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It is known that many diabetic patients experience testicular atrophy. This study sought to investigate the effect of 4-hexylresorcinol (4HR) on testicular function in rats with streptozotocin (STZ)-induced diabetes, focusing on testicular weight, sperm motility, histological alterations, and serum testosterone levels to understand the efficacy of 4HR on testes. Our findings reveal that 4HR treatment significantly improves testicular health in diabetic rats. Notably, the STZ group exhibited a testicular weight of 1.22 ± 0.48 g, whereas the STZ/4HR group showed a significantly enhanced weight of 1.91 ± 0.26 g ( p < 0.001), aligning closely with the control group's weight of 1.99 ± 0.17 g and the 4HR group's weight of 2.05 ± 0.24 g, indicating no significant difference between control and 4HR groups ( p > 0.05). Furthermore, the STZ/4HR group demonstrated significantly improved sperm motility compared to the STZ group, with apoptotic indicators notably reduced in the STZ/4HR group relative to the STZ group ( p < 0.05). These results underscore the therapeutic potential of 4HR for maintaining testicular function under diabetic conditions.

We studied the possibility of using 4-hexylresorcinol to increase the efficiency of anti-mycobacterial chemotherapy. In an in vitro experiment, 4-hexylresorcinol increased the efficiency of rifampicin, kanamycin, and isoniazid against Mycobacterium smegmatis by 3-5 times. Experiments in sanitation of BALB/c mice infected with M. smegmatis showed the best efficacy of the isoniazid and 4-hexylresorcinol combination in comparison with isoniazid monotherapy. The growth-inhibiting activity of the combination of antibiotic rifabutin with 4-hexylresorcinol was shown on 6 strains of M. tuberculosis. A 2-fold decrease in the minimum inhibitory concentration of this antibiotic in the presence of half-minimum inhibitory concentration of 4-hexylresorcinol was demonstrated for monoresistant strain M. tuberculosis 5360/42H r . On the mouse model of experimental tuberculosis caused by M. tuberculosis H37Rv, a 5-fold decrease in lung contamination and more rapid complete cure were achieved in animals treated with the combination of rifabutin and 4-hexylresorcinol in comparison with rifabutin monotherapy., (© 2024. Springer Science+Business Media, LLC, part of Springer Nature.)

To augment the functional attributes of pectin and expand its prospective utilization in food preservation, this research explored the enzymatic grafting of resorcinol and 4-hexylresorcinol onto pectin. Structural analysis verified the successful grafting of both resorcinol and 4-hexylresorcinol to pectin via esterification, with the 1-OH of resorcinol and 4-hexylresorcinol and the carboxyl group of pectin functioning as grafting sites. The grafting ratios of resorcinol-modified pectin (Re-Pe) and 4-hexylresorcinol-modified pectin (He-Pe) were 17.84 % and 10.98 %, respectively. This grafting modification notably enhanced the antioxidative and antibacterial properties of pectin. Specifically, DPPH clearance and the inhibition ratio in the β-carotene bleaching assay increased from 11.38 % and 20.13 % (native pectin, Na-Pe) to 41.15 % and 36.67 % (Re-Pe), and 74.72 % and 53.40 % (He-Pe). Moreover, the inhibition zone diameter against Escherichia coli and Staphylococcus aureus rose from 10.12 and 10.08 mm (Na-Pe) to 12.36 and 11.52 mm (Re-Pe), and 16.78 and 14.87 mm (He-Pe). Additionally, the application of native and modified pectin coatings effectively impeded pork spoilage, with the modified pectins demonstrating a more potent effect. Among the two modified pectins, He-Pe exhibited the most significant enhancement in pork shelf life., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Silk sericin is a degumming product used by the silk industry. The degumming process can affect the protein structure and molecular weight of silk sericin. The present study examined how pretreatment with 4-hexylresorcinol (4HR) affects the biomedical properties of silk sericin. Before the degumming process, silkworm cocoons were treated with 4HR solution. The protein structure of the final degumming product was evaluated by Fourier transform infrared spectroscopy (FT-IR) and scanning electron microscopy. Untreated silk sericin (S) and silk sericin pretreated with 4HR (S+4HR) were added to RAW264.7 cells, and the expression of BMP-2 was determined. The bone-regenerating capacity of S+4HR was evaluated using the critical-sized rat calvarial defect model. Compared with S, S+4HR showed an increase in β-sheet structures. Administration of S+4HR to RAW264.7 cells increased expression of BMP-2, mainly via the TLR-mediated signaling pathway. Bone volume, as measured by micro-computerized tomography, was significantly greater in the S+4HR group than in the S, gelatin alone, and unfilled control groups (p < 0.05 each). Expression of BMP-2 and runx2 in tissue specimens was significantly higher following treatment with S+4HR than with S (p < 0.05). Taken together, these findings show that 4HR pretreatment before the degumming process increased the β-sheet structure of silk sericin, as well as inducing BMP-2 expression and bone regeneration ability.

The p53 mutation is inherent in over 50% of human cancers. In head and neck squamous cell carcinoma, the p53 mutation is associated with a poor prognosis. 4‑Hexylresorcinol (4HR) is a pharmacologic chaperone. The present study aimed to investigate the effect of 4HR on p53 transcriptional activity in oral carcinoma cells with p53 mutations. To identify conformational changes induced by 4HR administration, peptides including the DNA‑binding domain from mutant and wild‑type p53 were synthesized, and Fourier transform infrared spectroscopy was performed. To determine the effect of 4HR on p53 mutant carcinoma cells, western blot analysis, p53 transcriptional activity analysis, MTT assay and apoptosis immunocytochemistry were performed. The YD‑15 cell line has a mutation in the DNA binding domain of p53 (Glu258Ala). When p53 Ala‑258 was coupled by 4HR, the p53 Ala‑258 structure lost its original conformation and approached a conformation similar to that of p53 Glu‑258. In the cell experiments, 4HR administration to p53 mutant cells increased p53 transcriptional activity and the expression levels of apoptosis‑associated proteins such as B‑cell lymphoma 2 (BCL2), BCL2‑associated X (BAX) and BCL2‑associated agonist of cell death (BAD). Accordingly, 4HR administration on YD‑15 cells decreased cell viability and increased apoptosis. In conclusion, 4HR is a potential substance for use in the recovery of loss‑of‑function in mutant p53 as a pharmacologic chaperone.

Background: 4-Hexylresorcinol (4HR) is a food additive and class I histone deacetylase inhibitor. In this study, we examined the effects of 4HR administration on the submandibular gland in a growing rat model., Methods: Four-week-old rats were used in this study. The experimental group (nine males and eight females) received 12.8 mg/kg of 4HR weekly for 12 weeks. Ten rats (five males and five females) were used as controls. The submandibular glands of rats were collected 12 weeks after the first administration of 4HR. The weight of the glands was measured. Histological analysis, immunoprecipitation-high-performance liquid chromatography (IP-HPLC), and western blotting were performed., Results: The weights of the rat submandibular glands were higher in the experimental groups than in the control group, especially in male rats (P < 0.05). The vascular endothelial growth factor (VEGF) and testosterone in the submandibular glands were more highly expressed in 4HR-treated male rats than in untreated rats, as detected by both western blotting and immunohistochemistry. The IP-HPLC results demonstrated that the expression levels of Ki67, epidermal growth factor, and testosterone in the submandibular glands were higher in 4HR-treated male rats than in untreated rats., Conclusions: This study demonstrated that the systemic administration of 4HR increased the weight of submandibular glands in male rats. In addition, the testosterone and VEGF expression levels in the submandibular glands increased owing to 4HR administration., (© 2022. The Author(s).)

4-Hexylresorcinol (4HR) has been used as a food additive, however, it has been recently demonstrated as a Class I histone deacetylase inhibitor (HDACi). Unlike other HDACi, 4HR can be taken through foods. Unfortunately, some HDACi have an influence on craniofacial growth, therefore, the purpose of this study was to evaluate the effects of 4HR on craniofacial growth. Saos-2 cells (osteoblast-like cells) were used for the evaluation of HDACi and its associated activities after 4HR administration. For the evaluation of craniofacial growth, 12.8 mg/kg of 4HR was administered weekly to 4 week old rats (male: 10, female: 10) for 12 weeks. Ten rats were used for untreated control (males: 5, females: 5). Body weight was recorded every week. Serum and head samples were collected at 12 weeks after initial administration. Craniofacial growth was evaluated by micro-computerized tomography. Serum was used for ELISA (testosterone and estrogen) and immunoprecipitation high-performance liquid chromatography (IP-HPLC). The administration of 4HR (1-100 μM) showed significant HDACi activity ( p < 0.05). Body weight was significantly different in male rats ( p < 0.05), and mandibular size was significantly smaller in 4HR-treated male rats with reduced testosterone levels. However, the mandibular size was significantly higher in 4HR treated female rats with increased growth hormone levels. In conclusion, 4HR had HDACi activity in Saos-2 cells. The administration of 4HR on growing rats showed different responses in body weight and mandibular size between sexes.

High efficiency of a combined preparation including synergistic polymyxin B and 4-hexylresorcinol was shown for treatment of experimental sepsis caused by an antibiotic-resistant highly virulent hypermucoid Klebsiella pneumoniae strain KPM9Pm r in mice. Complex therapy with polymyxin B (1 mg/kg) and 4-hexylresorcinol (30 mg/kg) led to cure in 80%; in 20% of these mice, no bacterial cells were found. After treatment with polymyxin B alone, only 50% animals survived and all of them contained bacterial cells. Comparative analysis of the results of monotherapy and combined treatment indicates that 4-hexylresorcinol not only increases the efficiency of antibiotic, but also minimizes persistence of the infection agent and therefore, the risk of development of antibiotic resistance., (© 2021. Springer Science+Business Media, LLC, part of Springer Nature.)

Inhibition of α-glucosidase as well as non-enzymatic glycation is thought as an effective method for treating type-2 diabetes mellitus. In this study, we investigated the inhibitory potential and mechanism of 4-hexylresorcinol against α-glucosidase and non-enzymatic glycation by using multispectroscopic analyses and molecular docking. The results of enzyme kinetics showed that 4-hexylresorcinol reversibly inhibited α-glucosidase activity in a noncompetitive way. Fluorescence quenching then revealed that it increased the hydrophobicity of α-glucosidase and changed the conformation of the enzyme by forming the α-glucosidase-hexylresorcinol complex. Thermodynamic analysis and molecular docking further demonstrated that the inhibition of 4-hexylresorcinol on the α-glucosidase was mainly dependent on hydrogen bond and hydrophobic interaction. Moreover, the 4-hexylresorcinol moderately inhibited the formation of fructosamine, and strongly suppressed the generation of α-dicarbonyl compounds and advanced glycation end products (AGEs). The interaction between 4-hexylresorcinol and bovine serum albumin was mainly driven by hydrophobic interaction. This study showed a novel inhibitor of α-glucosidase as well as non-enzymatic glycation, and provided a drug candidate for the prevention and treatment of type-2 diabetes., (Copyright © 2020 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

4-Hexylresorcinol (4HR) is used as a food preservative and an ingredient of toothpaste and cosmetics. The present study was performed using 233 antisera to determine the changes in protein expression induced by 4HR in human umbilical cord vein endothelial cells (HUVECs), and evaluated the 4HR-induced effects in comparison with previous results (Kim et al., 2019). Similar to RAW 264.7 cells, 4HR-treated HUVECs showed decreases in the expression of the proliferation-related proteins, cMyc/MAX/MAD network proteins, p53/RB and Wnt/β-catenin signaling, and they showed inactivation of DNA transcription and protein translation compared to the untreated controls. 4HR upregulated growth factors (TGF-β1, β2, β3, SMAD2/3, SMAD4, HGF-α, Met, IGF-1) and RAS signaling proteins (RAF-B, p38, p-p38, p-ERK-1, and Rab-1), and induced stronger expression of the cellular protection-, survival-, and differentiation-related proteins in HUVECs than in RAW 264.7 cells. 4HR suppressed NFkB signaling in a manner that suggests potential anti-inflammatory and wound healing effects by reducing M1 macrophage polarization and increasing M2 macrophage polarization in both cells. 4HR-treated HUVECs tended to increase the ER stress mediators by upregulating eIF2AK3, ATF4, ATF6, lysozyme, and LC3 and downregulating eIF2α and GADD153 (CHOP), resulting in PARP-1/AIF-mediated apoptosis. These results indicate that 4HR has similar effects on the protein expression of HUVECs and RAW 264.7 cells, but their protein expression levels differ according to cell types. The 4HR-treated cells showed global protein expression characteristic of anticancer and wound healing effects, which could be alleviated simultaneously by other proteins exerting opposite functions. These results suggest that although 4HR has similar effects on the global protein expression of HUVECs and RAW 264.7 cells, the 4HR-induced molecular interferences in those cells are complex enough to produce variable protein expression, leading different cell functions. Moreover, HUVECs have stronger wound healing potential to overcome the impact induced by 4HR than RAW 264.7 cells., Competing Interests: NO authors have competing interests.

Ever decreasing efficiency of antibiotic treatment due to growing antibiotic resistance of pathogenic bacteria is a critical issue in clinical practice. The two generally accepted major approaches to this problem are the search for new antibiotics and the development of antibiotic adjuvants to enhance the antimicrobial activity of known compounds. It was therefore the aim of the present study to test whether alkylresorcinols, a class of phenolic lipids, can be used as adjuvants to potentiate the effect of various classes of antibiotics. Alkylresorcinols were combined with 12 clinically used antibiotics. Growth-inhibiting activity against a broad range of pro- and eukaryotic microorganisms was determined. Test organisms did comprise 10 bacterial and 2 fungal collection strains, including E. coli and S. aureus, and clinical isolates of K. pneumoniae. The highest adjuvant activity was observed in the case of 4-hexylresorcinol (4-HR), a natural compound found in plants with antimicrobial activity. 50% of the minimal inhibitory concentration (MIC) of 4-HR caused an up to 50-fold decrease in the MIC of antibiotics of various classes. Application of 4-HR as an adjuvant revealed its efficiency against germination of bacterial dormant forms (spores) and prevented formation of antibiotic-tolerant persister cells. Using an in vivo mouse model of K. pneumoniae-induced sepsis, we could demonstrate that the combination of 4-HR and polymyxin was highly effective. 75% of animals were free of infection after treatment as compared to none of the animals receiving the antibiotic alone. We conclude that alkylresorcinols such as 4-HR can be used as an adjuvant to increase the efficiency of several known antibiotics. We suggest that by this approach the risk for development of genetically determined antibiotic resistance can be minimized due to the multimodal mode of action of 4-HR., Competing Interests: The authors have declared that no competing interests exist.

Surgical methods for accelerating orthodontic tooth movement are limited by possible damage to the tooth root and patient discomfort. 4-Hexylresorcinol (4HR) has been shown to increase bone remodeling and may potentially facilitate tooth movement. This study investigated the (1) effect of 4HR administration on osteoblast-like cells and (2) effect of 4HR administration on tooth movement in ovariectomized rats. Saos-2 cells were treated with either 4HR or solvent (control). Protein expression levels were investigated 2, 8, and 24 h after treatment. Thirty ovariectomized Sprague-Dawley rats were divided into two experimental groups (A and B) and one control group. After installation of an orthodontic tooth movement device, groups A and B received subcutaneous weekly injections of 4HR (1.28 and 128 mg/kg). Micro-computerized tomography and histological analyses were performed after 2 weeks of tooth movement. The application of 4HR elevated expression of osteogenic markers in Saos-2 cells. Movement of the first molars was significantly greater in rats administered 4HR. Furthermore, the expression of bone morphogenic protein-2, receptor activator of nuclear factor kappa-B ligand, osteocalcin, and tartrate-resistant acid phosphatase were increased after 4HR administration. 4HR application demonstrated increased expression of osteogenic markers in Saos-2 cells and accelerated orthodontic tooth movement in rats.

Angiogenesis plays an important role in active inflammation and wound healing. Our results showed that silk sericin and 4-hexylresorcinol (4HR) increased vascular endothelial growth factor (VEGF) expression in a dose-dependent manner in RAW264.7 cells. Unlike 4HR, silk sericin increased the expression of hypoxia inducible factor-1α (HIF-1α) and HIF-2α. Pretreatment with an HIF inhibitor decreased the sericin-induced increase in VEGF expression. However, the HIF inhibitor did not affect the 4HR-induced increase in VEGF expression. An inhibitor of matrix metalloproteinase (MMP) declined the 4HR-induced increase in VEGF expression. Silk sericin increased production of reactive oxygen species (ROS), whereas 4HR decreased ROS. M1 markers were increased by silk sericin treatment, and M2 markers were increased by 4HR treatment. VEGF and angiogenin expression were higher in rats treated with a 4HR-incorporated silk mat than in rats treated with a silk mat alone. In conclusion, silk sericin and 4HR increased VEGF expression in RAW264.7 cells via HIF-mediated and MMP-mediated pathways, respectively. Silk sericin exerted like pro-oxidant effects and 4HR exerted anti-oxidant effects. Rats treated with a 4HR-incorporated silk mat showed higher levels of VEGF and angiogenin than those treated with a silk mat alone.

4-Hexylresorcinol (4HR) is a small organic compound that is used as an additive antiseptic and antioxidant, but its molecular properties have not been clearly elucidated. The present study explored the cellular effects of 4HR on RAW 264.7 cells by immunoprecipitation high-performance liquid chromatography (IP-HPLC) using 216 antisera. 4HR-treated cells showed significant decreases in the expressions of proliferation-related proteins, cMyc/MAX/MAD network, p53/Rb/E2F and Wnt/β-catenin signalings, epigenetic modifications, and protein translation. Furthermore, 4HR suppressed the expressions of growth factors and proteins associated with RAS signaling, NFkB signaling, inflammation, and osteogenesis, but elevated the expressions of proteins associated with p53-mediated and FAS-mediated apoptosis, T-cell immunity, angiogenesis, antioxidant, and oncogenic signaling. In a 4HR adherence assay, TNFα, PKC, osteopontin, and GADD45 were strongly adherent to 4HR-coated beads, whereas IL-6, c-caspase 3, CDK4, and c-caspase 9 were not. Many 4HR adherent proteins were expressed at lower levels in 4HR treated RAW 264.7 cells than in non-treated controls, whereas 4HR non-adherent proteins were expressed at higher levels. These observations suggest 4HR affects the expressions of proteins in an adhesion-dependent manner and that its effects on proteins are characteristic and global in RAW 264.7 cells.

Objective: The objectives of this study were to evaluate the suppression of the nuclear factor kappa B (NF-kB) pathway by 4-hexylresorcinol (4HR), which was activated by tumor necrosis factor-α (TNF-α) in osteoblasts, and new bone formation by 4HR-incorporated porcine bone in an animal model., Study Design: For the confirmation of successful incorporation of 4HR into porcine bone, scanning electron microscopy (SEM) and Fourier transform-infrared (FT-IR) analysis were performed. High performance liquid chromatography was performed for the analysis of the 4HR release profile from porcine bone. MC 3T3-E1 cells were used for the analysis of the NF-kB signaling pathway activation by western blotting and real-time reverse transcriptase polymerase chain reaction. New bone formation and the analysis of marker protein expression were studied in a rat calvarial critical-sized defect model., Results: Both SEM and FT-IR analysis demonstrated successful incorporation of 4HR into porcine bone. Approximately 30% of 4HR was steadily released from porcine bone for 18 days. 4HR suppressed the NF-kB signaling pathway, which was activated by TNF-α application in MC 3T3-E1 cells. Histological analysis revealed that porcine bone particles with incorporated 4HR showed significantly greater new bone formation than those without 4HR at 4 and 8 weeks after operation (P < 0.05). The expression intensities of alkaline phosphatase, osteoprotegerin, and osteocalcin were also higher in the 4HR-incorporated group., Conclusion: The application of 4HR suppressed the NF-kB signaling pathway in osteoblasts and 4HR-containing porcine bone particles promoted new bone formation in a rat calvarial defect model.

Silk suture material is primarily composed of silk fibroin and regarded as a non-resorbable material. It is slowly degraded by proteolysis when it is implanted into the body. 4-Hexylresorcinol (4HR) is a well-known antiseptic. In this study, the biodegradability of 4HR-incorporated silk sutures were compared to that of untreated silk sutures and polyglactin 910 sutures, a commercially available resorbable suture. 4HR-incorporated silk sutures exhibited anti-microbial properties. Matrix metalloproteinase (MMP) can digest a wide spectrum of proteins. 4HR increased MMP-2, -3, and -9 expression in RAW264.7 cells. MMP-2, -3, and -9 were able to digest not only silk fibroin but also silk sutures. Consequently, 59.5% of the 4HR-incorporated silk suture material remained at 11 weeks after grafting, which was similar to that of polyglactin 910 degradation (56.4% remained). The residual amount of bare silk suture material at 11 weeks after grafting was 91.5%. The expression levels of MMP-2, -3 and -9 were high in the 4HR-incorporated silk suture-implanted site 12 weeks after implantation. In conclusion, 4HR-treated silk sutures exhibited anti-microbial properties and a similar level of bio-degradation to polyglactin 910 sutures and induced higher expression of MMP-2, -3, and -9 in macrophages.

Objective: The objective of this study was to evaluate (1) the effect of 4-hexylresorcinol (4HR) application on TNF-α expression in RAW264.7 cells and (2) the effect of 4HR ointment on burn wound healing in a rat burn wound model., Materials and Methods: RAW264.7 cells were cultured in the presence of different concentrations of 4HR, and the attached cells were sampled for western blot analysis at each time point. Animal studies were conducted on male Wistar rats (n=24). Three treatment groups were evaluated in this study: (1) treatment with ointment alone (negative control), (2) treatment with ointment containing 0.2wt% 4HR, and (3) treatment with ointment containing 2wt% 4HR. For the assessment of wound healing, digital photographs were taken at 1, 5, and 14 days. Animals were sacrificed at 1, 5, and 14 days for histological examination or ELISA analysis., Results: Western blot analysis showed that TNF-α protein levels were reduced following treatment with 4HR at concentrations between 1 and 10μg/mL. In animal studies, the average denuded area 14 days after burn was smaller in the 2wt% 4HR treatment group compared with the control group (P=0.022). Immunohistochemical analysis revealed lower TNF-α expression in the 2wt% 4HR treatment group compared with the control group 14 days after burn (P=0.027)., Conclusion: 4HR reduced TNF-α expression in RAW264.7 cells and in the burn wounds of rats. Burn wounds dressed with ointment containing 4HR also exhibited rapid epithelization and collagen regeneration., (Copyright © 2016 Elsevier Ltd and ISBI. All rights reserved.)

Insecticidal effects of 4-hexylresorcinol, a phenoloxidase (PO) inhibitor, were determined on Hyphantria cunea (Drury) under laboratory conditions. The LC50 for the 15-d-old larvae was estimated to be 2.95 g/L after 96 h exposure. The activities of glutathione S-transferase (GST) and PO showed a decrease in larvae treated with 4-hexylresorcinol, and the IC50 of GST and PO were estimated to be 0.8 and 0.43 g/L, respectively, 24 h after treatment. The PO from the hemolymph of fall webworm was purified by ammonium sulfate precipitation, gel-filtration, and ion-exchange chromatography, and then enzymatic characteristics and the mechanism of inhibition were determined using L-dihydroxyphenylalanine (L-DOPA) as the substrate. The purified PO showed a single band on SDS-PAGE with a molecular weight of about 70 kDa. The optimum pH for PO activity was observed at pH 7.0, optimum temperature was found to be 45 °C, and PO activity was strongly inhibited by Zn(2+) . IC50 values were estimated to be 8.2, 19.14, and 24.04 μmol/L for 4-hexylresorsinol, kojic acid, and quercetin, respectively. The inhibitory potencies (i.e., I50 of each compound/I50 of 4-hexylresorcinol) of kojic acid and quercetin on H. cunea PO were estimated to be 1.87 and 2.89, respectively. 4-hexylresorcinol was determined to be a competitive inhibitor, and kojic acid and quercetin were determined to be mixed inhibitors. PO is one of the most important enzymes in an insect's immune system, and the use of PO inhibitors seems to be a promising approach for pest control due to their potential safety for humans., (© 2014 Institute of Zoology, Chinese Academy of Sciences.)

Growth of members of most of the studied genera of gram-positive (Dietzia, Kocuria, and Rhodo- coccus) and gram-negative bacteria (Pseudomonas and Chromobacterium) in biofilms exhibited higher resistance to an translation inhibitor, azithromycin compared to the growth of planktonic cultures of the same strains. Low concentrations of azithromycin were found to stimulate biofilm formation by the studied saprotrophic strains. The rate of synthesis of the polysaccharide matrix component exceeded the rate of cell growth, indicating implementation of the biofilm phenotype under these conditions. It was found that an alkylhydroxybenzene (AHB) compound 4-hexylresorcinol was capable of almost uniform suppression of growth of both planktonic cultures and biofilms of the saprotrophic strains under study. In some cases, combined action ofazithromycin and AHB resulted in an additive inhibitory effect and prevented the stimulation of biofilm growth by subinhibitory azithromycin concentrations. Thus, AHB may be considered a promising antibiofilm agent.

Grafted macromolecules often induce granuloma formation with foreign body giant cell (FBGC) infiltration, and this is the main reason for graft failure. Diacylglycerol kinase (DAGK) is an important intracellular mediator of FBGC formation in macrophages. In this study, 4-hexylresorcinol (4HR) inhibited DAGKδ in a macrophage cell line (RAW264.7 cells). As a result of DAGK-δ inhibition by 4HR, FBGC formation was significantly inhibited in RAW264.7 cells. Silk fibroin is a well-known natural macromolecule, and when it is grafted into bone defects, it results in granuloma formation with massive FBGC formation. 4HR-incorporating silk graft materials displayed significant reduction of granuloma formation and increases in the extent of new bone formation in a rabbit calvarial defect model. In conclusion, 4HR could inhibit foreign body reaction via a DAGK-mediated pathway., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The present study was undertaken to examine the effects of cisplatin plus 4-hexylresorcinol (4-HR) combination therapy on oral mucosal melanoma (OMM) using cultured primary OMM cells in a tumour xenograft model. Cultured primary OMM cells were used for the MTT assay and DNA microarray. OMM cells were implanted into the submandibular glands of nude mice. The mice were then treated with cisplatin only or cisplatin plus 4-HR. Tumour size changes, survival rate and tumour metastasis were compared between the two groups by observation, micro-positron emission tomography (PET) and histological examination. In the MTT assay, the cisplatin plus 4-HR group showed significantly higher inhibition of OMM cell growth compared to the other groups (p<0.05). DNA microarray results showed significant inhibition of matrix metalloproteinase (MMP)-2 gene expression upon 4-HR application. The necropsy and micro-PET results showed that the mice from the cisplatin-only group had more distant metastases than the mice from the cisplatin plus 4-HR combination group (p=0.002). MMP-2 expression was lower in the primary tumours in the cisplatin plus 4-HR combination group than in the cisplatin-only group (p<0.001). Overall survival was longer in mice from the cisplatin plus 4-HR combination group than in the cisplatin-only group (p=0.049). In conclusion, the combined effect of cisplatin and 4-HR resulted in fewer metastases and longer survival than cisplatin-only treatment in the OMM xenograft model.

The bacterial dormancy-inducing factor 4-hexylresorcinol (4-HR) has been shown to have synergistic antitumor effects when used in combination with cisplatin. In the present study, 4-HR was used as a single agent in the squamous carcinoma cell line SCC-9. The results demonstrated that 4-HR suppressed SCC-9 cell proliferation compared to primary cultured gingival fibroblasts. 4-HR dose-dependently induced SCC-9 cell apoptosis as determined by caspase-3 activity, annexin V expression, as well as by scanning and transmission electron microscopy. 4-HR inhibited intracellular calcium oscillation in both SCC-9 cells and normal human dermal fibroblasts. 4-HR-induced apoptosis was partly reversed by calcium channel blockers. Of note, 4-HR reduced the tumor mass formed by SSC-9 cell implantation in BALB/cAnNCrj-nu/nu mice and mass size reduction was also partly reversed by the concomitant application of calcium channel blockers. Collectively, our results suggest that 4-HR has strong antitumor effects by inhibiting calcium channel oscillation and inducing apoptosis.

DNA topoisomerases are highly exploited targets for antimicrobial drugs. The spread of antibiotic resistance represents a significant threat to public health and necessitates the discovery of inhibitors that target topoisomerases in novel ways. However, the traditional assays for topoisomerase activity are not suitable for the high-throughput approaches necessary for drug discovery. In this study we validate a novel assay for screening topoisomerase inhibitors. A library of 960 compounds was screened against Escherichia coli DNA gyrase and archaeal Methanosarcina mazei DNA topoisomerase VI. Several novel inhibitors were identified for both enzymes, and subsequently characterised in vitro and in vivo. Inhibitors from the M. mazei topoisomerase VI screen were tested for their ability to inhibit Arabidopsis topoisomerase VI in planta. The data from this work present new options for antibiotic drug discovery and provide insight into the mechanism of topoisomerase VI.

Inhibitors of phenoloxidase are used routinely to characterise the structural and functional properties of phenoloxidases. Hemocyanin-derived phenoloxidase activity is also sensitive to standard phenoloxidase inhibitors. In this study, we characterise the effects of a number of phenoloxidase inhibitors on hemocyanin-derived phenoloxidase activity from the chelicerate, Limulus polyphemus. Both inhibition type and K(i) values were similar to those observed for hemocyanin-derived phenoloxidase from another chelicerate, Eurypelma californicum. In addition, substrate inhibition was observed at concentrations above 2mM dopamine. The conformation in which two of the inhibitors, namely tropolone and kojic acid, would bind near the Cu(II) centre of hemocyanin is proposed., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The dormancy-inducing factors of bacteria inhibit tumor cell growth. In the present study, we evaluated the antitumor effects of the dormancy-inducing factor 4-hexylresorcinol (4-HR) using real-time cell electronic sensing (RT-CES) in SCC-9 cells (tongue squamous cell carcinoma cells). Treatment with 4-HR suppressed the growth of SCC-9 cells in a dose-dependent manner. We used a DNA microarray to identify genes that showed a significant change in expression upon 4-HR administration in SCC-9 cells. Among the differentially expressed genes, the protein expression of several cell proliferation related factors, including E2F1, E2F2, E2F3, E2F4, E2F5, E2F6, Sp1 and Sp3, were determined by western blot analyses. Treatment with 4-HR strongly suppressed E2F2 and slightly suppressed E2F3 but did not change the expression of E2F1, E2F4, E2F5 and E2F6 relative to no treatment. Furthermore, 4-HR increased Sp1 expression in a dose-dependent manner and decreased Sp3 expression. Therefore, the ratio of Sp1 to Sp3, an important driving force of epithelial cell differentiation, was drastically increased. Consistent with this observation, 4-HR increased the expression of the epithelial cell differentiation markers involucrin and keratin 10. Together, our results indicate that 4-HR induces the differentiation of SCC-9 via the modulation of the E2F-mediated signaling pathway.

Lozenges for the treatment of sore throat provide relief of discomfort in cases of oral inflammation. This effect has not been fully explained so far. Here, we have examined the proposition that key components of pharmaceutical preparations for the treatment of sore throat which are routinely regarded antiseptics might have sodium channel-blocking, i.e. local anaesthetic-like effects. We investigated the effects of hexylresorcinol, amylmetacresol and dichloro-benzylalcohol on voltage-operated neuronal (Na(V)1.2) sodium channels heterologously expressed in HEK 293 cells in vitro. Hexylresorcinol, amylmetacresol and dichloro-benzylalcohol reversibly blocked depolarisation-induced whole-cell sodium inward currents. The half-maximum blocking concentrations (EC(50)) at -150 mV were 23.1, 53.6 and 661.6 microM, respectively. Block induced by hexylresorcinol and amylmetacresol was increased at depolarised potentials and use-dependent during trains of depolarisations applied at high frequency (100 Hz) indicating that both drugs bind more tightly to inactivated conformations of the channel. Estimates for the inactivated state affinity were 1.88 and 35 microM for hexylresorcinol and amylmetacresol, respectively. Hexylresorcinol and amylmetacresol are 10-20 fold more potent than the local anaesthetic lidocaine in blocking sodium inward current. Both drugs show an increased effect at depolarised membrane potentials or in conditions of high-frequency discharges.

The effect of hexylresorcinol (HR), a chemical analogue of microbial anabiosis autoinducers of the alkylhydroxybenzene (AHB) group, on the stability of biological membranes and monolamellar liposomes formed of egg phosphatidylcholine (ePC) was studied. According to spectrophotometry and electron microscopy studying of HR-loaded liposomes in the presence of a surface-active agent Tween 20, the critical ratio between HR and ePC for liposome preservation was found to be close to equimolar. The trends in HR influence on membrane structural organization and stability confirmed in experiments on liposomes were also reproduced on intact bacterial cells explaining non-species-specific effect of AHBs. The demonstrated high efficiency of AHB biocides may be used in material and equipment protection against biocorrosion.

Aim: To investigate the modifying effect of hexylresorcinol chemical analog on anti-lysozyme activity of bacteria., Materials and Methods: Influence of hexylresorcinol on anti-lysozyme activity (ALA) of bacteriawas studied on the examples of such strains as Bacillus cereus 279, Klebsiella pneumoniae 278, Escherichia coli 101, which are able to inactivate lysozyme. As a chemical analog of microbial regulators, C6-alkyloxybenzol (C6-AOB) was used, which was introduced in growth medium containing microorganisms, which than were cultivated until entering in the stationary growth phase. Isolation of clones was carried out by seeding the 24-hours broth culture on solid growth medium followed by measurement of ALA with photometric method., Results: Modifying effect of chemical analog of extracellular microbial regulators (C6-AOB) on population structure of bacteria on the sign of anti-lysozyme activity was revealed. Maximal effect was observed at C6-AOB concentrations 1 and 10 mcg/ml. Clonal redistribution on ALA manifested by decrease of proportion of clones with high and intermediate values of ALA and increase of proportion of clones with low ALA or its absence., Conclusion: Obtained data are perspective for development of measures for control of persisting pathogenic microorganisms by modification of persistence factors with microbial autoregulators.

The effect of pretreatment with pyrophosphate and 4-hexylresorcinol in combination with modified atmosphere packaging (MAP) (80% CO(2), 10% O(2), 10% N(2), or 80% CO(2), 20% N(2)) on the quality of white shrimp during storage at 4 degrees C was investigated. Shrimp pretreated with 2% pyrophosphate and 0.25% 4-hexylresorcinol and stored under MAP showed the lower microbiological and chemical deteriorations as evidenced by delayed microbial growth as well as lower trimethylamine (TMA) and total volatile base nitrogen (TVB) production (P < 0.05). Additionally, the growth of coliforms was inhibited effectively. White shrimp pretreated with 4-hexylresorcinol had the lower melanosis throughout the storage compared with those without treatment (P < 0.05). This was associated with the lowered polyphenol oxidase (PPO) activity in shrimp treated with 4-hexylresorcinol. Therefore, the effective retardation of microbiological and chemical deterioration of white shrimp stored under MAP with the decrease in melanosis could be achieved by pretreatment of the shrimp with pyrophosphate and 4-hexylresorcinol. Furthermore, decapitation could be another means to lower the microbial load and melanosis in white shrimp, particularly those stored under MAP.

The effects of ascorbic acid (AA) and 4-hexylresorcinol (4-HR) on pear polyphenoloxidase (PPO) activity and stability have been investigated in vitro. AA does not interact directly with PPO but prevents browning by reducing oxidized substrates. The 4-HR exerts a dual role on PPO. If no substrates are present, it interacts preferably with the deoxy form of PPO inactivating it. If substrates and 4-HR are both present they compete for the catalytic site. The 4-HR behaves then as a canonical enzyme inhibitor, binding to the met form of PPO. Simultaneous addition of 4-HR and AA has synergistic inhibition or inactivatory effects depending on the presence or the absence of PPO substrates.

The anthelmintic activity of the drupe extracts of Melia azedarach L. (Meliaceae) growing in Argentina was tested against tapeworms, hookworms, nodular worms and earthworms, and was shown to be better than the standards piperazine phosphate and hexylresorcinol against tapeworms and hookworms, respectively., (Copyright (c) 2006 John Wiley & Sons, Ltd.)

The fact of long-term preservation of the physicochemical properties of DNA molecules in aqueous solutions in complexes with methylresorcinol, hexylresorcinol, and tyrosol, the chemical analogues of microbial autoregulators (d1 factors) from the group of alkylhydroxybenzenes (AOB), was established. Compared to the control variants of storage of aqueous DNA solutions, the AOB influence consisted in the sum of correlating effects: the prevention of DNA degradation (according to spectrophotometric parameters) and the preservation of its viscous characteristics and electrophoretic mobility. The initial DNA properties were preserved to the greatest degree in the presence of hexylresorcinol, the compound with the longest alkyl radical. Possible mechanisms of the protective action of alkylhydroxybenzenes in relation to DNA are discussed, namely, the prevention of its hydrolysis due to isolation from the aqueous environment and maintaining DNA stability in the dormant forms of microorganisms.

We established that chemical analogues of alkylhydroxybenzenes (AHB), belonging to alkylresorcinols and functioning as microbial autoregulatory d1 factors, enhance the UV resistance of various DNA molecules of different origin and conformation. These include the linear DNA of the lambda phage, bovine spleen DNA, and the DNA of the pUC19 plasmid that is composed of a number of annular (supercoiled and relaxed) and linearized molecules. Irradiating DNA with UV light (lambda = 254 nm) in the presence of methylresorcinol (MR) or hexylresorcinol (HR) results in comparatively insignificant DNA destruction as evidenced by our data on the electrophoretic mobility pattern in agarose gel. Using the linear Hind III restricts of the lambda phage DNA, we revealed that the protective effect of AHB varies depending on their chemical structure (it is more manifest with HR than MR) and concentration. Importantly, the effect of HR on bovine spleen DNA was based on its protective activity and manifested itself after a long incubation period. Studies using the pUC19 plasmid demonstrated that AHB, apart from increasing the resistance of linearized DNA molecules to UV irradiation, prevented both the supercoiled annular-supercoiled relaxed and the supercoiled relaxed-linearized transitions. The possible mechanisms of the UV-protective effect of AHB on DNA and their contributions to the resistance of dormant microbial forms to environmental factors are discussed.

We conducted a comparative study of the effects of alpha-amino-gamma-butyrolactone, the common structural element of extracellular microbial regulators of the homoserine lactone (HSL) group, and of 4-n-hexylresorcinol, an autoregulator of the alkylhydroxybenzene (AHB) group, on the growth and development of gram-positive and gram-negative bacteria. We revealed non-species-specific effects of HSL and AHB and characterized their concentration dependencies. The addition of 10(-5)-10(-3) M HSL or 10(-5)-10(-4) M AHB during the exponential growth phase of the cultures grown on balanced media resulted in cell division arrest and accelerated the transition to the stationary phase that culminated in endospore formation in Bacillus cereus, Alicyclobacillus tolerans, and Sulfobacillus thermosulfidooxidans. When bacilli grew under the cultivation conditions that resulted in a low-zero spore percentage, 10(-4)-10(-3) M HSL cancelled the inhibition of spore formation. In the gram-negative bacteria Pseudomonas aurantiaca and Azotobacter vinelandii, AHB at concentrations of 10(-4) to (1.5-2.5) 10(-4) M induced the formation of dormant cells. Studies with the actinobacterium Streptomyces avermitilis revealed that the HSL effect varied depending on the age of the test cultures. The addition of 10(-4) M HSL during the lag phase of a submerged streptomycete culture accelerated its transition to the stationary phase and induced the formation of endospores, the dormant cells that are regarded as alternatives to exospores (conidia). If HSL (3.64 and 4.55 mg per 1cm2 disc) was locally added to a surface S. avermitilis culture, the growing mycelium formed rings that differed in their density, in the extent of the development of aerial mycelium, and in the presence/absence of exospores. Ring-shaped growth of streptomycete mycelia was also induced by 0.075-0.75 mg of AHB; however, unlike HSL, AHB repressed exospore formation. The data on non-species-specific effects of HSL and AHB suggest that they may perform regulatory functions on the microbial community level.

It has been shown that the phenotypic dissociation of Bacillus subtilis SK1 and S. typhimurium TA100 is induced by hexylresorcinol, an exogenous non-species-specific autoregulator of pleiotropic action, which is genotoxic for both pro- and eukaryotes. Nongenotoxic homoserine lactone, a chemical analogue of cell-density-responsive species-specific regulators, does not induce bacterial dissociation. The phage resistance of the S- and R-type variants of S. typhimurium TA100 induced by hexylresorcinol has been found to be the same as that of the S- and R-type salmonella variants obtained by the routine subculturing method.

We revealed a relationship between alkylhydroxybenzene (AHB)-induced changes in the structural organization of supramolecular complexes (SC) of the DNA of Pseudomonas auraniaca and the phenotypic dissociation of this bacterium. The addition of 0.1-0.3 mM hexylresorcinol (C6-AHB), a chemical analogue of microbial anabiosis autoinducers, caused the formation of cystlike refractile cells (CRC) in these gram-negative, nonsporulating bacteria. Inoculating pseudomonad CRC on solid nutrient media resulted in phenotypic dissociation of the microbial population that yielded several variants with different colony structure and morphology. This manifested itself in the conversion of the original S-colony-forming phenotype into the R form and in the formation of less pigmented colonies. These transitions were possibly linked to AHB-induced structural changes in the DNA. In vitro studies revealed that AHB could interact with DNA SC, resulting in their structural modification that manifested itself in changes in their elastoviscosity. DNA supramolecular complexes isolated from proliferating, stationary-phase, and anabiotic P. aurantiaca cells differed in their elastoviscosity and capacity to interact with AHB homologues with different hydrophobicity, such as hexylresorcinol and methylresorcinol (C1-AHB). The DNA SC from actively proliferating cells were characterized by smaller elastoviscosity compared with those from stationary-phase and anabiotic cells, due to the difference in the DNA superspiralization degree and the physiological age of the bacteria involved. C6-AHB produced a pronounced relaxing effect on the DNA SC from exponential-phase P. aurantiaca cells. The less hydrophobic C1-AHB produced a similar effect on the DNA SC from stationary-phase cells. The curve of the dose-effect dependence of C6-AHB had a breaking point within the submillimolar (10(-4) M) concentration range. These concentrations induce the formation of cystlike anabiotic pseudomonad cells that are characterized by an unstable genotype and dissociate into distinct variants upon inoculation on solid media.

A controlled atmosphere containing 48% CO2 and 7% O2 was used in association with refrigeration for storage of deepwater pink shrimp (Parapenaeus longirostris). Shrimp were treated with two different concentrations of sodium metabisulfite or 4-hexylresorcinol and subjected to the controlled atmosphere immediately after capture onboard ship or on arrival in port. Total volatile basic nitrogen, total viable counts, enterobacteria, lactic acid bacteria, and luminescent bacteria were determined, and black spot progression was evaluated. The combined effect of controlled atmosphere and melanosis inhibitors was used to delay black spot development as compared to the shrimp stored in ice alone. Storage under the controlled atmosphere without ice limited microbiological quality, namely, total viable counts, but enterobacterial growth was lower.

The microbial alkylhydroxybenzenes (AHB), autoinducers of anabiosis, or d1 factors, participate in stress response of mycelial fungi, as determined from changes in intracellular Ca2+ concentration. By using the genetically modified strain Aspergillus awamori 66A, which produces a recombinant Ca2+-dependent protein aequorin, the dynamics of Ca2+ was studied in the cytosol of cells exposed to mechanical shock in the presence of the protective doses (0.001-0.01% w/vol) of a chemical AHB analogue, 4-n-hexylresorcinol. Like under stressful conditions, Ca2+ concentration increases in the cell cytosol in response to enhanced AHB level in a growing fungal culture; thus, AHB is perceived by cells as a stress signal. The level of cell response, which was determined from the amplitude of luminescence dependent on the Ca2+ concentration in cytosol was related to the physiological age of the cells and AHB concentration. Micromycete preincubation with AHB was found to protect cells from subsequent stress; this was reflected in the Ca2+ response. The protective AHB effect was manifested as (1) a significant decrease in the amplitude of luminescence and, thus, in Ca2+ accumulation in the cytosol during subsequent mechanical stress (as compared to the control--mechanical stress only); (2) development of the secondary Ca2+ response, which was not observed in the control; (3) a high level of Ca2+ retained in the cytosol for a long time in the presence of AHB (as compared to the control without preincubation with AHB). The mechanisms underlying the AHB effect on the Ca2+ transport systems are discussed.

The effects of hexylresorcinol and dodecylresorcinol on the monophenolase and diphenolase activity of mushroom tyrosinase have been studied. The results show that hexylresorcinol and dodecylresorcinol can inhibit both monophenolase and diphenolase activity of the enzyme. The lag period of the enzyme was obviously lengthened, and the steady-state activity of the enzyme decreased sharply. Two microM of hexylresorcinol and dodecylresorcinol can lengthen the lag period from 98 s to 260 and 275 s, respectively. Both hexylresorcinol and dodecylresorcinol can lead to reversible inhibition of the enzyme. The IC50 values of hexylresorcinol and dodecylresorcinol were estimated as 1.24 and 1.15 microM for monophenolase and as 0.85 and 0.80 microM for diphenolase, respectively. A kinetic analysis shows that hexylresorcinol and dodecylresorcinol are competitive inhibitors. The apparent inhibition constant for hexylresorcinol and dodecylresorcinol binding with free enzyme has been determined to be 0.443 and 0.405 microM for diphenolase, respectively.

The protective effects of resveratrol and 4-hexylresorcinol against oxidative DNA damage in human lymphocytes induced by hydrogen peroxide were investigated. Resveratrol and 4-hexylresorcinol showed no cytotoxicity to human lymphocytes at the tested concentration (10-100 microM). In addition, DNA damage in human lymphocytes induced by H2O2 was inhibited by resveratrol and 4-hexylresorcinol. Resveratrol and 4-hexylresorcinol at concentrations of 10-100 microM induced an increase in glutathione (GSH) levels in a concentration-dependent manner. Moreover, these two compounds also induced activity of glutathione peroxidase (GPX) and glutathione reductase (GR). The activity of glutathione-S-transferase (GST) in human lymphocytes was induced by resveratrol. Resveratrol and 4-hexylresorcinol inhibited the activity of catalase (CAT). These data indicate that the inhibition of resveratrol and 4-hexylresorcinol on oxidative DNA damage in human lymphocytes induced by H2O2 might be attributed to increase levels of GSH and modulation of antioxidant enzymes (GPX, GR and GST).

The mutagenic activity of chemical analogues of microbial anabiosis autoinducers (the autoregulatory d1 factors of cell differentiation), which act to inhibit cell proliferation, to enhance cell tolerance, and to induce the transition of cells to anabiotic state, was studied using the Ames test. In the range of concentrations studied (0.1 to 100 micrograms/ml), alkyl-substituted hydroxybenzenes (AHBs) differing in hydrophobicity, i.e., methylresorcinol (C1-AHB) and hexylresorcinol (C6-AHB), as well as unsubstituted resorcinol, showed different growth-inhibiting and mutagenic effects. C6-AHB was found to inhibit the growth of Salmonella typhimurium TA100 and to induce its mutagenesis at a rate of 1.8 revertants/nmol. C1-AHB taken at low concentrations not only failed to inhibit bacterial growth but even stimulated it and exerted an antimutagenic effect. Unsubstituted resorcinol virtually did not influence bacterial growth and showed weak mutagenic activity. The growth-inhibiting effect of elevated C6-AHB concentrations correlated with the degree of the transition of the original phenotype producing S-type colonies to a phenotype producing R-type colonies. The role of AHB homologues, as microbial autoregulators with mutagenic activity, in the regulation and correlation of two processes (the phenotypic dissociation of microbial populations and the formation of resting microbial forms) is discussed. The accumulation of AHBs in senescent microbial cultures may induce adaptive mutations, change the expression of genes, and promote the development of minor cell subpopulations (phenotypes), thus providing for the adaptation of these cultures to varying environmental conditions.

Pressure inactivation of mushroom PPO was studied for pH values ranging from 4 to 8, and the effect of some antibrowning agents on the pressure stability of mushroom PPO at pH 6.5 was evaluated. pH reduction below 6.5 resulted in a lowered inactivation threshold pressure and an increase of the absolute value of the activation volume (or a decrease of the z(p) value), the latter two parameters reflecting the pressure dependency of the inactivation rate constant. An increase in pH from 6.5 to 8, on the other hand, did only marginally affect the pressure stability of the enzyme. Mushroom PPO at pH 6.5 was markedly sensitized toward pressure by the presence of 2.5 mM 4-hexylresorcinol and slightly stabilized by the presence of 5 mM EDTA. The presence of 5 mM glutathione, sodium chloride, or benzoic acid caused no significant alteration of the enzyme pressure stability. Only in the presence of 4-hexylresorcinol, significant changes of the activation volume and z(p) value were noticed.

The present study demonstrates the antiradical efficiency of fisetin, a flavonol widely distributed in fruits and vegetables, by its ability to react with two different free radicals, ABTS; and DPPH;. The polyphenolic nature of fisetin led us to consider whether it might be oxidised by polyphenol oxidase (PPO), and the results reported show that it can be oxidised by PPO extracted and partially purified from broad bean seeds. The reaction was followed by recording spectral changes with time, with maximal spectral changes being observed at 282 nm (increase in absorbance) and at 362 nm (decrease). The presence of two isosbectic points (at 265 and 304 nm) suggested that only one absorbent product was formed. These spectral changes were not observed in the absence of PPO. The oxidation rate varied with the pH, reaching its highest value at pH 5.5. The fisetin oxidation rate increased in the presence of sodium dodecyl sulfate, an activator of polyphenol oxidase. Maximal activity was obtained at 0.87 mM sodium dodecyl sulfate. The following kinetic parameters were determined: Vmax=49 microM/min, Km=0.6 mM, Vmax/Km=8.2x10-2 min-1. Flavonol oxidation was inhibited by selective PPO inhibitors such as cinnamic acid (a classical competitive inhibitor, Ki=1.4 mM) and 4-hexylresorcinol, which behaved as a slow-binding inhibitor. The results reported show that fisetin oxidation was strictly dependent on the presence of polyphenol oxidase.

The amebicides dehydroemetine and chloroquine diphosphate and the anthelmintic pyrvinium pamoate, previously reported to be mutagenic in Salmonella typhimurium (Cortinas de Nava et al., 1983), were clearly shown to be genotoxic in the Escherichia coli pol A+/pol A- assay. Two other antiparasitic drugs, diiodohydroxyquin and 4-hexylresorcinol, were also found to be genotoxic in E. coli, while iodochlorhydroxyquin preferentially inhibited the pol A+ strain. From the 3 alternative testing methods employed, the liquid suspension succeeded in detecting 5 antiparasitic drugs as genotoxic; the microsuspension identified 2, and the disc diffusion method only 1. However, the metabolic activation system could only be coupled successfully and in a reproducible way to the microsuspension assay.

The bactericidal effect of hexylresorcinol was assessed using bacterial aggregates. Increased drug concentration resulted in decreased survival of bacteria when the aggregates were exposed to hexylresorcinol for at least eight hours. This indicates that this drug may be effective against dental plaque only when available for long periods of time.

Effects of piperazine derivatives, especially of diethylcarbamazine (DEC) on adult Angiostrongylus cantonensis and Dirofilaria immitis were examined. Piperazine (3 X 10(-5)-10(-4) M) paralyzed A. cantonensis and the action was antagonized by picrotoxin. 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP) (10(-5)-10(-4) M) caused contraction but little effect was produced by strychnine. An inhibitory effect on untreated preparations was caused by lower concentrations (3 X 10(-6)-10(-5) M) of diethylcarbamazine citrate (DEC) and also on the preparations contracted by eserine. A stimulatory effect was also seen when higher concentrations (10(-4)-3 X 10(-4) M) of this drug were applied to both preparations. The inhibitory action of DEC was antagonized by gabergic antagonists such as picrotoxin and bicuculline, but not by alpha-adrenergic antagonists like dibenamine and phentolamine. When the worm preparation was paralyzed by strychnine or hexylresorcinol (inhibitors of the release of acetylcholine in this worm), the stimulatory effect of DEC was blocked, but pyrantel (a nicotinic cholinergic agonist) contracted the paralyzed preparation. However, the effect of DEC on D. immitis (10(-7)-3 X 10(-4) M) was inhibitory, and this action was also antagonized by picrotoxin. These results suggest that the DEC inhibitory and stimulatory action is through the gabergic and cholinergic mechanisms in adult A. cantonensis and D. immitis.