R. Michael E. Richards

Vancomycin resistance reversal in enterococci by flavonoids

The development of clinical vancomycin‐resistant strains of enterococci (VRE) is a major cause for concern. Here we show that a combination of galangin or 3,7‐dihydroxyflavone with vancomycin may be used to sensitize resistant strains of Enterococcus faecalis and Enterococcus faecium to the level of vancomycin‐sensitive strains.

A triterpene from Rubus pinfaensis.

A new pentacyclic triterpene, pinfaensin, was isolated from the roots of Rubus pinfaensis and the structure elucidated by 13C NMR methods. This triterpene, a glycoside of pinfaensic acid (glucosyl 23-formyl-2 alpha,3 beta,19 alpha-trihydroxyurs-12-en-28-oate), in order to achieve purification and characterization, was hydrolysed to previously unreported pinfaensic acid which on acetylation and methylation gave methyl diacetoxypinfaensate (methyl 2 alpha,3 beta-diacetoxy-23-formyl-19 alpha-hydroxyurs-12-en-28-oate).

Investigation of Synergism between Combinations of Ciprofloxacin, Polymyxin, Sulphadiazine and p-Aminobenzoic acid

Abstract— Subinhibitory concentrations of combinations of any two of ciprofloxacin, colistin (or polymyxin B), sodium sulphadiazine and p‐aminobenzoic acid were shown by checkerboard minimum inhibitory concentration determinations to have synergistic inhibitory activity against Pseudomonas aeruginosa and to have either synergistic or additive activity against Staphylococcus aureus. In addition, sulphadiazine plus either ciprofloxacin or polymyxin showed markedly enhanced killing activity against both P. aeruginosa and S. aureus. p‐Aminobenzoic acid plus either ciprofloxacin or polymyxin also demonstrated enhanced killing activity against P. aeruginosa but these combinations were less effective in enhancing activity against S. aureus. Ciprofloxacin in combination with polymyxin had a marked synergistic effect against P. aeruginosa but only a slight synergistic effect against S. aureus. These findings indicate a potential usefulness for the synergistic combinations against P. aeruginosa and S. aureus in the clinical situation; that is, they indicate an extended role for sulphonamides and support a potential role for p‐aminobenzoic acid as enhancers of the activity of primary antibacterial agents such as ciprofloxacin and polymyxin. We suggest that for a second antibacterial to enhance the activity of ciprofloxacin, it may be necessary for the second antibacterial to increase cell permeability so increasing bacterial uptake of ciprofloxacin.

Unsaturated E-ring triterpenes from rubus pinfaensis

Abstract Two new isomeric unsaturated E-ring pentacyclic triterpenoids have been isolated from the roots of Rubus pinfaensis . These triterpenes, glucosides of 2α,3β,23-trihydroxyurs- 12-en-28-oic acids with additional Δ 18 or Δ 19 unsaturation, were hydrolysed to the previously unreported pinfaenoic and isopinfaenoic acids, which, on acetylation and methylation, were converted into methyl triacetoxy-esters (methyl 2α,3β,23-triacetoxyursa- 12, 18-or -12,19-dien-28-oates), allowing purification and characterization by 13 C and 1 H NMR techniques. The possibility of these triterpenoids arising as artefacts of the isolation process has been addressed.

Enhancement of antibacterial activity by p-aminobenzoic acid and sulphadiazine

Abstract Subinhibitory concentrations of combinations of the sulphonamide antagonist p -aminobenzoic acid with either carbenicillin, dibromopropamidine isethionate, or polymyxin B were shown to have synergistic antibacterial activity against Pseudomonas aeruginosa, Enterobacter cloacae, Proteus mirabilis and Staphylococcus aureus . Sulphadiazine was compared with p -aminobenzoic acid for its ability to enhance the killing times of dibromopropamidine isethionate, polymyxin B and carbenicillin. p -Aminobenzoic acid enhanced the activity of all the antibacterials and sulphadiazine enhanced the activity of dibromopropamidine isethionate and polymyxin B against all the bacteria. These findings indicate a potential usefulness for both p -aminobenzoic acid and sulphadiazine in enhancing the antibacterial activity of antibiotics and primary antibacterial agents in both topical and systemic use. The findings also indicate that p -aminobenzoic acid and sulphadiazine have distinctly different mechanisms of action. The antibacterial enhancing activity of sulphadiazine is not unique to its use in combination with trimethoprim.

Investigation of the antibacterial activity of p-aminobenzoic acid against P. aeruginosa and E. cloacae

Abstract p -Aminobenzoic acid was found to have a bactericidal action against Pseudomonas aeruginosa and Enterobacter cloacae . This action was not the result of a reduction in pH. An open porin strain of P. aeruginosa was more sensitive to p -aminobenzoic acid than the mother strain, indicating uptake via the outer membrane porin protein. Subinhibitory concentrations of a combination of p -aminobenzoic acid with dibromopropamidine isethionate were shown to have synergistic antibacterial activity against P. aeruginosa . Bacterial uptakes of the antibacterials determined by an HPLC assay method combined with dry cell weight determinations indicated that enhancement of activity of the combination was related to the fact that both compounds increase the bacterial uptake of the other. P. aeruginosa cells grown in the presence of subinhibitory concentrations of p -aminoenzoic acid sustained damage which made the cells more sensitive to the action of both benzalkonium chloride and EDTA and to lysis by lysozyme plus EDTA. These cells also exhibited lysis when treated with lysozyme alone indicating the p -aminobenzoic acid causes damage to the outer membrane. Folinic acid was found to block the antibacterial activity of p -aminobenzoic acid and to increase bacterial resistance to benzalkonium chloride and EDTA.

The effect of formulation on the antimicrobial activity of cetylpyridinium chloride in candy based lozenges

AbstractPurpose. The purpose of this investigation was to determine the influence on the antimicrobial activity of cetylpyridinium chloride of the various components of the formulation of each of six candy based lozenges. Methods. In vivo activity was investigated using six volunteers by determining the reduction in colony forming units recoverable from the oropharynx after sucking each lozenge separately on different days. In vitro determinations investigated the relative activity of aqueous solutions of the lozenges, the effect on activity of additional active ingredients, pH and lozenge base ingredients against separate inocula of each of the test organisms Staphylococcus aureus, Streptococcus pyogenes and Candida albicans. Results. Both in vivo and in vitro results showed that the pH of the dissolved lozenge solution was the single most influential readily adjustable formulation parameter which significantly influenced the activity of cetylpyridinium chloride activity in candy based lozenges. Conclusions. Lozenges containing cetylpyridinium chloride as the active ingredient should be formulated at a pH greater than 5.5.

Excipient interaction with cetylpyridinium chloride activity in tablet based lozenges.

AbstractPurpose. The purpose of the investigation was to determine the effect of tablet excipients on the activity of cetylpyridinium chloride (CPC) and the relative interaction between excipients and CPC. Methods. An analytical assay was developed to evaluate the interaction between CPC and the excipients. In vivo activity was investigated using six volunteers by determining the reduction in colony forming units recoverable from the oropharynx after sucking each proprietary lozenge separately on different days. In vitro determinations investigated the relative antimicrobial activity of aqueous solutions of the lozenges and, the effect of pH and tablet base excipients on that activity against Staphylococcus aureus, Streptococcus pyogenes and Candida albicans. Results. Both in vivo and in vitro results showed that the tablet based lozenges had markedly reduced antimicrobial activities compared with previous results with a candy based lozenge (invivo and in vitro) or the same concentration of aqueous CPC (in vitro}. Magnesium stearate suspensions in CPC 250 µg/ml indicated that magnesium stearate adsorbed CPC and at 0.4% lozenge weight and above significantly reduced the antimicrobial activity of CPC 250 µg/ml. Conclusions. The reduced activity of CPC in tablet based lozenges resulted from a decreased availability of CPC in solution due to an adsorption of CPC on magnesium stearate. To avoid this reduction in activity tablet based lozenges containing CPC 250 µg/ml, or similar concentrations, plus magnesium stearate should contain not more than 0.3% w/w lozenge weight of the lubricant.

Simultaneous determination of some antibacterial drugs in Isosensitest broth using high-performance liquid chromatography with solid-phase extraction

A method is described for the simultaneous determination of combinations of some antibacterial drugs in a matrix of isosensitest broth. A double solid-phase extraction procedure is described in which trimethoprim and dibromopropamidine isethionate together with 4-chlorophenylbiguanide as internal standard are freed from endogenous components by a cation-exchange extraction cartridge and subsequently removed and individually separated by reversed-phase ion-pair chromatography. Sulfadiazine, sulfamerazine and p-aminobenzoic acid, unretained by ion exchange, are similarly isolated for chromatography by adsorption on a CH-bonded phase cartridge and individually assayed using the same chromatographic system. The rationale of the pre-treatment and chromatography is described and the quantitative aspects of the analyses of selected combinations of these drugs are reported.

Separation and quantification of murein and precursors from Enterobacter cloacae after treatment with trimethoprim and sulphadiazine

Abstract— The intracellular concentrations of the soluble murein precursors UDP‐Mur‐NAc‐pentapeptide in the cytoplasm, the membrane‐bound lipid precursor disaccharide pentapeptide and the muropeptides of Enterobacter cloacae cultures treated with trimethoprim (12·5μg mL−1) and sulphadiazine (250 μg mL−1) were determined by using capillary zone electrophoresis analysis. In the presence of trimethoprim, UDP‐Mur‐NAc‐pentapeptide as well as disaccharide pentapeptide accumulated. In the case of sulphadiazine‐treated cells, the concentration of UDP‐Mur‐NAc‐pentapeptide roughly paralleled the control cells but sulphadiazine caused a slow incremental accumulation of disaccharide pentapeptide. The muropeptide composition of the murein indicated that the differences between the peptidoglycans produced by the control cells and the cells grown in the presence of either trimethoprim or sulphadiazine alone or in combination were quite marked. The results suggest that the enhanced activity of trimethoprim plus sulphadiazine against E. cloacae is caused by an additional effect on the inhibition of the bacterial peptidoglycan biosynthesis and that this additional effect is a fundamental part of the antibacterial action of the antimetabolites. This effect leads to changes of cell morphology and resultant changes in bacterial cell permeability.