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Pediatric drug formulations of sodium benzoate: I. Coated granules with a hydrophilic binder.

High doses of sodium benzoate are applied in the treatment of some rare metabolic disorders. In most cases children are affected who often refuse the oral uptake of sodium benzoate as a powder or in solution due to its bad taste. Therefore, small-sized, saliva-resistant microcapsules have been developed containing high doses of the drug substance. Granules were produced by roller compacting of sodium benzoate powder without any additives, by solvent-free cold extrusion and hot-melt extrusion adding poly(ethylene glycol)s of different grades. The granules with a diameter of less than 1 mm were film-coated by an ethanolic solution of Eudragit E 100. The microcapsules from hot-melt extrusion containing 25% Macrogol 4000 were most stable during the coating process and showed the highest yields. Sodium benzoate is completely released from the microcapsules within 9 min into 0.1 N HCl and 0.01 N HCl whereas dissolution into buffer pH 6.8 is different in the initial phase and completed after 14 min. The bad taste of sodium benzoate is not recognized in the buccal space for at least 5 min. The microcapsules are stable during storage for at least 6 months.

Capsules↗

Ivermectin is better than benzyl benzoate for childhood scabies in developing countries.

OBJECTIVE: To compare single dose oral ivermectin with topical benzyl benzoate for the treatment of paediatric scabies. METHODS: An observer-blinded randomized controlled trial was undertaken at Vila Central Hospital, Vanuatu. One hundred and ten children aged from 6 months to 14 years were randomized to receive either ivermectin 200 micro g/kg orally or 10% benzyl benzoate topically. Follow up was at 3 weeks post-treatment. Primary outcome measures were the number of scabies lesions, the itch visual analogue score and nocturnal itch. Secondary outcome measures were the skin's reaction to treatment, the passage of worms in stool and other side effects. RESULTS: Eighty patients completed the study protocol. There was no significant difference between the two treatments; both produced a significant decrease in the number of scabies lesions seen at follow up. Ivermectin cured 24 out of 43 patients (56%), and benzyl benzoate 19 out of 37 patients (51%) at 3 weeks post-treatment. No serious side effects were noted with either treatment, but benzyl benzoate was more likely to produce local skin reactions (P = 0.004, OR 6.4, 95% CI 1.6-25.0) CONCLUSIONS: Ivermectin is cheap and effective in the treatment of paediatric scabies. Ivermectin has minimal observed toxicity and has the additional beneficial effects of antiparasitic action in onchocerciasis, filariasis and strongyloidiasis. Ivermectin is better than benzyl benzoate for the treatment of paediatric scabies in developing countries.

Adolescent↗

Use of stable isotopes to identify benzoate as a metabolite of benzene degradation in a sulphidogenic consortium.

An enriched sulphidogenic consortium capable of mineralizing benzene was used to study the metabolic pathway of anaerobic benzene degradation. Benzoate was detected in active cultures and benzene was confirmed to be the source of this benzoate by the addition of deuterated benzene (D6) and subsequent detection of deuterated benzoate (D5) in active cultures but not in autoclaved controls. Benzoate was utilized by this culture at 1/12 the rate of benzene, while its presence did not inhibit benzene utilization. The benzene utilization rate was reduced, however, in the presence of 2-fluorobenzoate. When the culture was supplemented with [(13)C]-bicarbonate, the carboxyl group on benzoate was not labelled with [(13)C]-carbon, suggesting that this transformation relies on a more complex set of reactions than simple addition of carbonate.

Bacteria↗

Chitosan potentiates the antimicrobial action of sodium benzoate on spoilage yeasts.

AIMS: The objective of this study was to determine whether low concentrations of chitosan and benzoate in combination could be used to enhance the antimicrobial action of either compound alone against three spoilage yeasts in saline solutions. METHODS AND RESULTS: Saccharomyces exiguus, Saccharomycodes ludwigii and Torulaspora delbrueckii were suspended in 0.05 and 0.005% chitosan glutamate and 0.025% sodium benzoate, alone or in combination, in 0.9% saline solutions at pH 6.2 and 4.5. Survivor curves were constructed from viable counts determined periodically for up to 120 min. Chitosan at 0.005% almost doubled the extent of death caused by 0.025% benzoate alone, from about 1-2 log to about 2-4 log cfu ml(-1), depending on pH and target organism. CONCLUSIONS: Chitosan (0.005%) and 0.025% sodium benzoate acted synergistically against spoilage yeasts in saline solutions. SIGNIFICANCE AND IMPACT OF THE STUDY: These results suggest that the natural compound chitosan may be useful as an adjunct in the potentiation of the biocidal efficacy of antimicrobial compounds such as benzoates.

Chitin↗

Final report on the safety assessment of Benzyl Alcohol, Benzoic Acid, and Sodium Benzoate.

Benzyl Alcohol is an aromatic alcohol used in a wide variety of cosmetic formulations as a fragrance component, preservative, solvent, and viscosity-decreasing agent. Benzoic Acid is an aromatic acid used in a wide variety of cosmetics as a pH adjuster and preservative. Sodium Benzoate is the sodium salt of Benzoic Acid used as a preservative, also in a wide range of cosmetic product types. Benzyl Alcohol is metabolized to Benzoic Acid, which reacts with glycine and excreted as hippuric acid in the human body. Acceptable daily intakes were established by the World Health Organization at 5 mg/kg for Benzyl Alcohol, Benzoic Acid, and Sodium Benzoate. Benzoic Acid and Sodium Benzoate are generally recognized as safe in foods according to the U.S. Food and Drug Administration. No adverse effects of Benzyl Alcohol were seen in chronic exposure animal studies using rats and mice. Effects of Benzoic Acid and Sodium Benzoate in chronic exposure animal studies were limited to reduced feed intake and reduced growth. Some differences between control and Benzyl Alcohol-treated populations were noted in one reproductive toxicity study using mice, but these were limited to lower maternal body weights and decreased mean litter weights. Another study also noted that fetal weight was decreased compared to controls, but a third study showed no differences between control and Benzyl Alcohol-treated groups. Benzoic Acid was associated with an increased number of resorptions and malformations in hamsters, but there were no reproductive or developmental toxicty findings in studies using mice and rats exposed to Sodium Benzoate, and, likewise, Benzoic Acid was negative in two rat studies. Genotoxicity tests for these ingredients were mostly negative, but there were some assays that were positive. Carcinogenicity studies, however, were negative. Clinical data indicated that these ingredients can produce nonimmunologic contact urticaria and nonimmunologic immediate contact reactions, characterized by the appearance of wheals, erythema, and pruritus. In one study, 5% Benzyl Alcohol elicited a reaction, and in another study, 2% Benzoic Acid did likewise. Benzyl Alcohol, however, was not a sensitizer at 10%, nor was Benzoic Acid a sensitizer at 2%. Recognizing that the nonimmunologic reactions are strictly cutaneous, likely involving a cholinergic mechanism, it was concluded that these ingredients could be used safely at concentrations up to 5%, but that manufacturers should consider the nonimmunologic phenomena when using these ingredients in cosmetic formulations designed for infants and children. Additionally, Benzyl Alcohol was considered safe up to 10% for use in hair dyes. The limited body exposure, the duration of use, and the frequency of use were considered in concluding that the nonimmunologic reactions would not be a concern. Because of the wide variety of product types in which these ingredients may be used, it is likely that inhalation may be a route of exposure. The available safety tests are not considered sufficient to support the safety of these ingredients in formulations where inhalation is a route of exposure. Inhalation toxicity data are needed to complete the safety assessment of these ingredients where inhalation can occur.

Animals↗

Active transport of benzoate in Pseudomonas putida.

Benzoate uptake in Pseudomonas putida is mediated by an active transport system capable of accumulating benzoate against a 150-fold concentration gradient when subsequent metabolism is blocked by mutation. Initial benzoate transport rates are inhibited by CCCP, sodium azide, arsenate and DCCD. Uptake is stimulated by including a respirable carbon source during preincubation of the bacteria. The initial uptake rate and the ATP pool levels are not correlated and no periplasmic components were found to bind benzoate. These observations indicate that benzoate uptake is energized by the membrane potential, rather than by ATP hydrolysis.

Adenosine Triphosphate↗

Regulation of circadian methyl benzoate emission in diurnally and nocturnally emitting plants.

Emission of methyl benzoate, one of the most abundant scent compounds of bee-pollinated snapdragon flowers, occurs in a rhythmic manner, with maximum emission during the day, and coincides with the foraging activity of bumblebees. Rhythmic emission of methyl benzoate displays a "free-running" cycle in the absence of environmental cues (in continuous dark or continuous light), indicating the circadian nature of diurnal rhythmicity. Methyl benzoate is produced in upper and lower snapdragon petal lobes by enzymatic methylation of benzoic acid in the reaction catalyzed by S-adenosyl-L-methionine:benzoic acid carboxyl methyltransferase (BAMT). When a detailed time-course analysis of BAMT activity in upper and lower petal lobes during a 48-hr period was performed, high BAMT activity was found at night as well as in continuous darkness, indicating that the BAMT activity is not an oscillation-determining factor. Analysis of the level of benzoic acid during a 24-hr period revealed oscillations in the amount of benzoic acid during the daily light/dark cycle that were retained in continuous darkness. These data clearly show that the total amount of substrate (benzoic acid) in the cell is involved in the regulation of the rhythmic emission of methyl benzoate. Our results also suggest that similar molecular mechanisms are involved in the regulation of methyl benzoate production in diurnally (snapdragon) and nocturnally (tobacco and petunia) emitting plants.

Activity Cycles↗

Developmental regulation of methyl benzoate biosynthesis and emission in snapdragon flowers.

In snapdragon flowers, the volatile ester methyl benzoate is the most abundant scent compound. It is synthesized by and emitted from only the upper and lower lobes of petals, where pollinators (bumblebees) come in contact with the flower. Emission of methyl benzoate occurs in a rhythmic manner, with maximum emission during the day, which correlates with pollinator activity. A novel S-adenosyl-l-methionine:benzoic acid carboxyl methyl transferase (BAMT), the final enzyme in the biosynthesis of methyl benzoate, and its corresponding cDNA have been isolated and characterized. The complete amino acid sequence of the BAMT protein has only low levels of sequence similarity to other previously characterized proteins, including plant O-methyl transferases. During the life span of the flower, the levels of methyl benzoate emission, BAMT activity, BAMT gene expression, and the amounts of BAMT protein and benzoic acid are developmentally and differentially regulated. Linear regression analysis revealed that production of methyl benzoate is regulated by the amount of benzoic acid and the amount of BAMT protein, which in turn is regulated at the transcriptional level.

Amino Acid Sequence↗

Tartrazine and benzoate challenge and dietary avoidance in chronic asthma.

This study undertook to determine the usefulness of tartrazine and benzoate challenge and dietary avoidance in the management of patients with chronic asthma. Double-blind ingestion-challenge tests were performed on separate days with lactose, tartrazine, benzoate and acetylsalicyclic acid (ASA). Of the twenty-eight subjects challenged, one responded to tartrazine and one to benzoate. Two additional subjects responded to ASA and a further eight were not tested with this material because of a definite history of sensitivity. Twenty-four subjects completed 1 month periods of observation while first on a normal diet and then while on a tartrazine-benzoate avoidance diet. No improvement occurred during the modified diet in anyone with positive challenge-tests or in all, but one, of those with a history of ASA idiosyncrasy; paradoxically, several of these subjects worsened during this period. We conclude that tartrazine-benzoate dietary avoidance was not of value in the management of the chronic asthmatic in this study, even among patients who respond to challenge with these substances or have ASA idiosyncrasy.

Adrenal Cortex Hormones↗

Gene clusters involved in anaerobic benzoate degradation of Geobacter metallireducens.

The degradation of aromatic compounds follows different biochemical principles in aerobic and anaerobic microorganisms. While aerobes dearomatize and cleave the aromatic ring by oxygenases, facultative anaerobes utilize an ATP-dependent ring reductase for the dearomatization of the activated key intermediate benzoyl-coenzyme A (CoA). In this work, the aromatic metabolism was studied in the obligately anaerobic model organism Geobacter metallireducens. The gene coding for a putative carboxylic acid-CoA ligase was heterologously overexpressed and the gene product was characterized as a highly specific benzoate-CoA ligase catalysing the initial step of benzoate metabolism. However, no evidence for the presence of an ATP-dependent benzoyl-CoA reductase as observed in facultative anaerobes was obtained. In a proteomic approach benzoate-induced proteins were identified; the corresponding genes are organized in two clusters comprising 44 genes. Induction of representative genes during growth on benzoate was confirmed by reverse transcription polymerase chain reaction. The results obtained suggest that benzoate is activated to benzoyl-CoA, which is then reductively dearomatized to cyclohexa-1,5-diene-1-carbonyl-CoA, followed by beta-oxidation reactions to acetyl-CoA units, as in facultatively anaerobic bacteria. However, in G. metallireducens the process of reductive benzene ring dearomatization appears to be catalysed by a set of completely different protein components comprising putative molybdenum and selenocysteine containing enzymes.

Acyl Coenzyme A↗

Moment analysis of metabolic heterogeneity: conjugation of benzoate with glycine in rat liver studied by multiple indicator dilution technique.

Metabolic zonation was assessed with the multiple indicator dilution (MID) technique in the single-pass perfused rat liver with use of moment analysis of the formed metabolite (M) data. During single-pass, retrograde rat liver perfusion with 17 Awake M benzoate, a bolus containing tracer preformed metabolite (PM) [(3)H]hippurate was injected rapidly into the hepatic vein at 20 min postperfusion, followed by injection of a second bolus containing [(14)C]benzoate at 30 min. Both doses also contained noneliminated reference indicators ((51)Cr-labeled RBCs, (125)I-labeled albumin, [(14)C]- or [(3)H]sucrose, and (2)H(2)O). The steady-state extraction ratio of benzoate, the area under the curve (AUC) and its mean transit time (MTT) during retrograde flow were identical to those previously observed for prograde flow. Values of AUC(PM) and MTT(PM) and AUC(M) were also similar to previously published prograde data, but the MTT(M) with retrograde perfusion was smaller than that for prograde perfusion. This, according to theory based on the tubes-in-series model, was consistent with perivenous enrichment of glycination activity when transport of drug was even and when the ratio of drug influx/efflux coefficient exceeded that for metabolite. Similar benzoate transport in periportal, homogeneous and perivenous isolated rat hepatocytes existed, and the influx/efflux coefficients (partition ratio) of benzoate from MID indeed exceeded that of hippurate. However, metabolism by zonal hepatocytes failed to reveal the anticipated metabolic zonation, and this is likely due to the shallow gradient of metabolic activity. The study demonstrates that moment theory is useful in delineating the perivenous enrichment of glycine conjugation activity.

Animals↗

Cloning and expression of the benzoate dioxygenase genes from Rhodococcus sp. strain 19070.

The bopXYZ genes from the gram-positive bacterium Rhodococcus sp. strain 19070 encode a broad-substrate-specific benzoate dioxygenase. Expression of the BopXY terminal oxygenase enabled Escherichia coli to convert benzoate or anthranilate (2-aminobenzoate) to a nonaromatic cis-diol or catechol, respectively. This expression system also rapidly transformed m-toluate (3-methylbenzoate) to an unidentified product. In contrast, 2-chlorobenzoate was not a good substrate. The BopXYZ dioxygenase was homologous to the chromosomally encoded benzoate dioxygenase (BenABC) and the plasmid-encoded toluate dioxygenase (XylXYZ) of gram-negative acinetobacters and pseudomonads. Pulsed-field gel electrophoresis failed to identify any plasmid in Rhodococcus sp. strain 19070. Catechol 1,2- and 2,3-dioxygenase activity indicated that strain 19070 possesses both meta- and ortho-cleavage degradative pathways, which are associated in pseudomonads with the xyl and ben genes, respectively. Open reading frames downstream of bopXYZ, designated bopL and bopK, resembled genes encoding cis-diol dehydrogenases and benzoate transporters, respectively. The bop genes were in the same order as the chromosomal ben genes of P. putida PRS2000. The deduced sequences of BopXY were 50 to 60% identical to the corresponding proteins of benzoate and toluate dioxygenases. The reductase components of these latter dioxygenases, BenC and XylZ, are 201 residues shorter than the deduced BopZ sequence. As predicted from the sequence, expression of BopZ in E. coli yielded an approximately 60-kDa protein whose presence corresponded to increased cytochrome c reductase activity. While the N-terminal region of BopZ was approximately 50% identical in sequence to the entire BenC or XylZ reductases, the C terminus was unlike other known protein sequences.

Benzoates↗

Biphenyl and benzoate metabolism in a genomic context: outlining genome-wide metabolic networks in Burkholderia xenovorans LB400.

We designed and successfully implemented the use of in situ-synthesized 45-mer oligonucleotide DNA microarrays (XeoChips) for genome-wide expression profiling of Burkholderia xenovorans LB400, which is among the best aerobic polychlorinated biphenyl degraders known so far. We conducted differential gene expression profiling during exponential growth on succinate, benzoate, and biphenyl as sole carbon sources and investigated the transcriptome of early-stationary-phase cells grown on biphenyl. Based on these experiments, we outlined metabolic pathways and summarized other cellular functions in the organism relevant for biphenyl and benzoate degradation. All genes previously identified as being directly involved in biphenyl degradation were up-regulated when cells were grown on biphenyl compared to expression in succinate-grown cells. For benzoate degradation, however, genes for an aerobic coenzyme A activation pathway were up-regulated in biphenyl-grown cells, while the pathway for benzoate degradation via hydroxylation was up-regulated in benzoate-grown cells. The early-stationary-phase biphenyl-grown cells showed similar expression of biphenyl pathway genes, but a surprising up-regulation of C(1) metabolic pathway genes was observed. The microarray results were validated by quantitative reverse transcription PCR with a subset of genes of interest. The XeoChips showed a chip-to-chip variation of 13.9%, compared to the 21.6% variation for spotted oligonucleotide microarrays, which is less variation than that typically reported for PCR product microarrays.

Bacterial Proteins↗

Catabolism of benzoate and phthalate in Rhodococcus sp. strain RHA1: redundancies and convergence.

Genomic and proteomic approaches were used to investigate phthalate and benzoate catabolism in Rhodococcus sp. strain RHA1, a polychlorinated biphenyl-degrading actinomycete. Sequence analyses identified genes involved in the catabolism of benzoate (ben) and phthalate (pad), the uptake of phthalate (pat), and two branches of the beta-ketoadipate pathway (catRABC and pcaJIHGBLFR). The regulatory and structural ben genes are separated by genes encoding a cytochrome P450. The pad and pat genes are contained on a catabolic island that is duplicated on plasmids pRHL1 and pRHL2 and includes predicted terephthalate catabolic genes (tpa). Proteomic analyses demonstrated that the beta-ketoadipate pathway is functionally convergent. Specifically, the pad and pat gene products were only detected in phthalate-grown cells. Similarly, the ben and cat gene products were only detected in benzoate-grown cells. However, pca-encoded enzymes were present under both growth conditions. Activity assays for key enzymes confirmed these results. Disruption of pcaL, which encodes a fusion enzyme, abolished growth on phthalate. In contrast, after a lag phase, growth of the mutant on benzoate was similar to that of the wild type. Proteomic analyses revealed 20 proteins in the mutant that were not detected in wild-type cells during growth on benzoate, including a CatD homolog that apparently compensated for loss of PcaL. Analysis of completed bacterial genomes indicates that the convergent beta-ketoadipate pathway and some aspects of its genetic organization are characteristic of rhodococci and related actinomycetes. In contrast, the high redundancy of catabolic pathways and enzymes appears to be unique to RHA1 and may increase its potential to adapt to new carbon sources.

Bacterial Proteins↗

Effect of sodium benzoate on the cutaneous bilirubin content of the adult Gunn rat.

Na benzoate, which dissociates intravascular bilirubin-albumin complexes, might thus have the potential for driving bilirubin from the plasma into tissues such as the skin. We investigated this hypothesis in the adult Gunn rat. As others, we observed an early fall in blood bilirubin concentration after intraperitoneal Na benzoate injection, without any change in skin bilirubin content. Nevertheless, we observed a significant decrease (p less than 0.01) in skin bilirubin content at 24 and then 48 h after Na benzoate injection, at which times blood bilirubin remained at a low level. The phenomenon was reversible since, at 8 days after Na benzoate injection, blood and skin bilirubin contents had returned to control values. Regarding the decrease in skin bilirubin content after Na benzoate injection we described a dose-effect relationship. So, we could not confirm our hypothesis. We concluded that the phenomenon we observed is probably due to a shift in the distribution equilibrium of the pigment between serum, skin and other tissues.

Animals↗

Improvement of aqueous solubility and rectal absorption of 6-mercaptopurine by addition of sodium benzoate.

The solubility of 6-mercaptopurine (6-MP) in water increased as the concentration of sodium benzoate or sodium hippurate in the solution increased. The solubility of 6-MP in 20% (w/v) sodium benzoate or sodium hippurate solution was about 6-fold larger than that of 6-MP alone. The stability constant of the soluble complex of 6-MP with sodium benzoate was estimated to be 2-8 M-1 from (1) phase-solubility study and (2) analysis of chemical shifts observed in 1H-NMR. Partition of 6-MP from the saturated solution to n-octanol was also greatly increased by the addition of sodium benzoate or sodium hippurate, the degree being less in the latter. Administration of 6-MP with 20% (w/v) sodium benzoate to rat rectum resulted in enhanced absorption and the area under the plasma concentration-time curve was comparable to that obtained by intravenous administration (bioavailability = 100%), while the bioavailability after intrarectal administration of 6-MP with 20% (w/v) sodium hippurate was only 9%. The reason for the difference was discussed.

1-Octanol↗

Production of catechol from benzoate by the wild strain Ralstonia species Ba-0323 and characterization of its catechol 1,2-dioxygenase.

Ralstonia sp. Ba-0323, a wild strain isolated from soil, produced catechol from benzoate and accumulated it outside the cells. The bacterium produced a maximal amount of catechol (1.6 mg/ml) from 3 mg/ml of sodium benzoate in a 20-h growing culture. The conversion rate of benzoate to catechol was 70% on a molar basis. The catechol production by the resting cells increased in the presence of glycerol, and the maximal amount of catechol produced from 3 mg/ml of sodium benzoate reached 1.9 mg/ml at the conversion rate of 83% after 8 h of incubation. Catechol 1,2-dioxygenase, which catalyzed the ring cleavage of catechol, was purified to homogeneity from a cell extract of Ralstonia sp. Ba-0323 growing on benzoate and characterized. The specific activity of the purified enzyme was much lower than those of the dioxygenases from other microorganisms reported. The Km for catechol of the purified enzyme was much higher than those of other dioxygenases. In addition, the NH2-terminal amino acid sequence of the enzyme was less similar to the other catechol 1,2-dioxygenases than they are to each other.

Amino Acid Sequence↗

Resistance of yeast flora of labaneh to potassium sorbate and sodium benzoate.

Ten yeast cultures belonging to eight species representing the yeast flora of labaneh were tested for their resistance to potassium sorbate and sodium benzoate. Changes in counts were monitored after 7, 14, and 21 d at 5 degrees C in yeast-free labaneh containing different concentrations of potassium sorbate or sodium benzoate. More than 400 mg/kg of sodium benzoate were needed to limit the counts of Saccharomyces cerevisiae (biovariants 1, 2, and 7) Cryptococcus curvatus, Pichia farinosa, Candida blankii, Debaryomyces hansenii, and Trichosporon brassicae to < or = 10(5) cfu/g after 14 d at 5 degrees C; 150 and 300 mg/kg were needed for Geotrichum candidum and Trichosporon cutaneum, respectively. When potassium sorbate was used, > 400 mg/kg were needed for P. farinosa and D. hansenii; 350 mg/kg were needed for S. cerevisiae biovar 1, Cr. curvatus, and C. blankii; 250 and 200 mg/kg were needed for S. cerevisiae biovariants 2 and 7, respectively; and < 100 mg/kg were needed for T. brassicae, T. cutaneum, and G. candidum. Less than 150 and < 250 mg/kg of potassium sorbate and sodium benzoate, respectively, were needed to limit yeast counts to < 10(5) cfu/g for 7 d at 5 degrees C in two commercial labaneh samples that had initial yeast counts of 4.8 x 10(2) and 9.0 x 10(2) cfu/g; 200 and > 400 mg/kg were required when the storage was extended to 14 d. In commercial labaneh with 9.6 x 10(3) cfu/g, > 300 and > 400 mg/kg of potassium sorbate and sodium benzoate, respectively, were needed to limit the yeast count to < or = 10(5) cfu/g after 7 and 14 d at 5 degrees C.

Benzoates↗