and political influence. Substantial increases in production can be expected from these countries. The microbe is entering lustily into a new field: that of amino acids: L-lysine of fermentation origin is noiv supplied in food: pharmaceutical, and feed grades: and r-glutamic acid may be expected to compete Ivith the more classical sources. Yet. production efficiency gains appear 10 be parallelling sales increases for the most part, as general industry-!vide domestic plant expansions are not in vogue. This may be partially influenced by production facilities installed in other countries, some of lvhich are subsidiaries of U. S. firms. Some unconventional applications of microorganisms are of interest. Microbial insecticides consisting of live spores of Bacillus thuringiensis are being tested against pests associated with alfalfa, corn: cabbage, tobacco, and other crops. Soil flora which thrive on hydrocarbon gases may indicate petroleum deposits. Certain bacteria are being used to liberate oil from oil-bearing sands, sediments. shales, and tars. Bacterial acids leach copper, zinc, and iron and manganese from many types of low grade ores. Even “coal fermentations’‘ are being studied as sources of ne\v organic intermediates. hlicrobes are being sought Ivhich can use noxious gases. Algae are being studied as oxygen, lvater, and food suppliers for future space travel. Bacteria are being exploited to remove grease from selver lines and to destroy and detoxify phenol and formaldehyde Fvaste liquors from chemical processes. I t is hoped they may also destroy detergents in selvage treatment plants.
Plant Expansion and Production T h e fermentation industry continues a world-wide expansion of operations. Domestically, expansions are small: except for research (Tables I and 11). The C . S. Commerce Department recently announced that it will now alloLv export licenses of antibiotics and similar drugs to Russia and its European satellites. Included in the list are penicillin, streptomyin and dihydrostreptomycin. chlorotetracycline, oxytetracycline, and all similar antibiotics including sulfonamides. A total embargo is still in
These antibiotics are produced and packaged behind the iron curtain. In most cases, they are rather primitive examples of packaging drugs. Note that the names are of U. S. origin on some packages
effect on Communist China? North Korea, and S o r t h 1Yetnam. T h e general production figures on antibiotics production in Russia and its satellites are not known. .4 recent statcment by Dremov of the RIoscoiv Pharmaceutical Institute says that by 1960 antibiotic production will increase by 2705 over 1955. I t is evident from this statement that present production is low as compared with U. S. Latest L.S. production figur-s are shown in Table 111.
General Revielvs and books of current biochemical interest are listed in Table I\-.
Solvents and Alcohols Fermentation production of solvents and alcohol continues to decline in the U. S . Foreign alcohol production such as in India has run u p against the problem of full utilization of the alcohol produced. India produced 15,500,000 gallons in 1958, u p 210% since 1957. A ne\v target date is 45,000,000 gallons by 1960. India is considering exporting its present surplus. All of its alcohol is produced by fermentation. VOL. 51,
Industrial alcohol production by fermentation and synthetic plants in the U. S. A . since 1950 is shoirn in Table 1’. A steady decline in fermentation alcohol is evident. A recent nexv use of alcohol in the U. S.has been in animal feeds, such as morea ( I B ) . Morea contains ethyl alcohol in a low concentration to provide the proper environment in the rumen for the most effective use of urea and other feed ingredients. I t is used specifically to promote the metabolism of rumen microorganisms which in turn provide necessary nourishment to the animal. Reports of current interest on alcohol fermentation are scarce; those on acetone-butanol fermentation are listed in Table V I .
Citric, Itaconic, Fumaric, and Gluconic Acids lfiles Laboratories plans for expansion of its present citric acid plant (Table I) will double present output of citric acid. T h e new plant will also utilize the submerged process. Recent publications on citric, itaconic, fumaric, and gluconic acids are shown in Table \ T I , NO. 9, P A R T II
SEPTEM5ER 1959
1087
Gibberellin continues to arouse interest in the agricultural field. A recent use involving potassium gibberellate on naval oranges increased the juice content 9% and the vitamin C content 13%. The FDA has cleared gibberellins for seed treatment of lima, snap and soy-
Table II. Parent Co. Allied Laboratories Bard Pharmaceuticals Dumas-Milner Corp. Glaxo Labs. Ltd. Hampden-Harvard Breweries Kingsbury Breweries
Household products Antibiotics, vaccines Beer Beet Mouth washes
Sestle-Lemur
Cosmetics, perfumes Beer
Pabst Bretving Co. Chas. Pfizer Br Co. Chas. Pfizer & Co. Plough. Inc. Renfield Importers, Ltd. Rexall Drug Co.
Chemicals, antibiotics Chemicals, antibiotics Drugs, pharmaceuticals Importing Co. Drugs
Schering Corp.
Pharmaceuticals Drugs
Sterling Drug Co.
Drugs
Vick Chemical
Drugs, chemicals, plastics Drugs, chemicals, plastics
A. H. Robbins
Vick Chemical
1""
P0uI1as.
IUS"
cleareu
UY
LIlc
Thompson seedless and Black Corinth grapes, with a concentration of 5 to 50 50 p.p.m.: a t rates of 5 to 50 gallons per acre.
Company Acquisitions
Products Drugs Pharmaceuticals
Lavoris Co
Per
FDA are dipping Of Potato seed Pieces in a solution of 1 P,P-m. and spraying of
Acquired Co Campana Sales Co. G. F. Harvey Co. (Saratoga Springs, N. Y . ) Taylor Labs, Inc. (Houston, Tex.) Allen and Hanburys, Ltd. Dobler Brewing (Albany, N. U.) Sioux City Brewing Co. (Sioux City, IoGa) Milton Antiseptic (England) E. L. Patch Co. (Boston, Mass.) Blatz Co. (Milwaukee, \Vis.) Dumex Co. (India, Ceylon, Pakistan) Kemball, Bishop & Co. (Folkstone, England) Creolin Co.
Products Proprietary, cosmetics Ointments Drug products Pharmaceuticals, insulin Beer Beel Mouth washes Pharmaceutical specialties Beer Pharmaceuticals, antibiotics, vitamins Tartaric, citric, gluconic acids Disinfectants
Medley Distilling Co. (Owensboro, K y . ) Carnegies, Ltd. (Welyn, London) Whittier Labs (Chicago) American Scientific Labs (Madison, \Vis.) Delaware Poultry Labs Inc. (Millsboro, Del.) Lavoris Co.
\$-hiskey iVhiskey Pharmaceuticals
Walker Laboratories (Mt. Vernon, N. Y . )
Vitamins, drugs
Pharmaceuticals Veterinary biologicals pharmaceuticals Poultry vaccines, animal medicines Mouth washes
U. S. Production and Sales of Some Fermentation Products" Production, Sales, Fermentation Product Year Lb. Lh. Value, $ Penicillin and salts 1956 631,000 593,000 63,529,000
Table 111.
Streptomycin Dihydrostreptomycin Neomycin and salts All others (broad spectrum excluding tetra-
cycline Tetracycline Antibiotics for feed supplements, food preservation, crops Total antibiotics for human, animal, and poultry Riboflavin for human, animal, and poultry use Vitamin Biz Hydrocortisone alcohol and acetate a
1957 1956 1957 1956 1957 1956 1957 1956 1957 1956 1957 1956 1957 1956 1957 1956 1957 1956 1957 1956 1957
U. S. Tariff Commission, Washington, D. C.
1088
INDUSTRIAL AND ENGINEERING CHEMISTRY
694,000 130,000 198,000 492,000 582,000 17,000 24,000 477,000 465,000 220,000 404,000 779,000 870,000 1,967,000 2,373,000 370,000 405,000 655 790 7,000 8,000
601,000 66,299,000 5,604,000 147,000 169,000 6,786,000 431,000 18,198,000 462,000 20,039,000 14,000 5,989,000 19,000 6,328,000 366,000 105,148,000 430,000 113,627,000 186,000 72,903,000 299,000 108,466,000 683,000 28,108,000 795,000 31,307,000 1,737,000 271,371,000 1,986,000 323,596,000 245,000 5,321,000 5,613,000 320,000 450 22,640,000 537 2 1,869,000 4,000 9,552,000 3,000 6,342,000
aiiu ~ c ; x d > iiiuiLaLc
LUCLUII
y i c i u ~C a i i
UL
increased 207, by the use of gibberellin. Recent publications of interest are shown in Table V-111.
6-Ca rote ne As pointed out in the last review, fermentative production of p-carotene is now receiving special attention. Several papers of current interest are tabulated in Table IX. Amino Acids Interest continues in the fermentative production of various amino acids. Merck & Co.: Inc., has been licensed by Kyowa Chemical Co. (Japan) to produce L-glutamic acid by fermentation. Researchers a t Kyowa also reported that the culture used in the L-glutamic process could be mutated to produce either L-lysine or L-ornithine (ZF, 3 F ) . Merck is now in production with this lysine process. Pfizer also continues to produce Llysine by its fermentation process. T h e present price of L-lysine hydrochloride is now $4.95 per pound. Studies of interest are shown in Table X.
Dextran Interest in dextran remains low. New uses are still being exploited. Some interest has developed in the production of other bacterial polysaccharides, and in gums from yeast. Some of these publications (7G-7G) are cited.
Vitamins Considerable interest is still maintained in vitainin BP and BIZ fermentations. A recent report shows that antimetabolites of vitamin B I ~may be more effective in treating leukemia and other forms of cancer than the antifolic compounds now used. Recent tests involve use of the anilide and the ethyl amide of the monocarboxylic acid of vitamin B ~ Q .Reports of current interest on the vitamins are listed in Tables XI and X I I .
Antibiotics Antibiotics remain the principle fermentation products and the mainstay of the pharmaceutical industry, particularly in the United States. T h e industry has grown rapidly in the United Kingdom, Western Europe, Eastern Europe, Middle East, and South Africa. Antibiotics a c c o u n ~ for more than 207, of the monetarv value of all pharmaceuticals produced in the United States. Pridham and others (74K) have prepared useful classification of selected groups of Streptomyces, containing an excellent bibliography on antibiotics.
.4 concept of "lymphotropic antibiotics" was presented ( 5 9 K ) . ,4ntibiotics a n d other crystalloid molecules circulate via the vascular system, with loiv levels in the lymph a n d lymph nodes. Macromolecules have a particular affinity for the lymph system. Combinations of certain basic antibiotics such as streptomycin. neomycin, viomycin, a n d streptothricin with macromolecules such as sulfonic or phosphorylated polysaccharides were claimed to lower toxicity and blood levels b u t prolong blood levels and increase lymph levels. Bile salts were reported to lower the minimum inhibitory concentrations (MIC) levels of neomycin for Staphylococcus aureus and penicillin for resistant strains b u t not for sensitive strains (781;). For some other antibiotics, MIC levels were raised. A conference on basic a n d clinical research on kanamycin was held in New York, July 1958 ( 4 K ) . Waksman (27A) served as editor to present a summary of the practical applications of neom)-cin. In spite of hundreds found in the past decade, several new antibiotics were reported during the year. These a r e summarized in T a b l e XIII. Additional production information concerning some of the older antibiotics is found in the following references (9K, 79K, 29K, 371;39K, 67K, 76K. and 7121;).
Antibiotic Chemistry. T h e chemistry of complex antibiotic structures continues to yield to the efforts of chemists. Particularly striking is the work of Patrick a n d others (72K) who have for the first time determined the structure of a polyene antifungal antibiotic, the tetraene pimaricin, which was shown to be a member of the macrolide family. H a r m a n and others (42K) have reported the structure of another antifungal, eulicin (ClaHjzOiSs) which is not a polyene, but rather a complex amide.
Table VI. Subject Butanol-2-propanol fermentation industry use of sugary raw materials Starting materials and fermentation equipment Fermentation and distillation Shift of solvent ratio in subculture Fermentation of molasses with Weizman strain Fermentation of sugar solutions to butanol-acetone and ethanol Inhibitory factors in acetone-butanol fermentation Butanol-acetone fermentation of sugar cane juice Battery method of fermentation in acetone-butanol production
Table VII. Table IV. General Reviews and Books of Biochemical Interest Subject
Ref.
Penicillia ( 7-4) Yeasts (2,4, 2 7 A ) Oxidative fermentations by bacteria (3A) Homemade wines ( 4 A1 Sewerage and sewage treatment ( 5 A , 7 4 A ) Industrial and economic microbiology in North America (6A) Microbiology (7A) Continuous cultivation of microorganisms (8.4) Industrial utilization of yeasts (9.4) Antibiotic manufacture (7 0 ~ Microbial biochemistry (17A) .4ntibiotics 72A Biological laboratory data ( 73A Genetic basis of selection ( 75A Biochemistry (76.4) Biological treatment of sewagc and industrial wastes ( 7 7.4 ) Enzymology (78.4) Microbiological fermentation and chemical biosynthesis of medicines (79A) Enzyme chemistry (20A1 Environmental sanitation (22A) Steroid chemistry (23A) Biochemical engineering (25~) Gibberellin (24.4 ) Chemical transformation by microorganisms (26A) Neomycin (27~) British fermentation industries (28A)
Table V.
Industrial Alcohol Production"
'iear
Fermentation, Million Gal.
Synthetic, Million Gal.
1950 1951 1952 1953 1954 1955 1956 1957
61.2 120.7 120.2 108.1 45.5 44.5 74.9 31.3
104.0 113.5 125.9 147.5 153.7 172.3 180.6 192.6
a
Stanford Research Institute.
Reports on Acetone-Butanol Kef.
Results
Commercial production using C ~ O J (2B) tridium t o m u m ; yields of 30-340/, on initial sugar in 30-40 hr. Uses 6.5-7'.070 sugar 13B) (4B-6B) SubculturinT \L ithout heat shock changes ratio of solvents and yield 7B Uneconomical Additions of Ca(0Ac)r or AcOH : yield 40y0 on sugar utilized. Contaminants affecting fermentation 16 new \.arieties of c. acetohzlty~icum were isolated : effectiveness of 86 additives First-order reaction
SB ) 9B ) IOB)
70B)
Studies on Citric, Itaconic, Fumaric, and Gluconic Acids
Subject Autolysis of ' 4 . foenicis, a citric acid strain Formation of 2-keto-1-idonic acid from gluconate and lime idonate by Pseudomonas chromospirans Faerber Production of citric acid by submerged fermentation process Increasing citric acid production of active strain of A . niger Fumaric acid fermentation process
Citric acid production and changes of potential in surface and deep culture Submerged fermentation for citric acid Production of calcium 2-ketoyluconate by fermentation Lvith Pseudomonas species Changes in mineral composition of .4. nzger during biosynthesis of citric acid Production of itaconic acid
Calcium gluconate by fermentation Industrial production of gluconic acid by bacteria Calcium gluconate by fermentation
Results Methods of inducing autolysis
U.S.S.R. prncess Culturinq in molasses solution in presence of citric acid increases activit). Process for production of fumaric acid using Rhizopus oryzae grown on inverted molasses, salts, media containing Ni ions at about 10-300 p.p.m. media A . niger produced citric acid from sugar only at oxidation-reduction potentials below 11.5 Process utilizes A . saitoi; media changes, p H control, and use of oxygen improved yields Fermentation and recovery process, including costs of plant to produce 1,000,000 pounds yearly .4nalysis of mat during various stages for mineral composition Addition of finely divided (1.50 microns) adsorbents to broth-CaS04, kaolin, Fuller's earth, etc. improves yield 0.5 to 5 gramsjliter Description of a patented process using Pseudomonas Juorescem Used P . ovalis; ferments 13yoglucose in 32 hours; yield 94-95y0 oftheoretical; possibility of continuous process Rate of transformation faster in recycling drum 7's. vat
VOL. 5 1 , NO. 9, P A R T I I
SEPTEMBER 1959
1089
UNIT PROCESSES T h e isolation a n d structure of acetomycin ( C I O H ~ I O Lwas ) also reported
(XK). Amecetin B has been distinguished from amecetin (80K). The former yields cytosamine o n alkaline hydrolysis a n d p-arninobenzoylcytosine o n mild acid hydrolysis. Amecetin also yields D-a-methylserine. T h e structure of puromycin is 6 dimethylamino - 9 - [ 3 - deoxy - 3 - (pmethoxy - L phenylalanylamino) - /3-
-
~-ribofuranosyl]-P-purine(40K).
A
structure has been proposed for the antibacterial macrolide, pikromycin (31;). Blastmycin (C&36N2o9) (77OK),o n alkaline hydrolysis, yields blastmycinone, 2 - ti - butyl - 3 - hydroxy 4 - methyl - 4lactone, a n d blastmycic acid, ,V-(formyl3-aminosalicylal)-L-threonine. Chartreusin, (C32H 3 4 - 3 6 0 16) (88K), contains D-fucose, D-digitalose, and aglycone (CI9H10-1206),which is a derivative of A-methyl-2-phenyl naphthalene or x-methyl-2,3-benzofluorene containing a lactone group, two phenolic hydroxyls, and t\vo oxygen atoms of unknown
-
Table VIII.
Gibberellins assay Effect of gibberellic acid on soil microorganisms Effects of soil placement and dosage of gibberellins to plant response Action of gibberellins on bre\ser's yeast Physiological effects of gibberellins Giberellin and higher plants
Biosynthesis of p-carotene from DL-Bhydroxy-@-methyl-6[2 '4CI-valerolactone by Phycomyces biakesleanus and carrot slices Effect of various grains on production of @-caroteneby microorganisms
Influence of oils and fatty acids on production of p-carotene by microorganisms Production of @-carotene by species in Rhodotorula
Biosynthesis of p-carotene
1090
to be a n octa- or heptacyclopeptide with two or three amino acids on a side chain branched onto a n a,y-diaminobutyric acid end of the chain, joined to the carboxyl of isopelargonic acid ( 7 4 K ) . A neiv structural formula for gliotoxin is considered consistent with all chemical facts (72K). A six-membered carbocylic ring occurs in fumagillin (57K). Other products were discussed also ( I P K , 2 8 K ) . Dutcher (371;) has proposed a revised formula for aspergillic acid and hydroxy aspergillic acid. Trichoderma Liride uses phenylalanine as a precursor to the indole moity of
Results Process of preparing gibberellic acid Lvith Gibberella jujikuroz in an aerated nutrient solution characterized by addition of carbon dioxide ( 5 to 10% of air used) Fluorometric technique for determining gibberellin and gibberellic acids Induced some stimulation and inhibition not consistent with application rate Used granular potassium gibberellate at
Ref. (7Di
various rates
No value on Saccharomyes cersisiae Chemical nature, distribution, and physiological action of Field observations with vrgetable crops
Investigations on p-Carotene
Subject Slicrobiological production of /3carotene in shaken flasks
Biosynthesis of p-cz1r0tene-C'~
(+)cis-2,4-dimethylcyclohexanone(33K). The polypeptide, polymyxin B, is thought
Investigations on Gibberellin
Subject Metabolic process for preparation of gibberellic acid
Table IX.
nature. O n e phenolic group is glycosidically linked to a disaccharide. The pentaene fungichromin ((235HsoOlJ, o n periodic oxidation, yields the chromophore 2-methyl-2,4,6,8,1O-dodecapentaenedial which is readily reduced to the diol with the same ultraviolet absorption as fungichromin. T h e alkaline degradation product of actidione is
Results Carotene production with and strains of Choanephoraceae increased 4-5 times over that of unmated strains; yields further enhanced by addition of vegetable oils, detergent, and /3-ionone Growing shake cultures of Phycomyc~s blakesleanus will incorporate mevalonic acid into @-carotene
+
Hexane-extracted soybean meal in place of grain resulted in higher yields (8 mg. of carotene per gram of dry fermentation solids) Used choice white grease to improve yields ; yield directly proportional to percentage of grease added, up to 457, Kutritional factors which influence production by species of Rhodotorula in nonsynthetic medium; yields of 5.2-120.8 mMg./gram of dry cells; thiamine stimulated synthesis Synthesis by algae Anabaena cjiindrica and A . uariabilis Asparagine (0.2'36) in culture medium stimulates production of @-caroteneby Phycomyces blakesleanus 30-fold
INDUSTRIAL AND ENGINEERING CHEMISTRY
Ref. ( 7E)
gliotoxin. Tryptophan and acetate bvere not used. The carboxyl of indole-2carboxylic acid is the C1 of phenJIalanine (89K). T h e structure of kanamycin is essentially complete (201;). T w o hexosamine units occur, one of which is 6-deoxy-6amino-D-glucose (6-glucosamine) (26A: 27K). Kanamycin B has been isolated by countercurrent distribution and Amberlite XE-64 chromatography. Hydrolysis of .T-acetyl kanamycin B yields 2-deoxystreptamine and kanosamine ; it does not contain 6-glucosamine. An unidentified ninhydrin-positive substance was encountered (77K). Tetracyclines. T h e past year has seen several new developments in the t e t r a q c l i n e series of antibiotics. Snell (86K)described a method for preparing radioactive oxytetracycline by using carbon-14 labeled acetate precursors. Boothe a n d others ( 7 5 K ) reported the total synthesis of a four-ring compound
(& dedimethylamino-12a-deoxy-6-demethylanhydrochlorotetracycline) prcviously obtained as a degradation product from chlorotetracycline. T h e synthesis required some 30 steps. By mutation of Streptomyces aureofaciens. the Lederle group has prepared other interesting analogs in this famil>-. Active 6-demethylchlorotetracyclinc analogs, and 6-demethylbromotetrac~cline were obtained from the media containing the appropriate inorganic halides; 6-deinethyltetracycline \vas prepared from both halotetracyclines by catalytic hydrogenation (7K, 52K, 56K). T h e clinical effectiveness of 6-demethylchlorotetracycline !vas reported ( 3 7 K ) . A biologically inactive analog uf chlorotetracycline was found to be the product of another mutant of S. aurenJaciens (53K): 7-chloro-5a(l la)-dehydrotetracycline. On catalytic hydrogenation this substance was converted to tetracycline and a new isomer, 5a-epitetrac!-cline, the latter being biologically inactive. Of further interest is the report ( 5 4 K ) that "normal" S. aureoJacim r effectively hydrogenates 7 - c h l o r o - h (1 la) dehydrotetracycline to the parent antibiotic. 7-chlorotetracycline. Stephens and others (871C): by hydrogenolysis of tetracycline under acid conditions with a palladium catalyst. obtained 6-deoxytetracycline among the products. Oxytetracycline yielded 6deoxyoxytetracycine, and O-demethyltetracycline yielded 6-deoxy-6-demethyltetracycline. This series of compounds does not undergo acid degradation to the respective anhydro products. T h e 6deoxytetracycline and 6-deoxy-6demethylretracycline have been reported by others ( 2 K ) . T\vo heptaene antifungal antibiotics kvere isolated from S. aureofacienr (461;).
FERMENTATION Shaposhnikov and others (871;)have reported a synthetic medium for the biosynthesis of oxytetracycline, containing starch, ammonium salts, and trace elements tvith yields of 1500 to 1300 pg./ml. Doskocil a n d others (30K) studied the biochemistry of S. rimosus fermentations in relation to oxytetracycline production. Considerable pyruvic acid accumulates in the early phase (Gram-positive hyphae). H y p h a e fragmentation follows, with pyruvic acid consumption. T h e fragments form secondar>- hyphae (Gram-negative) which produce the antibiotic and other pigments. Tetracycline predominates in 5‘. aureomiens broths containing as little as 0.0025% 2-mercaptobenzthiazole, particularly in the presence of bromide ion (6E;). McCormick a n d others (551;) ha\,e used 2,5-dimercapto-1,3,4-thiadiazole for the same purpose. hfinieri a n d others (621;) employed deionized corn steep liquor. Various recovery procedures have been developed (76K,35K, 571;, 73K, 703K, 706K). Lseful fermentation media and procedures a r e summarized in Table XI\’. Streptomycin. S1:orkers a t Takeda Pharmaceutical Industries (651;, 961;, 98K) have found a strain of Streptomyces, S. humidis n . sp., which produces dihydrostreptom>cin as a direct fermentation product. This is a particularly interesting discovery. Another reduction product, dihydrodeoxystreptomycin? can be prepared from streptomycin by a n improved process if hydrogenation occurs in the presence of a n amalgamated aluminum catalyst a t p H 2.2 (7081;). Dihydrodeoxystreptomycin is a clinically effective form of the antibiotic. Penicillin. T h e biogenetic origin of the entire penicillin ring structure is known, Lvith the exception of the thiazolidine nitrogen atom. T h e sulfur atom, carbon and hydrogen atoms of the p-lactone: and side-chain nitrogen atoms come from L-cysteine. T h e penicillamine moiety is synthesized from valine, but i t is not known whether D- or r-valine is used. Xothing is known about the intermediates (81;). Arnstein and S l a r gareiter (8K)attempted to produce neir pencillins using substituted cystines, n i t h o u t success. Derivatives of 13.8dimethyllanothionine and DL-penicillamine \vere not active as precursors ( 7 K ) . T h e biosynthesis and properties of 4benzylphenouymethyl penicillin ivere described. At p H 3 the neiv product was more stable than penicillin G or V, but blood levels were only about half those of penicillin V ( 4 8 K ) . A particularly interesting finding was announced from the Beecham Research Laboratories in England ( 7 0 K ) . I n the absence of acyl side-chain precursors
(phenylacetic
acid,
etc.) Penicillium 6-aminopenicillanic acid, the nucleus of the antibiotic. This is consistent with the view that the acyl side-chain introduction occurs in the last stage of the biosynthesis. T h e nucleus compound has previously been encountered in small quantities in broths. Further processing details, including ion-exchange recovery m a y be found in a recent patent (771;). Although 6-aminopenicillanic acid itself is essentially \vithout antibiotic activity, new pencillins are possible by using different acidic side chains. Perhaps another route to 6-aminopenicil-
chrjsogenum accumulates
Table X. Subject Production of L-glutamic acid by various microorganisms
Fermentative production of Lornithine (preliminary report) L-Lysine production using microbial auxotroph Fermentation process for production of L-lysine
Preparation of amino acids
Glutamic acid by use of microorganisms
Table XI. Subject Biosynthesis of riboflavin ; purine metabolism and riboflavin synthesis in Eremotheciitm ashbyti Riboflavin biosynthesis in Cnndida jnreri
Influence of sterilization on biosynthesis of vitamin Bs with E. ashbyii
Influence on p H on biosynthesis of vitamin B? with E. ashbyii using serum and skim milk Recovery of riboflavin for animal feed products Slaintenance of E. ashbyiz
Riboflavin production Riboflavin production
lanic acid is via side-chain cleavage from penicillin G .
Steroids T h e year 1958 brought several novel developments in the steroid drugs. Some were strictly chemical, some biochemical, and others combined both synthetic methods. These developments have probably caused a shift in production and marketing from the “wonder drugs” of only a few years ago to new candidates with improved pharmacological properties.
Studies on Amino Acids Results Screening tests with media containing carbohydrate and ammonia; highest level of L-glutamic acid was produced by new species of ‘Micrococcus (0.25 mole of acid from 1 mole of glucose) hlutant of above ‘Micrococcus produced Lornithine on gluocse, corn steep liquor, salts media; 0.36 mole L-ornithine produced from 1 mole of glucose Mutant strain of -14. glutamicus yielded Llysine when grown on glucose, mineral salts s o h . ; L-lysine yield as high as 29% from consumed glucose on weight basis Fermentation process for L-lysine using Escherichia coli which initially requires Llysine for growth under aerobic conditions. After growth p H adjusted to 7.28.4 with KHIOH, rupturing cell walls; reaction then proceeds under anaerobic conditions Fermentation process utilizing Aspergillus terreus on a distiller’s solubles, glucose, corn steep liquor medium to produce L-glutamic acid, L-tryptophan, L-lysine Ctilizes Bacillus cereus to produce L-glutamic acid by fermentation
Ref. (7F)
(3F)
(W
Reports on Vitamin BZ Results Possible mechanism of conversion of purine into riboflavin Importance of Fe confirmed; 0.1-p.p.m. Fe levels inhibit in presence of ( “ 4 ) ~ SO?; xanthine strongly flavogenic; hypoxanthine and uric acid as effective as (NHa)lS04; while adenine and adenosine ineffective Sterilization of nutrient media half serum, half skim milk, 5YGsucrose may not exceed 1 hour at 1 2 0 ” C. Optimum p H after sterilization \vas 6.5-6.7 before sterilization 8.0 Recovery process for riboflavin from E. ashbyii fermentation Complete maintenance of flavogenic activity over 8 months obtained by freezedrying cultures previously frozen at -68’ C. in maltose or glucose solns. in water or serum Process involving E. ashbyii Process utilizing A s h b y gossjpii on medium containing starch or starchy material as sole source of carbohydrate; yields as high as 1200 -;/ml. obtained on 1.5% starch; diastatic mold added to convert starch to sugar at 72 hours
VOL. 51, NO. 9 , P A R T II
SEPTEMBER 1959
Kef. (Iff)
(-If€)
(511)
( 6 Hj
1091
UNIT PROCESSES
.____.__.______--_-_____________________------------
Of significance was a new chemical synthesis of aldosterone by a University of Sl’isconsin team (70L). T h e Upjohn group began chemical testing of Gamethyl-l7a-acetoxyprogesterone (Provera) for the prevention of miscarriage and premature birth and as a n oral contraceptive. I t is claimed to be 300 times more potent than drugs now in use for this purpose. ‘The Merck group (ZL,3L) reported a new series of highly potent anti-inflammatory steroids, the lba-methyl analogs of the cortisone family. Of these, 16amethyl,9a-fluoro-A1-hydrocortisone (dexamethasone, Decadron) is claimed to be the most active nonsalt retaining agent known. I t was more potent than 16a-methyl derivatives of prednisolone, hydrocortisone, and h f l u o r o cortisone ( 7 7L). Schering, too: entered the market with the same compound
(Deronil) after announcing synthesis of the drug (37L, 33L). Until the 1ba-methy1,la-fluoro derivatives were introduced the older antiinflammatory agents, cortisone, hydrocortisone, prednisolone, and prednisone, were in substantial competition with each other; 6-methylprednisolone (Upjohn, hledrol) a n d triamcinalone (16a-hydroxy,9a - fluoroprednisolone-Lederle: Aristocort ; Squibb? Kenacort) intensified competition factors. A simultaneous announcement (9L) from Merck and Schering teams introduced another new family of steroids, those with C, hydroxyls. T h e Schering group, by chemical means: prepared ?ahydroxycortisone and ‘a-hydroxyprednisone. T h e hIerck biochemical syntheses resulted in 70- and 7D-hb-droxylations. T h e 7a-hydroxylations are accomplished by a Fielminthosporium strain.
Table XII. Studies on Vitamin B12 (Cyanocobalamin) Subject Results Absorption of vitamin B12 on synthetic catVitamin B12 products and preparation ion exchange resin of low toxicity with subsequent high stability Production of vitamin BIZ from Medium for industrial production of vitamin B12 with Bacillus pro,bionici (Shermanii) propionic acid bacteria Production of vitamin containing Culturing of S. griseusforma farinosus NRRL nutritional composition and B-1354 in aqueous medium of soybean antibiotics meal, glucose, mineral salts, C o r + ; yields of BIZ were 0.2-0.6 gram/ml., along with antibiotic complex Marine plants and sapropels as Dtscription with yields sources of vitamin BIZ Production of vitamins B12 by enCulture gro\sn on Barker’s synthetic media richment culture of Methanubncproduced single factor called “met” in terium omclianskii presence of C o - 7 ; factors B and 2 also produced. B and 2 may be precursors of factor “met” and cyanocobalamin. When supplied with 5,6-dimethylbcnziminazole and Co culture produced only cyanocobalamin yields of 80 y/ml. Utilization of vitamin B1l factors I n presence of 5,6-dimethylbenziminazole, factors B, Z1,22,23 and part of X 4 conby enrichment culture of ’11. verted to cyanocobalamin omelianskiz Vitamin B12 Process utilizing Actinuplanes subtropicalis, new species from sediment of the Amazon River. Max. yield 5.80 mg./liter in 66 hours Biosynthesis of cobalamin analogs Only adenine hydroxocobalamin found in by Propionibacterium arabinosum medium containing glucose? yeast extract, salts, and fermented with P . arabinosum. Addition of 5,6-dimethylbenz. imidazole resulted in cobalamin containing this substance in nucleotide portion of molecule Preparation of new cobalamins Preparation of 4-bromo-6-methoxybenzimidazole hydroxycobalamin and salts using arabinosum fermentation with precursors Vitamin BIZformation in cultures Optimum Co(N’03)2 con. 0.05 mg. 7 0 of antibiotic-producing Actinomycetes Conditions for formation of vitaIn glucose medium B. megatherium stops growing rapidly owing to acid developmin BI2 by Bacillus mpgalherium ment. At a p H 4.5 most BIZleaves cells and passes into fluid; addition of chalk gives lower yield but larger number of cells. Organic acids added with glucose give better p H regime and raise vitamin yields Production of vitamin B,*-active Process and equipment involved product from sewage sludge I
1092
INDUSTRIAL AND ENGINEERING CHEMISTRY
I
Xvhile the 7P-hydroxy is introduced by a Cladosporium strain. Substrates successfully hydroxylated include progesterone, desex)-corticosterone: Reichstein‘s substance S, a n d cortisone. Another simultaneous announcement by Upjohn a n d Syntex (6L,72L) brought forth the 6a-fluor0 steroids. T h e U p john report indicated that 6a,9a-difluoroprednisolone was 400 times more active than hydrocortisone as a n antiinflammatory, but salt retention was measurable. ,4 powerful glucocorticoid (liver glycogen deposition activity) is 6a-fluorohydrocortisone, which does not cause salt retention ; ba-fluoroprednisolone, another potent anti-inflammatory: causes little or no salt retention. LVhile the several fluorination reactions a r r accomplished chemically, useful hydroxylations required in the fluorosteroids are probably introduced by the several fermentation procedures of the industry. T h e mechanism of biological hydroxylations remains of interest. T h e 7ahydroxylation in the biosynthesis of cholic acid from cholesterol involves a direct replacement of the 7a-hydrogen ( 5 L ) . T h e same opinion is expressed for other hydroxylations (26L). If deuterium is introduced into the 116 position, Rhizopus nigricans readily accomplishes an 1la-h>-droxylation without affecting the marked 11/3position (8L). Japanese workers have surveyed large numbers of fungi for steroid hydroxylations. Some 473 Aspergilli were screened for lla-hydroxy- or bfi,llPhydroxylations of progesterone (28L). Only 21 out of 284 Rhizopus strains hydroxylated progesterone. R. chinensis gave a 507, yield of 1 la-hydroxyprogesterone (4L). A Syncephalastrurn sp. oxidized progesterone to dihydroxyprogesterone in t\vo days (29L). Corynebacterium simplex can oxidize a 17-h>-droxyl to 17-keto; 19-nortestosterone was converted to 19-11or-L~androstene-3,17-dione j37L). Several Mucorales-Mucor parasiticus, XI. ,griseocyanus, a n d Helicostylum piriforme oxygenated progesterone: deoxycorricosterone? 11-deoxycortisol, and testosterone to the corresponding 14a-hydroxylated products (27L); Sp- (or 9a-) derivatives of deoxycorticosterone and of 1 l-deoxycortisol were encountered. A n excellent review is devoted to microbial transformations of steroids (78A). Some current patent literature is summarized in Table XV. By strain selection, Sarcina cereuisiae can be propagated to contain as much as 117, ergosterol (78L). Fermentation as a Unit Process Microorganisms. During the past year, several reports have been issued on microbial genetics, as applied to indus-
FERMENTATION strains. -4 patent was issued (35iU) relating to manufacture of new
trinl
strains of microorganisms in which reproduction is normally asexual and to processes in which strains of a microorganism with desired genetical factors are combined. These strains have genetical markers which enable the strains to be easily distinguished and possess complementary nutritional requirements or sensitivity to poisons. T h e strains are inoculated into the complementary media with or without the poisons, a n d the heterokaryon so formed, or its conidia, is grown in a t least a similar medium, thereby favoring multiplication of heterozygous nuclei in which the factors of the marked strain are combined. T h e marked strain is thereupon reisolated to form a stable recombinant strain possessing the different genetical factors of the originally selected strains. .4n improved method for isolating biochemical mutants of Streptomyces griseoJ a w s has been reported by Saito a n d Auxotrophic mutants Ikeda ( 3 8 M ) . ivere isolated by irradiating condiospores with ultraviolet light. Survivors (equivalent to 0.1 to 1% of the conidia irradiated) were incubated for a week, a n d a filtration technique was applied to the newly grown conidia to concentrate growth factor mutants. A review of 40 years of research o n the variability of microorganisms in the U.S.S.R. has been prepared by Imshenetski? (Z5M). Maintenance of Strain. Mainrenance of industrial strains of microorganisms continues to be a problem in industrial plants. T h e classic methods, such as storage o n slants, colony selection from plates, a n d sand or soil storage, lyophilization, spray drying, or slant storage under oil continue to be used: with company experience usual1)- dictating t h e method of maintenance. Several studies were published dealing with preservation of Serratia marcescens as a representative bacterium. Continuous freeze-drying was studied by Maister a n d others ( 3 7 M ) . S. marcescens was dried successfully to a moisture content of 1% suitable for satisfactory storage stability without excessive loss of viable cells. Viability was 8070 or higher in the final product. A later study ( 3 0 M ) investigated conditions most fairorable to the storage of S. murcescens. Results indicated that various factors were important, such as percentage of moisture, time of harvest, p H , concentration of cells, storage of dried material, a n d temperature of storage. Another study by Benedict and others ( 6 M ) describes in detail a carefully standardized procedure for preparing and drying cells of S.marcescens. T h e second international symposium
Table XIII. .4ntibiotic Aburamycin
Actinobolin .4lbomycetin .4lomycin Amaromvcin
.4midomycin
.\lthiomycin
Cellocidin
Croceoniycin
Desertomvcin
Durainycin Enteromycin Erythronicyin .4 Eurocydein Fermicidin
Flavofungin Grasseriom ycin Hygromycin B
Hydroxymycin 3iitomycin C hf ycobacillin
Producing Organism
New Antibiotics Ref.
Properties
Acidic yellow crystals, n1.p. 163 "-5 C . ; no N, S, halogen; solvent sol.; OID 24.5 in methanol; UV 0.01,VNaOH 234, Orally tolerated; 278, 316, 410 m p . toxic I.P., I\': Gram positive; C, 55.57; H, 7.54; 0, 36.89; similar to aureolic acid Streptomyces C13H20-22h7206; Gram positive and negative; active us. certain tumors Stre,btomyces T- C2jH14iVOi; a*; -48.7 (lscCHClp); 12-5650-1 15 gram positive; acidic; toxicity, mice, oral 1500 mg./kg. ; I P 500 mg./kg. Streptomyces sp. Antifungal in broth and mycelium S. flavochromo- Active Gram-positive and Gram-negative coccii; not active us. Gram-negative bacgenes teria, molds, yeasts. CnjHs90,N; m.p. 164.5 C. a z z 6.2 (C 15>,, ethanol) Streptomyes Antifungal, solvent extracted. Degraded to D-valine and 3,6-diisopropyl-2,5-diketomorpholine. Inhibits : C'stilago, Candida, Pvccinia, Horniiocium. Not inhibited : Oidium, Oosporu, .4spergillus, RhiroStreptomyces aburamicnsis
(661;)
+
(221;) (651;)
( IOiK) (43K)
+
Pus
LYhite cr).stalline; sol. dioxane, ethyl cellosolve, pyridine; insol. \iater, benzene ; sl. sol. lower alcohols: decomp. 120"160 ' C. ; L V (0.OSNHCI) 220-3,285-90, 220-3: 235, 335 m p ; C l ~ H i 4 N & 0 6 (C4H4O2N2),;absorbcd on carbon, eluted S. chibnensis with 807, methanol, concentrated, crysn. sp. tallized from hot aqueous methanol; m.p. 216" C . decomp. E;c7; 290 at 299 mp; active tuberculosis BCG 3 pg./ml: Structure : acetylene dicarboxamlde S.arabicus C22H1806; yellow needles: m.p. 325' c., decomp. cy2& - 32 i 4'; sublimes in vacuo 240 i 260" C . ; mg./kg. lethal for mice Ca3H60-6201&, m.p. 189 "-190 j' contains S. Jauogungini C-methyl, no N-methyl; active Grampositive bacteria 1-25 pg./ml. ; very toxic, cytotoxic ; coproduct of flavofungin S.cinnaniomeus Polypeptide from antibiotic complex ; active Gram-positive rods, some yeast and fungi; contains 9 amino acids, 2 with S S. albireticuli C s H 8 0 s N 2 ; contains -COOH and -COCHa; UV max. 300-320 m p ; a': 0 " ; L D =~ 1.3 ~ mg./lO gram mouse Separated from erythromycin B by recrysS.eryythrcus tallization in nitromethane S. abkilecticuli Antifungal ; yellow, amorphous powder ; UV 318, 332, 380 m p ; no S, halogen. LDjo = 0.22 mg./lO gram mouse I P S. giiseolus ClrHplOJV: 0'; 52.3; m.p. 96.8" C., LDjo = 180 mg./kg. mouse IV; 2 mg./ kg. rat : active us. yeasts and Trichomonas vaginalis, 0.05-0.2 mg./ml. S. flaaojutigini Crystalline antifungal isolated from mycelLDSoI P = 25 mg./kg. ; nontoxic oral or SQ Active us. jaundice virus; mol. wt. hydroS. griseolaoendus chloride 610. Recovery via carbon absorption, alumina chromatography S. h,vdroscopicus Coproduct of hygromycin. Polyhydroxy S . one K-methyl, no base C I S H 2 8 N 2 010, 0-methyl or C-methyl; Gram-positive and -negative activity, 6-100 y/ml. Streptomyces Similar to streptomycin and neomycin; differs by having pseudoneomine; low toxicity S. caespitosus C ~ ~ H ~ l N 1 p OUV 1 g ; 216, 360, 560 mp. Exhibits delaved toxicitv Bacillus subtilis Mol. wt. 1800; 11.9% K,no S. halogen: ninhydrin-negative cyclic polypeptide. Acid hydrolysis yields aspartic acid, glutamine, serine, alanine, tyrosine, leucine, proline
+
(941;
(109K
(7011;)
(85A)
(841;) (251;)
(82h.)
(7041;)
( 7OOh.) ( 9OK ) (SOK)
(471;) ( 7051;)
(58K)
(Continued)
VOL. 51, NO. 9, PART II
SEPTEMBER 1959
1093
~
Table XIII. Antibiotic Mycospocidin
Producing Organism S. bobiliae
Nymbomycin Paromomycin
5'. rimosus
forma paromomycinus Streptomyces
Phytoactia, phytostreptin Pyoluteorin Pseudomonas aer uginosa Ramycin
Mucor ramannianus
Raromycin Teruchiomycin
Streptomyces S. eurodicus
Toyokainycin
S. toyokaenris
Tubercidin
Streptomyces
\'i
L't os i n
S. olivochromogenous
Properties ( C ~ U H ~ Z N Z Owhite ~ ) ~ ;crystal, decomp. 233-4' C. Active us. spore former and acid-fast bacilli. LDbo = 1-2 mg./kg. I P C16HlaT\T204. Highly insoluble antiphage, Streptomyces, Gram-positive bacteria C&g- iiN02 Polypeptides ; prepared for plant disease control CIIH703NC12; m.p. 174"-5 C., decomp. UV255,310 mp; mol. wt. 268; structure given Optically inactive, unsaturated hydroxy acid; m.p. 158 '-1 60 O C. ; pK 4.6 ; mol. wt. 478; no UV Tumor inhibitory C28H43N010;needle crystals; m.p. 202 '4" C. decomp. Cl2HlrNaOa; UV max. 230, 279, 339 mp; LDloa IV = 20 mg./kg. Active hi.tuberculosis, Candida albicans C11HlaN404; UV in 0.001 NHC1: 225, 270 M . tuberculgsis Colorless plates; m.p. 142.5" C., 0'2 80.5; C27H4009N2; antifungal; LDju I P
+
=
S-948
S-1008 Streprovitacin A and B L.A. 7017 PA-128 PA-1 47
n.8 mu ...D ' ,/ku --0-
Streptomyces sp. May be identical with echinomycin of Carboz; C-7H~oOgKaS, m.p. 237"-8' C. ; 02,3 -308 in CHCla; Ba(OH)2 reflex yields 2-quinoxaline carboxylic acld, NH3 Streptomyces sp. C2gH380;N&; m.p. 209 '-16' C., a*: -282 ; Ba(OH)2 reflex yields 2-quinoxaline carboxylic acid C l j H 2 3 N 0 5 ;isomeric compounds, used on experimental tumors Streptomyces sp. Gram-positive bacteria; yellow-green PO\ nil.. S. viridofaciens Fermentation media for chloro-
,'
tetracycline
Recovery of tetracycline as organic salts: citrate, tartrate, ascorbate Benzyl-penicilloate salts of tetracy c1in e Chloride-deficient medium, especially low chloride soybean meal, for tetracycline fermentation
Biological Conversions of Steroids Microorganism Pycnospium Stachylidium sp. Dolhichiza ferruginosa Curcularia sp. .Vocardia sp. Yeasts Cyhdrocephalum aureum Corynebacterium simplex Trichoderma viride Hendersonia herpotricha, M.'ojnoLeicia graminis Fusarium solani, F. caucasium, Rhizopus
Ref. ( I L , 1 7 L , 22L) (76L) (3UL) (35L) (34LI
(7L)
(36L) ( Z L , 32L I
172 ( 73L)
sumis
Streptomyces roseochromogenes S.uiridis, s. oiiuaecus, S.argenteohs C~lindrocarponrodicola, F. jaoonicuni var. enaijorme Trichothecium roseum, Leptosphaeria macul a m , Cucurbitaria laburni, Acrospeira leisis, Lophotrichus hartinii, Melanospora parasitzca, Thieiavia terricola R. nigricans Cunninghameila blakesleeana
VOL. 51, NO. 9 , PART II
(23L'l ( 24.L )
(Z5L)
( 75L
1)
(74L)
(74L)
SEPTEMBER 1 9 5 9
1095
now manufactured by Staplex Co.? Brooklyn, N. Y . T h e unit samples large volumes of air for microorganisms or particulate matter. Disposal of Fermentation Wastes. Disposal of wastes from a fermentation plant still poses numerous problems. A two-stage filter operation a t the Upjohn waste treatment plant, which handles wastes from antibiotics and steroids, was described by Tompkins ( 4 5 M ) . Problems such as those occasioned by antifoam greases or oils from the spent fermentation broths appeared to d o the greatest h a r m to the system. An interesting feature of the plant is that during M a y to late fall, about 100,000 gallons per day of unchlorinated effluent is sprayed on plant lawns, keeping lawns green, conserving water and chlorine, and reducing B.O.D. input to the waste stream. Cushman and Hayes ( 7 3 M ) have described a pilot plant study of multistage trickling filter treatment of various blends of sanitary sewage, chemical process wastes, and fermentation spent broths. First-stage B.O.D. removal exceeded 807, a t loadings of from 400 to 3500 pounds,’day/acre foot. Secondary filter removal exceeded GOY0 a t loadings of 500 to 300 pounds/day/acre foot. A report by Paradiso and How? (36.M) deals with sewage treatment a t the Tippecanoe plant of Eli Lilly & Co. where spent antibiotic broth is handled. Jeffreys ( 2 7 M ) has patented a simultaneous aerobic and anaerobic composting process which may be of interest in composting of fermentation residues as a means of disposal. Recovery of Fermentation Products. During the past year a novel ion exchange method for isolating streptomycin was disclosed by Barrels and others ( 3 M ) . T h e basic idea is an ion exchange isolation of streptomycin from the whole fermentation broth, eliminating the previous and expensive filtration step. I n addition, a purer product was claimed with higher over-all yields. T h e process is now in use a t E. R. Squibb & Sons. Table XVI lists some recent patents on recovery of fermentation products.
Table XVI.
Fermentation Product Recovery
Product Bacitracin Basic antibiotics Neomycin Nystatin Riboflavin Streptomycin and dihydrostreptomycin Vitamin B12
1096
Ref. ( 11ZK) (4 0 ) ~
Bibliography (1.4) Abe, S., “Penicillia Atlas of Microorganisms,” Shuppan Co., Tokyo, 1957. (2A) Arima, K., “Yeasts,” W. Junk, The Hague, 1957. (3A) Asai, T., .’l‘ippon N8gei-kagaku Kaishi 21, A25 (1957). (4A) Aylett, M., “Encyclopaedia of Home -Made Wines,” Odham Press, London, 1957. (5A) Babbitt, H. E., Baumann, E. R., “Sewerage and Sewage Treatment,” 8th ed., Wiley, New York, 1938. (6‘4) Buffton, .4.W. J., “Industrial and Economic Microbiology in North America,” Gt. Brit. Overseas Tech. Rept. No. 4 , 1958. (7AJ Clifton, C. E., others, “Annual Review of Microbiology,” vol. 11, Annual Reviews Inc., Palo Alto, Calif., 1958. (8A) “Continuous Cultivation of Microorganisms,’’ publishing House of Czechslovak .4cademy of Sciences, Prague, 1958. (9.4) Dunn, C. G., Econ. Botany 12, 145 (1958). (10A) Federal Trade Commission, “Economic Report on Antibiotic Manufacture,” U. S. Govt. Printing Office, Washington, D. C., 1958. (11A) Galanti, h?., Ann. Gemblonx 63, 33 (1957). (12.4) Guharay, S. V., “.Intibiotics. A Symposium,” Sree Sarawaty Press, Ltd., Calcutta, 1958. (13A) Hale, L. J., “Biological Laboratory Data,” Wiley, New York, 1958. (14.4) Imhoff, K., others, “Sewage Treatment,” 2nd ed., Wiley, New York, 1957. (15A) Lerner, I. M., “The Genetic Basis of Selection,” Wiley, New York, 1958. (16A) Luck, J. M., ed., “Annual Review of Biochemistry,” vol. 27, Annual Reviews, Palo Alto, Calif., 1958. (17A) McCabe, J., Eckenfelder, W’. N., Jr., “Biological Treatment of Sewage and Industrial Wastes,” vol. 11, Reinhold, New York, 1958. (18A) Nord, F. F., “Advances in Enzymology,” vol. 20, Interscience, New York, 1958. (19.4) Penau, H., J o u m d s $harm. franc., Paris; Coni. sot. tech.pharm. [Pub. in Tech. pharm. (Paris) 5, Yo. 4 (1958)l. (20A) “Proc. Intern. Symposium on Enzyme Chemistry, Tokyo-Kyoto, 1957,” Academic Press, New York, 1958. (21.4) Roman, W., ed., “Yeasts,” Academic Press, New York, 1958. (22A) Salvato, J. A., “Environmental Sanitation,” Wiley, New York, 1958. (23’4) Shoppee, C. W., “Chemistry of the Steroids,” -4cadernic Press, New York, 1958. (24.4) Steel, R., ed., “Biochemical Engineering,” Macmillan, New York, 1958. (25.4) Stodola, F. H., “Chemical Transformations by Microorganisms,” Wiley, New York, 1958. (26.4) Stodola, F. H., “Source Book on Gibberellin 1828-1 957,” L. S. Dept. Agr., ARS 71-11, 1958. (27’4) Waksman, S. .4.,ed., “Neomycin, Its Nature and Practical Application,” Williams & rVilkens, Baltimore, Md., 1958. (28.4) Whitmarsh, J. hi., “British Fermentation Industries,” Pitman, London, 1958.
(26iM)
( 17M, 4 6 M ) (14M, 32M) (49M, 5 0 M ) ( Z M ,5 7 M )
Acetone-Butanol (IB) Anderson, P. C., Rapp, Janet L. C. (to Feed Service Corp.), U. S. Patent 2,808,332 (Oct. 1, 1957). (2B) Baba, T., Hiroshima Daigaku K6gaku6u Kenju H8hoku 7, 51 (1958).
INDUSTRIAL AND ENGINEERING CHEMISTRY
(3B) Ibid., p. 59. (4B) Hongo, M., J . Agr. Cheni. Sac. ( J a j a n l 31, Item 7, p. 485 (1957). (5B) Ibid., Item 1, p. 615. (6B) Ibid., Item 2, p. 618. (7B) Ibid., 32, Item 2 (1958). (8B) Karsch, W., Schoedler, K. (to Tornescher Hefe G.m.b.h.), Ger. Patent 941,184 (April 5, 1956). (9B) Nemoto, S., J . Agr. Chem. Sac (Japan) 32, Item 5, p. 283 (1958). (10B) Perdomo, E. V., Inst. Cubano Inuest. Technol. Ser. estud. trabajos inuest., N o . 3, 32 11958). (11B) Yarovenko, V. L., others, Spirtouuya Prom. 24, 5 (1958). Citric, Itaconic, Fumaric, a n d Gluconic Acids (1C) Emiliani, E., Retamar, J. .4, Rev. f a c . ing. quim. (Uniu. Nacl. litoral, Santn Fl, Arg.) 26, 18 (1957). (2C) Faerber, G., others, Ceskuslou. mikrobioi. 3, 133 (1958). (3C) Kovats, J., Prremyst Spozywrzy 8, 303 (1958). (4C) Leopold, H., h‘ahrung 2, 140 (1958). (5C) Lubowitz, H. R., La-Roe, E. G. (to National Distillers Corp.), U. S. Patent 2,861,922 (Nov. 25, 1958). ( 6 C ) Matkovics, B., Kovacs, E., ’Vaturwissenschaften 44, 447 (1957). (7C) Ozaki, A , , others, HakkB Kyokaishi 15, 287 (1957). (8C) Pfeifer, V. F., others, IND. ENG. CHEM.50, 1009 (1958). (9C) Raina, P. N., Ramakrishnan, C. V., Current Sei. (India)26, 285 (1957). (1OC) Royal Norwegian Council Sci. Ind. Research, Brit. Patent 295,401 (May 21, 1958). (11C) Ueda, K. (to Chichikuma Miso Brewing), Japan Patent 6799 (’57) (.4ug. -7,
A/
1.
(12C) Ueda, K., others, HakkB Kyokuishi 15, 437 (1957). (13C) Zaheer, S. H., others, Prods. pharm. 13, 201 (1958). Gibberellin (1Di Borrow, A , others (to Imperial Chem. Ind., Ltd.) U. S. Patent 2,865,812 (Dec. 23, 1958). (2D) Kavanagh, F., Kuzel, N. R., J . Agr. Food. Chem. 6 , 459 (1958). (3D) Lu, K. C., others, Nature 181, 189 (1958). (4D) Manzelli, M. A,, Division of Agricultural and Food Chem., 134th Meeting, ACS, Chicago, lll., September 1958. (5D) Owades, J. L., Chiano, G., A m . Brewer 91, 39 (July 1958). (6D) Phinney, B. O., West, C. A,, Division of Agricultural and Food Chem., 134th Meeting, ACS, Chicago, Ill., September 1958. (7D) Wittwer, S. H., Bukovac, M. J., Mich. State Univ. Agr. Expt. Stat. Quart. Bull. 40, 352 (1957). 0-Carotene (1E) .4nderson, R. F., others, J . Agr. Food Chem. 6 , 543 (1958). (2Ej Braithwaite, G. D., Goodwin, T. W., Biochem. J . 67, 13P (1957). (3E) Ciegler, A,, others, 58th Meeting, SOC.Appl. Bacteriol., Chicago, April 27May 1, 1958, Abstracts, p. 13. (4E) Ibid., p, 13. (5E) Deufel, R . D., Clark, F. M., Ibid., p. 13. (6E) Glover, J., Shah, P. P., Biochem. J . 67, 15P (1957). (7E) Reichel, L., Wallis, M., Naturwissenschaften 45, 130 (1958).
FERMENTATION Amino Acids (1Fj Kinoshita, S.,others, J . Gen. .Microbiol. 3. 193 (19571. .._. ,. (2F) Zbzd., 3, p. 276. O F ) Zbzd., 4, 128 (19581. (4F) Kita, D. A , , Huane, H. T. (to Chas. Pfizer & Co.); L.S.*Patent 2,841,532 (July 1, 19.58). (5Fi Pfizer, Chas., 8: Co., Brit. Patent 795,178 (May 21, 1938). t6Fj .Tada, S., others (to .Ajinomoto Co.), Japan. Patent 9393 (’57) (Nov. 7,. - I
~
~
Dextran (1G) Kooi, E. R. (to Corn Products Co,) U.S.Patent 2,833,695 (PILay 6, 1958). (2Gi Leach; J. G., others, P/zytoflaI/iolo:y 4 i , 113 (1957). (3G I iilly, V. C., others, -4flpi. .Microbial. 6 , 1b5 (1958). (4Gl Novak, L. J., Stoycos, G. S. (to the Commonwealth Eng. C O . JU. , S. Patent 2,841,578 (July 1, 1958,~. (5Gj Rogovin, 5 . P., others, Division of ;Igricutural and Food Chem., 134th Ileeting, ACS, Chicago, September 1958, (6Gj Shinoda, X., J . Gen. .1lz~robzol. 4, 8 5 (1958). Zief, h l . , Stevens, J . R. (to Baker C.hemica1 C o . ) , U . S. Patent 2,807,610 (Sepr. 24, 19573.
(TF)
Vitamin B2 ( l H ) Brown, E. C., others, Biochenz. J . 68, 40 (1958). (2H Goodwin, 1‘. \V.: >lcEvoy, D., Zbid., 67, 16 (1957). (3H j Hendrickx, H., DeVlceschauwer, A , , Mededcl. Landbouwhogeschool e n OproeXzngsts. Stuat. Gent. 23, 65 (1958). (4H i Hendrickx, H., DeVleeschauwer, A,, MilchwzssenschaJt 13, 249 (1958). (5H)Holdsworth, H., Chern. 3 Ind. (London) 1958, 284. (6H) HoStalek, Z., J . Gen. .Mzcrobiol. 17, 267 (1957). l7H) Speedie, J. D., Cheni. @ Ind. (Lond o n ) 1958. 283. (8H)’Szu&ki, S. A. (to .American Cyanamid Co.1: Can. Patent 559,187 (June 24, 1958). Vitamin BIB [lJ) Boushard, E. F., others (to Chas. Pfizer 6r Co.), U. S. Patent 2,830,933 (.April 15, 1958). (25) Goncharoira, V. I., others, hZicrobzologzya (U.S.S.R.) 27, 224 (1958). (35) Hall, H. H., Benedict, R . G. (to U. S. X., Secy. of Agr.), U.S. Patent 2,846,310 (Aug. 5, 1958). (45) Kutseva, L. S., Bukin, V. N., Proc. Acad. Sci. U.S.S.R., Biochem. Sect. 115, 195 (1957). (5J) Neujahr, H. Y . , Callieri, D. A.: Acta. Chem. Scand. 12, 1153 (1958). (6J) Ibzd., p. 1167. (7J) Pascual, J., Span. Patent 236,386 (Sept. 30, 1957). (85) Perlman, D. (to Olin hfathieson Chemical Co.), L. S. Patent 2,842,540 (July 8, 1958). (95) Perlman, D., Barrett, J. ?*I.,Can. J . .Wicrobiol. 4, 9 (1958). (10J) Surikova, E. I., Popova, L. A,, .Vikrobzologiya (U.S.S.R.) 26, 432 (1957). ( l l J ) Ushakova, V. I., Doklady Akad. 1Vauk U.S.S.R. 122, 520 (1958). (125) Wolnak, B., Zinn, R . E. (to Sewage Comm. City of Milwaukee, Wis.), U. S. Patent 2,833,692 [May 6, 1958). Antibiotics (1K) American Cyanamid Co., Austral. Patent Appl. 27,952 (May 21, 1957).
(2K) .American Cyanamid Co., S. Afrlcan Patent -4ppl. 513/58. (3K) .\nliker, R., Gubler. K , Helu. Chznz. Acta 40, 1768-’2 (19581. (4K) A m . -V. 1.~ .had. 5’61. 7 6 . 17-408 (1958). (5K) Anzai, K., J . ilntzbioiics (Japan) 10A, 201-4 (1957). (6K) hrishima, M., others, AVzppon .\-dgeikagaku Kaislzi 30, 407-9 (1956). (7K) Arnstein, H. R. V., Clubb, hl. E., Biochem. J . 68, 528-35 (1958). (8K) Arnstein, H. R. V., hlargareiter, H.: Ibid., 68, 339-48 (1958). (9K) Ball, Stanley, Hughes, I. W. (to Glaxo Laboratories, Ltd.;, Brit. Patent 799,053 (July 30, 1958). (10K) Batchelor, F. R., others, LVature 183, 257-8 (1959). (1 1 K ) Beecham Research Laboratories, Belg. Patent 569,728 (July 25, 1958,. (12K) Bell, hl. R., others, J . A m . Chem. Sor. 80, 1001 (1958). (13Ki Berger, J., others, Exfleritntia 13, 434-6 11957). (14K) Biserte, G., Dautrevaux, M., Bull. suc. chirn. b i d . 39, 795412 (1957). (15K) Boothe, J. H., others, J . Am. Chem. SUG.81, 1006-7 (1959); Chenz. Ene. Il‘ews 39, 36 (March 9, 1959). (16K) Bristol Laboratories, Inc., Brit. Patent 794,738 (May 7, 1958). (17K) Brock, T . D., Sakolski, W. T., Antibiotics €3 Chemotherapy 8, 631 (1958). (18K) Chapmann, D. D., Tarbell, D. S., J . A m . Chem. Soc. 80, 3679-82 (1958). (19K) Charney, Jesse, Fisher, W. P. M.: Tytell, A. A. (to Merck & C o . ) , Can. Patent 554,991 (March 25, 1958). (20K) Chtm. Eng. .Vews 36, 24 (July 28, 1958). (21K) >bid., 37, 27 (Jan. 26, 1959). (22K) Chem. U’eek 83, p. 65 (Oct. 18, 1958). (23K) Ibid., p. 66. (24K) Cheney, L. C., Gottstein, W. J. (to Bristol Laboratories). U. S. Patent 2,824,877 (Feb. 25, 1958). (23K) Clark, R. K., Jr., U. S. Patent 2,823,203 (Feb. 11, 1958). (26K) Cron, M. J., others, J . A m . Clzem. SOC. 80, 2342 (1958). (27K) Ibid., pp. 4741-2. (28K) Cross, 4 . D., Stanley, D. S., J . A m . Chem. Soc. 80, 3682-6 (1958). [,29K) Denison, F. W., Jr., Friedland, W. C., Peterson, M. H. (to Abbott Laboratories), U. S. Patent 2,834,714 (May 13, 1958). (30K) Doskocil, J., others, J . Gen. Microb i d . 18. 302-14 11958). (31K) Dutcher, J. ‘D., J . Bid. Chem. 232, 785-95 (1958). 132K) Eble, T. E., others. ..lntibiotzcs €3 Chemotherapy 8, 627 (1958). (33K) Eisenbraun, E. J., others, J . Am. Chem. Soc. 80, 1261-2 (1958). 134K) Els, H., others, Ibid., 80, 878-80 11958). (35K) Ensminger, S. W., others (to Chas. Pfizer & C o . ) , U. S. Patent 2,831,878 (April 22, 1958). 136K) Ettlinger, L., others, Helu. Chim. ilcta 41, 216-19. 220-8 (1958). (37K) Freanev. T. E. (to Commercial Solvents Co;p.), U . S. Patent 2,828,245 (March 25, 1958). (38K) Friedland, W. C.. Dension. F. W.. Jr., Peterson, M. H. (to .4bbott ’Labora: tories), Zbid., 2,833,696 (May 6, 1958). (39K) Friedman, I. J., Martin, E. G., Taylor, R . J., Ibid., 2,827,417 (March 18, 1958). (40K) Fryth, P. W., others, J . A m . Chem. SUG.80, 2736-40 (1958). (41K) Hagemann, G., others, Ann. #harm. f r a n ~ 16, . 585 (1958).
(42K) Harman, R. E., others, J . Am Chem. Soc. 80, 5173-8 (1958). (43K) Hata, F., Sano, T., Japan. Patent 449 (‘57) (Jan. 25). (44K) Heinemann, B., others, Can. Patent 556,229 (.April 22, 1958). (45K) Ibzd., 566,663 (Nov. 25, 1958). (46K) Kaplan, A t . A., others, Antibiotzcs & Chemotherapj 8 , 491-5 (1958). (47K) Kanin, C. M.,Finland, M., ‘Vortheast. J . M e d . 559, 999-1004 (1958). (48Kj Lein, J., others, Antibzotzcs & Chemotherapy 8 , 99--103 (1958). (49K) Lepetit, S.p..k, Brit. Patent 787,791 (Dec. 17, 1957). (50Kj Zbid., 799,051 (July 30, 1958). i(51K) Lindner, Fritz, others (to Farbwerke-Hoechst A-G.), Ger. Patent 952,632 (Xov. 22, 1956). (52K) McCormick, J . R. D., others, J . Am. Ciiem. Soc. 79, 4561 (19571. (53Kj Zbid., 80, 5572 (1958). (54K) Ibzd., pp. 6460-1 (1958). (55Kj SlcCormick, J. K. D., others, S. African Patent Appl. 881/58. (56K) McCormick, J . R. D., others, U. S. Patent 2,878,289 (March 17, 1959). (57K) LlcNally, 3. G., Tarbell, D. S., J . Am. Chem. Soc. 80, 3676-7 (1958). (58K) htajumdar, S. K., Rose, S. K., Nature 181, 134-5 (1958). (59K) Malek, P., others, Zbid., 181, 706-7 (19583. (60K) Mann, R. L., others, J . Am. Chem. SOC.80, 2714 (1958). (61K) Merck & Co., Inc., Brit. Patent 791,731 (March 12, 1958); Can. Patent 558,021 jhfav 27, 1958). (62K) hfinieri,’P. P., others, U. S. Patent 2,866,738 (Dec. 30, 1958). (63K) hliyake, A,, Japan. Patent 6149 (‘57) (Aug. 10). (64K) Nakamura, S., others, J . Antibiotics ( J a p a n ) 10A, 248-53 (1957). (65K) Nakazawa, K., others, ll‘zjpon h’ogeikagaku Kaishi 32, 321 (1958). (66K) Niedercorn, J . G., Can. Patent 566,331 ( S o v . 18, 1958). (67K) Nishimura, H., others, Japan. Patent 3049 (’57) (May 21). (68K) Nishimura, N., others, J . Antibiotics 1Jabanl 10A. 205-12 (19571. (65Kj Nlshiya: M., Nishimura, H., Japan. Patent 7750 (’57) (Sept. 17). (70Kj Olin Mathieson Chemical Corp., Brit. Patent 796,493 (June 11, 1958). (71K) Parke, Davis & Co., Zbid., 797,568 (July 2, 1958). (72K) Patrick, J. B., others, J . Am. Chem. SOG.80, 6688-9 (1958). (73K) Pfizer. Chas.. & Co.. Brit. Patent . 787,895 (Dec. 18,‘1957). (74Kj Pridham, T. G., others, Appl. Mtcrobiol. 6 , 52-79 (1958). (75K) Rao, K. V., Lynch, J. E., Antibiotics 3 Chemotherapj 8 , 437-40 (1958). (76K) Rhodes, Alan, others (to Glaxo Laboratories. Ltd.1. ,, Brit. Patent 784,618 (Oct. 9, 1957). (77K) Schmitz, H., others, J . Am. Cliem. SOC.80, 2911-12 (1958). (78K) Schneirson, S., Amsterdam, D., ’Tuture 182, 56-7 11958). (79K) Sensi, P., others, Antzbiottcs €3 Chemotherapy 8, 241-4 (1958) (80K) Zbzd., 7, 645-52 (1958). (81K) Shaposhnikob, V. N., others, Doklady Akad. Nauk S.S.S.R. 121, 366-9 (1958). (82K) Shibata, M., others, Japan. Patent 4995(’56) (June 25). (83K) Shibata, M., others, Ibzd., 9396(’57) (&-ob. 7 ) . (84K) Zbzd., 4994(’56) (June 25). (85K) Shotwell, 0. L., others, J . A m Chem. Soc. 80, 3912-18 (1958).
VOL. 51, NO. 9, PART II
’
SEPTEMBER 1959
1097
UNIT PROCESSES (86K) Snell, J. F., U. S. Patent 2,843,526 (July 15, 1958). (87K) Stephens, C. R., others, J . A m . Chem. Sac. 80, 5324-5 (1958). (88K) Sternbach, L. H., others, Ibid., 80, 1639--47 (1958). (89K) Suhadolnik. R . J.. Chenoweth. R . G.. Ibid.. 80.’4391-2 ( i 9 5 8 ) (9OK) Sumiki, Y:, others,’ Japan. Patent 6296(’57) (Aug. 15). (91K) Suzuki, S., others, J . Antibiotics ( J a p a n ) 11, 81-3 (1958). (92K) Ibid., pp. 84-6. (93K) Szumski, S. A,, Can. Patent 560,576 (July 22, 1958). (94K) Taber, W.A , , Vining, L. C., Can. J . Microbiol. 3, 953-65 (1957); Can. J . Chem. 35, 1109-16 (1957). (95K) Takeda, R.: J . A m . Chem. SUC.80, 4749-50 (1958). (96K) Takeda Pharmaceutical Industries, Belg. Patent 553, 388 (March 31, 1957). j97K) Tanaka, h-., others, J . Antibiotics ( J a p a n ) 10A, 189-94 (1957). (98K) Tatsuoka, S., others, Phurm. Bull. ( T o k y o ) 5, 343-9 (1957). (99K) Umazawa, H., others, Japan. Patent 850(’58) (Feb. 13). (100K) Uri, J.: Bekesi, I., ,Tuture 181, 908 (1958). (lOlK) Ibid.,182, 401 (1958). (102K) Van Dijck, P. J., DeSomer, P., J . Gen. Microbiol. 18, 377-81 (1958). (103K) Vandeputte, John, heuser, L. J. (to Olin Mathieson Chemical C o . ) ,U. S. Patent 2,847,471 (Aug. 12, 19583. (104K) Wada, S., others, Japan. Patent 9399(’56) (Nov. 2). (105K) Wakaki, S.,others, Antibiotzcs E Chemotherapy 8, 228-40 (1958). (106K) Weidenheimer, J. F., Kitter, Lawrencc (to American Cyanamid C o . ) ,U. S. Patent 2,820,824 (Jan. 21, 1958). (107K) LVoznicka, \V., others, -Wed. Dosciondetalnai Mikl-obiol. 9, 293,441-5 11957). (108Kj ’Yabuta, T., others, U. S. Patent 2,837,510 (June 3, 1958). (109K) Yamaguchi, H., others, J . ilntibzotics ( J a p a n ) 10A, 185-94
(112K)‘Zinn, Eli, Chornock, F. 11;. (to Commercial Solvents C o r p . ~ U. , S.Patent 2,834,711 ( M a y 13, 1958). Steroids (1L) American Cyanamid Co., Brit. Patent 789,862 (Jan. 29, 1958). (2L) Arth, G. E., others, J . A m . Chern. Sac. 80, 3160-1 (1958). (3L) Ibid., pp. 3161-3. (4L) Asai, T., J . Gen. Appl. Microbiology ( T o k y o ) 4, 63-6 (1958). (5L) Bergstrom, S., others, J . A m . Chem. Sac. 80, 2337-8 (1958). (6L) Bowers, A , , Ringold, €5. J., Ibid., 80, 4423-4 (1958). (7L) Camerino, Bruno, Vercellone, Alberto (to Societa Farmaceutici Italia), U. S. Patent 2.866.736 iDec. 30. 1958). (8L) Carey, E. i., dthers; J . A m . Chem. SOC.80, 2338 (1958). (9L) Chem. Eng. 1TeeZc.r 36, 54 (March 31. 1958). 36, ,...-\ 17 (April 21, 1958); 43
1098
(16L) Davisson, J. W. (to Chas. Pfizer & C o . ) , U. S. Patent 2,830,937 (April 15, 1958). (l7L) Davisson, J. W., Kita, D . A , , Routten, J . B. (to Chas. Pfizer & Co.), Ibzd., 2,848,370 (Aug. 19, 1958). (18L) Dulaney, E. L. (to Merck & C o . ) , Ibid., 2,817,624 (Dec. 14, 1957). 119L) Dulanev. E. L.. McAleer. W.J. (to
American Cvanamid’ C o . ) . U. S. Patent 2,789,940 (.April 23, 1957j. (23L) Fried, Josef, others (to Olin Mathieson Chemical Co.), Ibid., 2,855,343 (Oct. 7, 1958). (24L) Ibid., 2,855,410 (Oct. 7 , 1958). (25L) Fried, Josef: Thoma, R. W.(to Olin Mathieson Chemical Co.), Ibid., 2,868,694 (Jan. 13, 1959). (26L) Hayano, hI.; others, J . , l m . Chem. Sac. 80, 2336-7 (1958). (27L) Hershberg, E. B. (to Schering Corp.), U. S.Patent 2,833,797 (May 6, 1958). (28L) Izuka, H., others, J . Gen. Appl. Microbiol. ( T o k y o ) 4, 67-78 (1958). (29L) Ibid., pp. 79-84. 130L) Kita. D . A,. Shull. G. hf. (to Chas. ‘ Pfizer a’ Co.). U. S.’Patent 2,830,936 (Apri 115, 1958). (31L) I(ushinskv, S , J . Bid. Chem. 230, 31-9 (1958). (32L) Kobile, Arthur (to Schering Corp.), U. S.Patent 2,837,464 (June 3, 1958). (33L) Oliveto. E. P.. others. J . -4m.Chem. . Soc. 80, 4428 (1958). (34L) Pfizer. Chas.. & Co.. Brit. Patent ‘ 789,363 (Jan. 22,’1958). ’ (35L) Shull, G. hf., Bloom, B. hf. (to Chas. Pfizer & C o . ) , U. S. Patent 2,830,935 (April 15, 1958). (36L) Shull, G. M . , Routten, J. B. (to Chas. Pfizer 6: Co.), Ibid., 2,866,737 (Dec. 30, 1958). Fermentation
as a
Unit Process
(1M) Anderson, A. A . , J . Bacferioi. 76, 47 (1958). (2M) Baron, A. L., hfaxion, E. J. (to Chase Chemical Co.), U. S. Patent 2,861,025 (Nov. 18, 1958’1. (3M’i Bartels, C. R., others, Chem. Eng. Progr. 54, 49 (1958). (4M) Bartlett, XI. C., Gerhardt, P., Division of Agricultural and Food Chem., 134th Meeting, ACS, Chicago, September 1958. (5M) Belozerskii, A . N., Kulae\-, I. S.. Biochernistry (U.S.S.R.) 22, 27 1,1957). (6M) Benedict, R. G., others, A$$. -Wicrubiol. 6, 401 (1958). (7M) Bhuyan, B. K . , Johnson, M. J., J . Baderiol. 76, 376 (1958). (8M) Brierley, M. R., Steel, R., Appl. .Miwobiol. 7 , 57 (1959). (9M) Brinsberg, S. L., Grabovskaya, 0.Z . , Mikrobiologya 27, 407 (1958). (10M) Calderbank, P. H., Trans. Inst. Chem. Engrs. 36, 443 (1958). (11M) Carpani, R . E., Roxburgh, J. M., Can. J . Chem. Eng. 36, 73 (1958). (12bI) Chari, C. N., Hindustan Antibiotic Bull. 1 , 25 (1958). ( 1 3 W Cushman, J. R., Hayes, J. R . , Proc. 11th Ind. Waste Conf. (Purdue Univ., 1956, Purdue Eng. Extension Dept., Extension Ser. No. 91, 62 (1957). (1441) Dale, J. K. (to Commercial Solvents Corp.): U. S. Patent 2,815,341 (Dec. 3, 1957). (1514) Deindoerfer. F. H., Humphrey, A . E., Division of Agricultural and Food
INDUSTRIAL AND ENGINEERING CHEMISTRY
Chem., 134th Meeting, ACS, Chicago, September 1958. (16M) Deindoerfer, F. H., Humphrey, A. E., IND.ENG. CHEM.51, 809 (1959). (17M) Dutcher, 3 . D., Vandeputte, J. (to Olin Mathieson Chemical Corp.), U. S. Patent 2.865.807 iDec. 3. 19581. (18M) D\&rschack,‘R., Proc. Natl. Conf. Instrument Methods Anal., Paper No. .4157-6-3, Chicago, 1957. (19M) Dyr, J., Kuasny’ pr?imsjl 4, 112 119581. (2641) -Gallicchio, V., Gottlieb, D., .Mycologia 50, 490 (1958). (21hI) Gergle, R. C., others, Proc. Sci. Sect. Toilet Goods Assoc. 28, 1 (1957). (22M) Heden, C. G., S o r d . .Wed. 60. 1090 (1958). (23M) Hedtn, C. G., Malmgren, B., Intern. Congr. Microbioi., 7th Congr., Stockholm, 1958, Abstracts VII, p. 411. (24M) Hough, J. S.,Rudin, A. D., J . Inst. Brezoing 64, 404 (1958). (25M) Imshenetskii, A . A , , .\ficrobiologiya 26, 632 (1957). (26M) Jackson, W. G. (to Upjohn C o . ) , U. S. Patent 2,848,365 (Aug. 19, 1958). (27M) Jeffreys, G. A , , Ibid., 2,867,521 (Jan. 6, 1959). (28M) Konovalov, S. X., .2licrobiologija 27, 232 (1958). (29M) Luedeking, R., Piret, E. L., Division of Agricultural and Food Chem., 134th Meeting, ACS, Chicago, September 1958. j30M) Maister, H. G., others. Appl. hlicrobiol. 6 , 413 (1958). (31hI) Maister, H. G., others, IXD.EXG. CHEM.50, 623 (1958). (3251) Morehouse, .A. L. (to Grain Processing Corp.), U. S. Patent 2,822,361 (Feb. 4, 1958). (33M) Mukhamedzyanov, B. S., others, U.S.S.R. Patent 167,900 (Nov. 27, 1957). (34M) Pan, S.C., Division of Agricultural and Food Chem., 134th Meeting, ACS Chicago, September 1958. ( 3 j M ) Pelligrino, G., others (to National Research Div. Corp., London), U. S. Patent 2,820,742 (Jan. 21, 1958). (36M) Paradiso, S. M., Howe, R. H. L., Ind. Wastes 3, 101 (1958). (37M) Rhodes, A,, others (to Glaxo Laboratories, Ltd.), U. S. Patent 2,843,527 (July 15, 1958). (38M) Saito, H., Ikeda, Y., J . Gen. Appl. Microbiol. ( T o k y o ) 3, 240 (1958). (39hi) Sobotka, H., others (to U. S.Secy. of Kavy), U. S. Patent 2,844,515 (July 22, 1958). (40M) Solomons, G. L., Perkins, M. P., J . AppI. Chem. (London) 8 , 251 (1958,. (41M) Ibid., p. 445. (42M) Steel, R., Brierley, M. R., A$$/. .Vlicrobioi. 7, 51 (1959). (43M) Stefanye, D., Dews, J. N., U. S. Patent 2,865,816 (Dec. 23, 1958). (44M) Strohm, J., others, Division of Xgricultural and Food Chem., 134th Meeting, XCS, Chicago, September 1958. (45M) Tompkins, L. B., S f w a g e and Ind. Wastes 29, 1161 (1957). ( 4 6 M ) VandeDutte. J. (to O h Mathieson ’ Chemical Cbrp.),’U. S . Patent 2,832,719 (April 19: 1958). (47M) Wasilewski, Z . , Prremsyl. Spo3y~’cty 12, 7 (1958). (48hI) Yamada, K., Takahashi, J., J . Agr. Chem.Soc. ( J a b a n ) 32.Item1, p. 344(1958). (49M) Ziegier, F. (to Schenley Industries, Inc.)? U. S. Patent 2,857,375 (Oct. 21, 10581. (50hI) Ibid.,2,857,376 f,Oct.21, 19581. 151hI) Ziegler, r V . 51.(to American Cyanamid C o . ) , U. S. Patent 2.830,009 (.\pril 8. 1958). l
.
I
