Proteus sp.y Klebsiella -pneumoniae, Aerobacter aerogenes, and Pseudomonas aeruginosa; and cases of meningitis due to susceptible strains of Salmonella. T h e agent h a s been found helpful in t h e t r e a t m e n t of t h e following diseases, b u t its clinical position h a s n o t been definitely defined: peritonitis, liver a b scesses, bile d u c t infections, wound infections, and empyema due to susceptible g r a m - n e g a t i v e organisms; tuberculosis; chronic lung infections d u e to mixed gramnegative flora; inflammation of t h e heart valves caused b y penicillin-resistant b u t streptomycin-sensitive pathogens; and pneumonia due to Klebsciclla pneumoniae. S t r e p t o m y c i n is of questionable value in t h e t r e a t m e n t of typhoid fever, brucellosis, and salmonella infections. In preliminary trkils, it h a s been found ineffective in t r e a t i n g Clostridia infections, rickettsial infections, virus malaria, diseases, a n d infections due to molds and fungi.
Production of Streptomycin H E R B E R T SILCOX,
Merck & Company, Inc., Rahway, N. J.
Chemical Tfie new finishing Here streptomycin
plant of Merck Sz Co., Inc.* at Rahway, is dried, milled to powder, weighed, and
Ar. J. packed
S t r e p t o m y c i n , a ne^v a n t i b i o t i c e f f e c t i v e a g a i n s t c e r t a i n d i s e a s e s n o t controlled b y penicillin, is n o w b e i n g produced o n a large s c a l e . D i s c o v e r e d i n 1944, s t r e p t o m y c i n h a s b e c o m e a c o m m e r c i a l reality7 a s t h e r e s u l t of r a p i d a n d e x t e n s i v e p r o c e s s d e v e l o p m e n t a c c e l e r a t e d b y repeated confirmation of m e d i c a l efficiency S T R E P T O M Y C I N is an a n t i b i o t i c agent produced b y certain strains of Streptomyces griseus. I t was found as the result of a search for an agent t h a t would b e active against gram-negative bacteria y e t "nontoxic to 'the body a n d would therefore offer possibilities as a chemotherapeutic agent. I t was first obtained a s a crude concentrate in 1944 b y SeLman A. W a k s m a n and Albert Schatz at t h e New Jersey Agricultural E x p e r i m e n t Station, R u t g e r s University (5). T h e concentrate isolated by Dr. W a k s m a n showed sufficient therapeutic value to stimulate considerable effort by Merck & Co., Inc., t o w a r d t h e establishment of its chemical composition, the development of m e t h o d s for its production, and t h e s t u d y of its medical performance. Encouraged b y t h e success of these combined evaluations, Merck s t a r t e d plans for large scale production in July 1945, and, in August of the same year, t h e construction began on a $3,500,000 p l a n t in E l k t o n , Va., and R a h w a y , N . J. Although the installation h a d n o t a s yet been completed, the first production of streptomycin c a m e out of t h i s plant during M a y 1946. Application
of
Streptomycin
Although streptomycin, like penicillin, is produced from organisms commonly found in t h e soil, streptomycin is n o t a s u b s t i t u t e for penicillin. W h e r e a s p e n i cillin is primarily effective a g a i n s t g r a m positive bacteria, streptomycin is useful 2762
Structure
of
Streptomycin
Considerable progress h a s been m a d e in t h e determination of the chemical s t r u c t u r e of streptomycin. The formula of t h e antibiotic has been established as C 21 H 39 N 7 Oi 2 (£). It consists of a h y droxylated base, streptidine (4), linked glycosidically to a disaccharidelike molecule, streptobiosamine ( i ) . Aqueous acid hydrolysis of streptom y c i n yielded streptidine, which h a s been s h o w n to b e a l,3-diguanidino-2,4,5,6tetrahydroxycyclohexane. Degradation of s t r e p t o m y c i n with methanol and hydrogen chloride (1) h a s given streptidine and a derivative of streptobiosamine. F u r t h e r h y d r o l y tic t r e a t m e n t of a streptobiosamine d e r i v a t i v e led to t h e isolation of iV-methyl/-glucosamine (2, 3). T h e degradation of streptomycin t o s t r e p t i d i n e a n d streptobiosamine and t h e further hydrolysis of t h e disaccharide t o yield A r -methyl-Z-glucosamine m a y b e represented b y t h e following e q u a t i o n s :
mainly in t h e control of infections d u e t o susceptible strains of gram-negative b a c teria. I n t h i s respect, it m a y be said t h a t streptomycin is s u p p l e m e n t a r y to p e n i cillin. Streptomycin has found established use in the t r e a t m e n t of t u l a r e m i a ; all t y p e s of infection caused by Hemophilus influenzae (meningitis, endocarditis, laryngot r a c h e a l , urinary t r a c t and p u l m o n a r y infections); cases of meningitis, b a c t e remia, and urinary t r a c t infections due to susceptible s t r a i n s of Escherichia coli,
NH
NHC-
-NH2
AH C 21 H 39 N 7 0 12 -f- H 2 0
CHOH/\CHOH I
I
+
N H
CsHaaNOa
C H O H V C H N H C - NH2 Streptomycin
CHOH Streptidine
+
Streptobiosamine
—CHOH I CH.NHCI-I C13H23TsT09 -f- H2O •
CeHioOs
Streptobiosamine
+
+ C H E M I C A L
O
IICOH I HOCII I CH
CHoOH iV'-methyl-Z-glucosamine
A N D
ENGINEERING
NEWS
Complete knowledge of t h e chemical s t r u c t u r e of streptomycin awaits further information p e r t a i n i n g to the structure of t h e product CsH^Os and the positions of linkages between t h e three fundamental p a r t s of the molecule. S t r e p t o m y c i n forms a complex salt with calcium chloride a n d a photomicrograph of crystals of this double salt is shown. Production of Streptomycin All t h e streptomycin produced to d a t e by Merck h a s been m a d e by aerobic ferm e n t a t i o n followed by recovery from t h e fermentation liquors and purification. As currently conceived, the production process engenders nearly all of the classical chemical engineering operations in t h e large n u m b e r of steps necessary. I n certain instances, during the development of the process, it was found necessary t o extend t h e engineering research fundamentals bej'ond those currently available and, in one i n s t a n c e , a new t y p e of chemical engineering process was developed for production. Of the large number of raw materials required for t h e process, m a n y are unique a n d are being used in a production process for the first t i m e . T h e work o n streptomycin, as in t h e development of t h e penicillin recovery processes, was hampered by limitations such as t h e e v a l u a t i o n of experiments b y biological m e t h o d s and t h e fact t h a t streptomycin, as now produced, is n o t a p u r e chemical c o m p o u n d t h a t can be controlled b y a few rigid specifications. A n y contemplated variation in process, r e gardless of how insignificant it m a y seem, m u s t be evaluated a s to its effect not only on t h e remaining s t e p s of the process b u t also on t h e clinical behavior of the final product, a s t h e clinical product m u s t be carefully guarded against the development or introduction of any impurities t h a t produce toxic effects on administration. Evaporators streptomycin
V O L U M E
for
2 4,
concentrating solutions
NO.
20
crude
The Merck plant produces s t r e p t o mycin in 15,000-gallon fermenters. T h e recovery of streptomycin is .then a c c o m plished by a continuous process w h i c h results in a crude, d r y s t r e p t o m y c i n hydrochloride. T h e isolation of this c r u d e material is accomplished by m e t h o d s similar to those published by \ V a k s m a n (6). T h i s crude material is then s u b jected to extensive purification and finishing operations. Streptomycin Fermentation Although strcptonr _in is produced b y aerobic fermentation, it is possible, a s with penicillin, to carry out the f e r m e n t a tion in t a n k s equipped with agitation d e signed for optimum gas-liquid c o n t a c t i n g . In order to obtain a maximum p r o d u c t i o n of streptomycin per gallon of f e r m e n t a t i o n capacity, a system based on mass i n o c u l a tion is used. T h i s system involves six steps from the soil tubes to t h e 15,000gallon fermenters, all operating u n d e r aseptic conditions. T h e composition of the f e r m e n t a t i o n medium has a marked effect upon the p r o duction of streptomycin. T h e m e d i u m , a s proposed by W a k s m a n (tf), consists of: Glucose Peptone M e a t extract Sodium chloride T a p water
10 5 5 5 1,000
grams grams grams grams ml.
I t h a s been shown possible to u s e various substitutes for glucose, p e p t o n e , a n d m e a t extract. These s u b s t i t u t e s , however, have varying effects upon t h e recovery and purification process. Frequently, considerable foaming i s experienced during the fermentation. T h i s situation can be alleviated b y t h e use of antifoam agents which are not t o x i c to the fermenting organisms and do n o t interfere with t h e recovery and purification processes.
Crystalline streptomycin chloride—calcium chloride Filtration
of Fermented
trihydrodouble salt Broth
T h e insoluble solid in the fermented broth is composed primarily of mycelium which is extremely fine and gelatinous in character. I t is difficult to separate this solid b y centrifugal force and i t also blinds any filtration medium very rapidly. Filter aids m a y be used to improve the filtration characteristics; however, their choice m u s t be carefully made, in t h a t certain of these filter aids tend to adsorb streptomycin. The method chosen for handling b r o t h in large volumes involves the filtration of admixed b r o t h on pressure r o t a r y precoat filters with advancing knives and water washing. I n spite of great variations in t h e filterability of individual fermentation batches, i t is possible to operate this type of u n i t in t h e continuous process b y making compensating variations i n the operating conditions. T h e organic material in the filter cake is disposed of b y means of incineration. Adsorption
of
Streptomycin
Following broth filtration, the strepto-
A section of the Elkton plant. Here streptomycin is manufactured in the first mass-production unit in the world. Starting from tiny test tubes, the microorganisms multiply so rapidly that in a few days they fill 15,000-gallon tanks
» > OCTOBER
2 5,
1946
2763
mycLn-rieh lilt rate i> admixed with art 1vated carbon on a continuous basis. T h e carbon is introduced by a measuring mecha,sm which integrates the amount of carbon added with the flow of broth fill r a t e and the streptomycin concentration in t h e filtrate. The introduction of carbon must be very closely controlled, as insufficient carbon results in incomplete adsorption and excess carbon reduces the elation yields. The strcptoinycin-rich carbon is separated from the spent broth by filtration. T h e spent broth has a very high biological oxygen demand, and it is d i s posed of by quadruple effect evaporation followed by inciner.-it ion of the resulting concent rate. Following separation from the spent broth, the rich charcoal adsorbate is washed first with alcohol and then eluted with acid-alcohol so hit ion. T h e acid eluate is neutralized and the streptomycin concentration in soluti'-n has now been increased to 0. lO to O.LV, and con. iderable preliminary purification has taken place. Evapora tion of Neutral El note The neutral eluate is now concentrated to a solution from which the streptomycin
Scaborg Announces Fissionable Neptunium Tla v discovery of a fissionable isotope of neptunium was announced by Glenn T. Seaborg, professor of chemistry at the University of California, at a meeting of the California Section of the AMKRICAX CMJVMICAL SO0-ineh cyclotron at Berkeley, but its fissionable tendency and low fission cross section (2 X 10 2G sq. cm.) were discovered by A. (Jhiorso, D. "W. Osborne, and L. H. Magnusson, work-
can be precipitated by solvents as the crude hvdrochloride and be recovered
ing; with Dr. Seaborg at the iikotallurgical laboratory, University olChicngo. \"p-' 17 is among t h e by-products of p l u 'oniuni production a< the Clinton a n d ITanford plants. Dr. Senborg also reported the d i s covery of a new isotope of plutoniuni-, Pu 241 , and announced the mass n u m bers of the elements, ainericium a n d curium. Pu-' 1 i- produced by a l p h a particle bombardment ofl~- : i \ It h a s a relatively long half-life and
