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Lederle Synthesizes Tetracycline Proof of structure confirmed by researchers as threey e a r p r o g r a m duplicates derivative of natural antibiotic J\.
TETRACYCLINE antibiotic has
now
been synthesized by American Cyânamid a t its Lederle Laboratories» in Pearl River, N. Y., w h e r e chlorotetraeycline was first produced commercially a b o u t 10 years ago. Total synthesis consists of 24 discrete chemical transformations, begins with a common industrial cresol, and yields a tetracycline derivative identical with one produced by partially degrading material made by fermentation (/ACS, F e b . 2 0 ) . Degradation methods at Lederle and elsewhere had determined structures of oxytetracycline and chlorotetracycline. Since then, Cyanamid's researchers h a v e pursued fermentation studies to seek other useful tetracycline derivatives. M u c h of this work involves us-
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ing various strains of the microorganisms Streptomyces (uireofaciens. Mutants of iiies»tr iniu Outrs», feeding Oil corn-steep liquor as nutrient, can produce compounds having different structures, Lederle explains. W h i l e continuing its fermentation studies, Lederle decided three years ago to try synthesizing a tetracycline. The synthesis project, which has heen carried on uninterruptedly since 1955, has n o w produced its first tetracycline: biologically active riLceinic dedimethylamino - 12a - deoxy-6-dernethylaiihydrochlorotetracycline. By analyzing infra-red and ultraviolet absoiption spectra, l>ioassays, and chromatographic behavior, Lederle chemists have shown it t o be the same as a
sample derived from the natural antibiotic, 6-demethylchlorotetracycline ^L/AlV^l
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and acetic acid, followed by dehydration with mineral acid. T h e only perceptihle difference between the natural and synthetic materials, according to group leader James H . Boothe, is that the man-made compound is racemic; the fermentation product is dextrorotatory. • Start w i t h Cresol. Boothe, with coworkers Andrew S. Kende, Thomas L. Fields, a n d Raymond G. Wilkinson, begin their synthesis with commercially available p-chloro-m-cresol. They convert this first to the anisole then to 2-chloro-5-metlioxybeiizyl bromide, which they use to prepare the benzyl-
RESEARCH malonic ester and then the homologous dinitriie. Alkaline hydrolysis to a substituted benzylglutaric acid is followed by eîosuire of the second ring upon treatment with polyphosphoric acid. Key t o making the third ring turns out to b e preparation of an aldehyde by red ι irin g an intermediate acid chloride. This opened the door after a great deal of fruitless knocking, says Samuel Kushner, department head who has been associated with Lederle's tetracycline research since it started. Condensing the aldehyde with excess cyanoacetamide, hydrolyzing to a diacid, a n d then esterifying, gives a diester which is conventionally cyclized by sodium hydride. The resulting tricyclic ester is par tially aromatized, converted to the carboxylic acid and then to the acylmalonate which gives a fourth ring, closed again b y reaction with sodium hydride. The ethyl ester is transformed smoothly into the desired amide by fusion with ammonium formate. Direct dealkylation by hydrochloric acid in acetic acid removes stabilizing groups to form the end product. • Sequential Çyclization. The concept of sequential ring closure was the cornerstone of this successful synthesis, says J. J . Denton, director of Lederle's organic chemicals research section. Major h u r d l e was finding how to leave the proper substituents in position ready for subsequent ring closures. T h o u g h this program cost "hundreds of thousands of dollars" in direct expense, Denton foresees no immediate return from the investment. But the synthesis does confirm the accepted framework of tetracycline antibiotics. F r o m here, Lederle may: • Prepare naturally occurring (fermentation^ tetracyclines, such as DMCT itself. • Make new tetracyclines not produced oy fermentation. • Determine how tetracycline acts as an antibacterial in animals. « Synthesize smaller antibiotic molecules containing elements of the tetracycline structure. Work i n various stages of progress is said t o be under way at university laboratories in Germany, England. U.S.S.R., and the U. S. But Lederle is the first to publish the total synthesis of tetracycline, it says.
COLOR COMPUTATIONS CUT· These NBS color specialists and mathematicians, (left to right) J. C. Sehleier, W . C. Rheinboldt, H . J. Keegan, and J. F. M e n a r d , use an electronic digital computer to convert spectral reflectance data into colorimetric terms. Color tree (center) is a three-dimensional array of the spectra
Computer Defines Color The couturiers of Paris, Rome, and New York decree what shade the ladies' hose shall be this spring. But fashion would not be so splendid if laboratories such as the National Bureau of Standards—which lately has brought a computer into the act—did not define, refine, and catalogue all the hues with which man arrays himself and his environment. But the job of defining color has involved tedious computations requiring hours to pinpoint a single color and determine its spectral data. T o lessen the color specialist's burden, NBS started a program last year to reduce this time to fractions of a second by using an electronic digital computer. In 1931, the International Commission on Illumination resolved that the color of any light is defined by specifying the amounts of primary red, green, and blue light which, when added together, produce a particular color. The fundamental measurement of a color standard is the spectrophotometric determination of the fraction of incident light transmitted or reflected by the standard in successive parts of the visible spectrum. This procedure means that the color
specialist must first compute the amount of light transmitted or reflected by the standard for each wavelength of the visible spectrum. This relative intensity is then multiplied by factors for the red, green, and blue primaries of the average human eye. These products when added together, give a specification of a color. This must b e done for each wavelength, and the work involves using desk computers, slide rules, planimeters, and nomographs. The final data that t h e spectroscopist comes u p with are points on a spectral curve which exactly pinpoint a color. Now, with the digital computer, NBS can do th° "*oh » aster and with few*3!* errors. Also programmed for the bureau's computer is the conversion of colorimetric data to Munsell terms—the scales of hue, value, and chroma used by industry and science to specify color and color tolerance. The Munsell scales correspond closely to the hue, lightness, and saturation of a color seen under daylight viewing conditions. To get the Munsell notation of a color requires a tridimensional interpolation among 5000 points which define the Munsell scales; this takes about 20 minutes. The computer does the job in 20 seconds. MARCH
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