Shigeru Yamabe

Comparative activities of the beta-lactamase inhibitors YTR 830, clavulanate, and sulbactam combined with ampicillin and broad-spectrum penicillins against defined beta-lactamase-producing aerobic gram-negative bacilli.

The in vitro synergistic activities of the beta-lactamase inhibitors YTR 830, clavulanate, and sulbactam, combined with ampicillin, ticarcillin, mezlocillin, azlocillin, piperacillin, and apalcillin, were determined against 34 strains of members of the Enterobacteriaceae family, Pseudomonas aeruginosa, Aeromonas hydrophila, and Haemophilus influenzae with characterized plasmid or chromosomal beta-lactamases or both. Strains were tested against fixed concentrations of beta-lactamase inhibitors (8 micrograms/ml) combined with doubling dilutions of beta-lactams. Synergy was defined as a fourfold or greater decrease in the MIC of the beta-lactam. Against Enterobacteriaceae producing Richmond and Sykes class III and V plasmid-mediated beta-lactamases, synergy was obtained against most strains with YTR 830- and clavulanate-beta-lactam combinations, with sulbactam being less effective. Against Enterobacteriaceae producing class I chromosomal beta-lactamases, combinations containing YTR 830 or sulbactam were more synergistic than combinations containing clavulanate. Against strains producing class V PSE enzymes, all three inhibitors were synergistic with piperacillin and apalcillin against strains producing PSE-1, -3, and -4 enzymes, while the PSE-2-producing strain was resistant to all inhibitors. YTR 830-beta-lactam combinations were also synergistic against strains producing the novel beta-lactamases OHIO-1, TLE-1, AER-1, and ROB-1. Overall, YTR 830 with piperacillin or apalcillin was the most effective combination.

Comparative activities of the beta-lactamase inhibitors YTR 830, sodium clavulanate, and sulbactam combined with amoxicillin or ampicillin.

YTR 830, a new beta-lactamase inhibitor, was compared with clavulanic acid and sulbactam against aminopenicillin-resistant clinical isolates. At a concentration of 8 micrograms/ml, YTR 830 was as effective as clavulanate or sulbactam in reducing the aminopenicillin MICs. Combined with amoxicillin, YTR 830 is a potentially useful agent for therapy of many bacterial infections.

Comparative activity of beta-lactamase inhibitors YTR 830, clavulanate, and sulbactam combined with beta-lactams against beta-lactamase-producing anaerobes.

The in vitro activities of the beta-lactamase inhibitors YTR 830, clavulanate, and sulbactam combined with six beta-lactams against 88 beta-lactamase-producing anaerobes were determined. When combined with the beta-lactams, the three beta-lactamase inhibitors showed no synergy against the 10 Bacteroides fragilis homology group II strains. When the beta-lactams were combined with the inhibitors, their geometric mean MICs against the remaining 78 strains were reduced from 4.2 to 150.2 micrograms/ml to 0.2 to 12.9 micrograms/ml. The activity of the beta-lactams combined with the beta-lactamase inhibitors was significantly greater than that of the beta-lactams alone against all groups except B. fragilis homology group II, with 76 to 100% of the strains susceptible to ampicillin plus inhibitor and greater than or equal to 90% susceptible to the other combinations.

A fluorospectrophotometric study on the binding of acridine orange with DNA and its bases

Abstract To confirm the nature of the binding of acridine orange (AO) with DNA, a fluorospectrophotometric study was made on the AO-DNA system as well as on the AOmononucleoside and AO-mononucleotide systems. In the (I) region of higher AO/DNA ratios, the fluorescence intensity of AO decreased markedly, while in the (II) region of lower AO/DNA ratios it increased markedly. Similar changes were observed with mononucleosides and mononucleotides, though their effects on fluorescence intensity were weak. There is a good parallel between these fluorospectral changes and the changes in the absorption spectrum already reported. Therefore, it was confirmed that there are two types of mechanism of AO-DNA binding, namely, self-association, in the (I) region and intercalation in the (II) region. Comparison of the effects of the four nucleosides upon the fluorescence intensity indicated that the contribution of an A:T nucleotide pair to the enhancement of fluorescence due to intercalation is more important than that of a G:C nucleotide pair. The reversing effects of urea, KC1, and ethylene diamine on AO-DNA binding indicate that in the (I) region, ionic bonding between AO and the phosphate groups of DNA is predominant, while in the (II) region, hydrophobic bonding between AO and the bases of DNA is predominant. A comparative study was made on a typical metachromatic system of AO and heparin.

Further fluorospectrophotometric studies on the binding of acridine orange with DNA. Effects of thermal denaturation of DNA and additions of spermine, kanamycin, dihydrostreptomycin, methylene blue and chlorpromazine

Abstract Fluorospectrophotometric studies on the binding of acridine orange (AO) with calf thymus DNA showed that the thermal denaturation of DNA reduced markedly the fluorescence of Complex II and the extent of this decrease depended on the temperature to which the DNA solutions were heated. The denaturation was carried out in the absence and presence of AO (methods A and B, respectively), and then fluorescence measurements of solutions were carried out at 23 °C. The fluorescence intensity-heating temperature curves obtained by methods A and B were similar in shape to the usual melting curves of DNA and AO-DNA solutions, respectively. The higher midpoint value obtained with method B indicates the stabilizing activity of AO against denaturation. These findings support an intercalation model for Complex II and an external self-association binding model for Complex I. A high concentration of ethylene diamine (EDA) restored the fluorescence of denatured Complex II to about 80% of the intensity value of native Complex II. The effects of spermine, kanamycin and dihydrostreptomycin were much stronger than that of EDA. Methylene blue (MB) and chlorpromazine (CP) reduced the fluorescence of native Complex II markedly. Since the analysis of the difference absorption spectra declared that MB and CP were intercalated without release of bound AO, the interacting MB and CP were considered to weaken the interaction between AO and DNA bases, that made AO more fluorescent. Free radical (CP·) of CP was prepared by a new method using H 2 O 2 , peroxidase, and ascorbic acid. Intercalated CP· showed a much stronger quenching effect on Complex II, indicating that unpaired electron spin contained in the costacking unit between CP· and DNA bases might affect the fluorescence of the adjacent AO molecule by paramagnetic perturbation.

Synergistic Effects of Chlorpromazine and Perphenazine on Several Chemotherapeutic Agents

The combination effects of chlorpromazine (CPZ) and periphenazine (PPZ) with beta-lactam antibiotics (ampicillin, carbenicillin, cefazolin) and nalidixic acid group compounds (nalidixic acid, piromidic acid and pipemidic acid) have been estimated to be synergistic by the filter paper strip-agar diffusion method (Dyes method) with Escherichia coli and Pseudomonas aeruginosa as test organisms. The observed synergism might be associated with their inhibition of various enzymes including ATPase and DNAase as well as with their specific binding to DNA. Similar synergistic effects of CPZ and PPZ have been shown by the broth dilution method. Based on these findings, it seems to be a fascinating project to devise a new phenothiazine drug without influence in mental disease that will have a greater measure of synergistic effect when combined with the above-studied antibacterial agents.

In Vitro Activity of YTR 830

YTR 830, now known as tazobactam, is a new penicillanic acid sulfone beta-lactamase inhibitor. The in vitro activity of YTR 830 combined with various penicillins was determined and compared to that of clavulanate and sulbactam combined with the same agents. Combined with ampicillin or amoxicillin, all three inhibitors were active against beta-lactamase producing strains of Staphylococcus aureus, Haemophilus influenzae, Klebsiella, Citrobacter diversus, and all anaerobes except for Bacteroides fragilis homology group II. YTR 830 was also effective against Escherichia coli and indole-positive Proteus. The inhibitors had no effect against Enterobacter or Serratia. Overall, the activity of YTR 830 was comparable to that of clavulanate, and superior to that of sulbactam.

Inhibitory Effects of Aminoglycoside Antibiotics on Reduction of Cytochrome c from Candida krusei

Five aminoglycoside antibiotics (AGAs)--kanamycin (KM), bekamycin (AKM), dibekacin (DKB), ribostamycin (RSM) and paromomycin (PRM)--were studied for their effects on the nonenzymic reduction of cytochrome c by FeSO4 (Yamabes system). Their inhibitory activity was in the order: DKB greater than AKM greater than KM greater than RSM greater than PRM. As this order correlated closely with that of the antibacterial activity of AGAs, Yamabes system has proved useful in predicting the latter activity. Divalent metal ions other than Fe2+ enhanced the AGA-dependent inhibition of Yamabes system in the order: Cu2+ greater than Mn2+ greater than Zn2+ greater than Co2+ greater than Ni2+ greater than. This order was similar to that of stability constants with general chelators except for the low positions of Ni2+ and Co2+. These findings suggested a metal chelation with free or bound Fe2+ for the action mechanism of AGAs on Yamabes system and the bacterial growing system. The antagonistic effects of exogenous Fe2+ on the antibacterial activity against Staphylococcus aureus and Escherichia coli as measured by the agar dilution method supported this concept. A dual relationship of molecular structure with chelating and antibacterial activities demonstrated the importance of high molecular basicity in a potent AGA. However, the combination effect of pipemidic acid (stimulator on Yamabes system) with KM was different from that with 1,10-phenanthroline (inhibitor on Yamabes system) as measured by Dyes method using Pseudomonas aeruginosa.

Effects of pyrrol-carboxylic acid derivatives on the growth of coliphages.

Effects of 14 pyrrol-carboxylic acid derivatives and analogues (PY-compounds) on the growth of coliphage MS2 using E. coli E102 (Hfr) as the host were measured by the agar double-layer method. Enlargements of plaque size were observed with 7 PY-compounds but increase in plaque numbers was not induced. These enlargements of plaque size were specific to RNA coliphages MS2, GA and qbeta and not found with DNA coliphages delta AC and T4. Furthermore, the interaction between PY-compound PY-10 and the coliphage MS2 was dependent on the host bacterium (indicator strain). When E102 (Hfr) was used, the enlargement was marked, in the case of substrain W1895 (Hfr) it was less, while in the case of substrain W6 (F+) it was undetectable. The one-step growth of the phage MS2 and the production of intracellular phage MS2 were little affected by the PY-compound PY-10. However, the rate of one-step growth was increased in the early stage after infection. Accordingly, the enlargements of plaque size by the PY-compounds might be correlated with an increase in rate of release of phage particles.