Kanamycins A and

Among the older aminoglycoside derivatives, kanamycin A and sisomicin were, at one time, a significant part of medical practice, but have now been largely replaced by the compounds Hsted in Table 1. Streptomycin is stiH used in a few restricted situations. 

Dideoxy-2 -fluorokanamycin A (Ref. 170), a kanamycin derivative in which the OH-2 or NH2-2 of kanamycin A and B, respectively, is replaced by a fluorine atom, was prepared by condensation of 6-azido-4-0-benzoyl-2,3,6-trideoxy-2-fluoro-a-D-n77ohexopyranosyl bromide, derived from 179 (see Section 11,2), with the pseudodisaccharide component of kanamycin. The compound showed mediocre antibacterial activity however, on attachment of an (5)-4-amino-2-hydroxybutanoyl residue, as in amikacin, to the NHj-l group, the derivative showed considerably enhanced activity. ... 

Unit, [tg/mL Kan, kanamycin S. aureus, Staphylococcus aureus P. aeruginosa, Pseudomonas aeruginosa AAC, aminoglycoside acetyltransferase ANT, aminoglycoside nucleotidyltransferase Perm, mutant, mutant strain impermeable to kanamycin A and B. 

Cowan and co-workers ° have extensively characterized the Cu " complexes of two different classes of aminoglycosides—namely, kanamycin A (4) and neomycin B (2)—in addition to the neamine (1) class of aminoglycosides. Their findings, based upon the interaction of CUSO4 with kanamycin A and neomycin B, identified a blue monomeric 1 1 complex of Cu -kanamycin A as well as a dark green 1 2 complex Cu +-(kanamycin A>2 and a 1 1 Cu +-neomycin B complex, although the latter two were found to be less stable in aqueous solution with observable precipitation within 4 h. The Cu +-kanamycin A complex was found to be most stable in aqueous solutions over a period of one day. Based on C-13 and H-1 NMR relaxation experiments, the Cu -kanamycin A solution structure at pH = 7.9 (12) is described as Cu + coordinated to the... 

The MASS assay was used to determine the Kd values for paromomycin, kanamycin A, and neamine. While the aminoglycoside anhbiotics were screened at 250 nM, 750 nM. 2.5 /trM, and 7.5 /rM, 16S was held at 100 nM. Figure 3.2 shows a plot of the frachon of 16S bound by the hgand (fraction bound) versus the total hgand concentrahon for the three aminogylcosides. The Kd values determined for paromomycin (83.9 nM), kanamycin A (7.8 I M), and neamine (13.1 /zM) are consistent with soluhon phase determina-hons [26,30] and our previous ESI-MS measurements [18]. Each Kd determinahon took 380 seconds of data acquisihon. Thus, the MASS assay provides a robust way to determine Kd values for 100 compounds a day. 

FIGURE 3.2 Kd determination for paromomycin, kanamycin A, and neamine. 16S was held at 100 nM concentration, while the aminoglycosides were screened at 250 nM, 750 nM, 2.5 gM, and 7.5 gM. The fraction hound is plotted versus the initial hgand concentration. Nonlinear regression was used to calculate the Kd and the curves were fit with > 0.99. 

During tliis synthetic work, Umezawa developed several tecluiiques for the selective protection of hydroxy 1 and amino groups in aminoglycosides. These include the carbamate - and zinc-chelate protection of kanamycins, developed in collaboration with Tsucliiya and Takagi. The latter procedure became crucial for the industrial production of amikacin from kanamycin A and arbekacin (habekacin) from dibekacin. 

An attempt to make 3, 4 -dideoxy analogs of kanamycin A and B by mutational biosynthesis in the presence of gentamicin-producing Micromonospora organisms... 

The same group also has described the syntheses of 6"-deoxy-6"-fluorokana-mycin A, 6"-deoxy-6"-fluoroamikacin, and l-A-[(/ )- and (/ 5 )-3-amino-2-fluoropro-panoyl] kanamycin A, using the DAST reagent for fluorination of the corresponding hydroxyl groups of kanamycin A and amikacin derivatives [112],... 

A-Benzamido derivatives of kanamycin A and tobramycin having a chromium complex in the benzene ring were prepared for use as metallotracers in im-mimoassay studies. To make these compoimds, A-hydroxysuccinimide-activated esters of these organochromiums were treated with kanamycin A or tobramycin to selectively produce the 6 -N-acylated derivative [138]. 

Different amino acid and peptide derivaties were used to synthesize a series of aminoacyl and dipeptidyl derivatives of kanamycin A and netilmicin modified at the 1-, and 6 -amino groups by selective acylation of the copper complexes of these antibiotics. However, none of these modified compounds were active against either gram-positive or gram-negative bacteria [139]. Recently, in a report from Abbott... 

Kanamycin. Antibiotic complex produced by Streptomyces kanamyceticus Okami A Umezawa from Japanese soil Umezawa el ai. J. Amibiot. 10A, 181 (1957) U.S. pat. 2,931,798 (1960). Comprised of three components, kanamycin A, the major component (usually designated as kanamycin) and kanamycins B and C, two minor congeners, lsoln and purification of kanamycins A and B and their salts Johnson er at., and Johnson, Hardcastle. U.S. pats. 

The reaction of204 with kanamycin A and subsequent deprotection affords 6 -N-L-(-)-HABA-kanamycin A (BB K6) 182. If the 6 - and 1-positions are previously protected the 3-N-L-(-)-HABA-kanamycin A (BB-K29) is isolated. Following the additional protection of the 3-position, the reaction with the active ester leads to 3"-N-L-(-)-HABA-kanamycin A (BB-K 11) These isomers are thirty to a hundred times less active than BB-K8 (Amikacin). 

Amikacin is more potent than kanamycin A and superior to BB-K19 (D,L-HABA) and BB-K31 (D(+)-HABA). Amikacin is the only important semisynthetic aminoglycoside on the market. In this connection two possibilities to synthesise the HABA moiety should be described. One of these syntheses starts with butyrolactone and uses the steps shown in Fig. 46 a. Another method utilizes the 1,3-dipolar addition of the nitrone ester 205 to ethyl acrylate (Fig. 46b). 

It has been known for a long time that the removal of both the amidino groups in streptomycin and the acylation of the amino groups destroys the antibiotic activity, whereas the reductive amination, the reduction to dihydrostreptomycin, the monosubstitution of an amidino function by a carbonyl group as in bluensomycin and the demethylation of the N-methylamino group do not affect the effectiveness of streptomycin. The differences in the activity of kanamycin A and B and the neomycins have shown that the number and the position of the amino groups are of importance for their efficacy. 

N-Methanesulfonate derivatives of kanamycin A and neomycin have been prepared, - and studied biologically in detail. - ... 

At first, as just described, the structures of the inactivated kanamycin A and paromamine were elucidated chemically, but they were also... 

Chemical Shifts and Coupling Constants for the 6-Amino-6 eoxy D>gluco e Moiety in Kanamycin A and Kanamycin A 3 -Fho phate... 

The lividomycin A-Sepharose 4B was packed in a column (1.0 X 11.7 cm 9 ml) and washed with 20 mM Tris hydrochloride buffer containing 10 mM magnesium acetate, 60 mM potassium chloride, and 10 mM 1,4-dithiothreitol, and 5 ml of the supernatant liquor from the disrupted cells was passed through the column at the rate of 40 ml/hour. The enzyme adsorbed was successively eluted with a gradient of sodium chloride (0 to 0.8 M) in 20 mM Tris-hydrochloric add buffer, pH 7.2. The enzyme that phosphorylates and inactivates both lividomycin A and kanamycin appeared in the eluate made with 0.4 to 0.6 M sodium chloride. By this procedure, the enzyme was purified 25- to 40-fold in phosphorylating activity for both kanamycin A and lividomycin A. The purified fraction obtained showed two bands in disc-gel eledrophoresis, and one of the bands had the activity as regards phosphorylating kanamycin A and lividomycin A. 

Refluxing of the inactivated kanamycin A in 2 M sodium hydroxide for 9 hours gave kanamycin A, and under these conditions, no decomposition of kanamycin A occurred therefore, this method is useful in determination of the inactivated kanamycin. The inactivated kanamycin melted at 176-180° (bubbling). The formula of monoacetylkanamycin A was obtained by elementary analysis. The amide bond was shown by i.r. spectroscopy. Mono-N-acetylkanamydn was shown by the presence of three nitrogen atoms (Van Slyke) and by the behavior in high-voltage electrophoresis, in comparison with mono-, di-, tri-, and tetra-V-acetyl-kanamycin. The amount of periodate consumed per mole at pH 4.3 in 1 M sodium acetate buffer at 24° was the same as that per mole of kanamycin A. The inactivated kanamycin A was deaminated by the Van Slyke... 

Escherichia coli K12 R5 (which produced this enzyme) was weakly resistant to kanamycins A and B, but sensitive to kanamycin C, which has no 6 -amino group to be acetylated. The involvement of this enzyme in the mechanism of resistance was shown by the resistance of this organism to kanamycin A and 3, 4 -dideoxykanamycin B, and by its sensitive behavior to the 6 -iV-methyl derivatives of these antibiotics. 

The involvement of this enzyme in the mechanism of resistance of E. coli carrying R factor is certain, because the Escherichia are inhibited by kanamycins A and B, and 3, 4 -dideoxykanamydn B. 

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