Therapeutic Implications of Phenolic Acids for Ameliorating Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD) is a chronic inflammatory intestinal disorder, and its complex etiology makes prevention and treatment challenging. Research on new drugs and treatment strategies is currently a focal point. Phenolic acids are widely present in plant-based diets and have demonstrated the potential to alleviate colitis due to their powerful antioxidant and anti-inflammatory properties. In this review, we provide an overview of the structures and main dietary sources of phenolic acids, encompassing benzoic acid and cinnamic acid. Additionally, we explore the potential of phenolic acids as a nutritional therapy for preventing and treating IBD. In animal and cell experiments, phenolic acids effectively alleviate IBD induced by drug exposure or genetic defects. The mechanisms include improving intestinal mucosal barrier function, reducing oxidative stress, inhibiting excessive activation of the immune response, and regulating the balance of the intestinal microbiota. Our observation points towards the need for additional basic and clinical investigations on phenolic acids and their derivatives as potential novel therapeutic agents for IBD.

Potent Efficacy of 3-Amino-4-hydroxy Benzoic Acid, a Small Molecule Having Anti-fibrotic Activity, in a Mouse Model of Non-alcoholic Steatohepatitis.

Non-alcoholic steatohepatitis (NASH), which is on the rise due to the increasing obese population and changing lifestyles, causes fibrosis over time and carries the risk of progression to cirrhosis and hepatocellular carcinoma. However, there are no approved effective treatments for NASH. Recent studies suggest that increased lipid metabolism and reduced nitric oxide content are responsible for NASH; 3-amino-4-hydroxy benzoic acid (AHBA) was identified as an inhibitor for the phosphatase activity of soluble epoxy hydrolase, which in turn inhibits lipid metabolism and endothelial nitric oxide synthase activity. The aim of this study was to assess the efficacy of AHBA in a mouse model of NASH. NASH was induced in mice by streptozotocin administration and a high-fat diet loading. The efficacy of AHBA was determined by measuring liver function using serum and liver samples and conducting a morphological assessment. AHBA considerably attenuated the increase in the liver weight and alkaline phosphatase content, which occurred due to the progression of NASH. Hepatocellular steatosis, inflammatory cell infiltration, and hepatocellular ballooning of hepatocytes remained unaltered. In contrast, AHBA treatment significantly ameliorated the fibrotic alterations within liver tissue that were induced by the onset of NASH. These results demonstrate the potential of AHBA as a therapeutic pharmaceutical compound that can treat NASH.

The food and pharmaceutical additive benzoic acid induces amyloid fibrillation of an intrinsically disordered protein.

Benzoic acid (BA) is a microbe-inhibiting flavoring agent used extensively as an additive in foods, pharmaceuticals, and hygiene and cosmetic products. The level of BA in foodstuffs prescribed by world bodies and governmental agencies is assumed to be safe so as to prevent adverse health effects. The safety level of BA is however controversial, and whether different conditions of its use would be generally regarded as safe (GRAS) has been rarely determined. In the quest of how food additives affect the structure and conformation of proteins, this study evaluates the interaction of BA with an intrinsically disordered protein (IDP) at pH 4.2 that matches the pH conditions applicable for the commercial use of benzoate preservatives, and examines its structural transformation by NMR, fluorescence, and high-resolution microscopy. The interaction with BA transforms the protein to a denatured aggregated mesophase that undergoes reconfiguration to yield rigid amyloid fibrils. Significantly, fibrils are observed even with 0.1 mM BA while the recommended level of its use as a preservative is in the 0.4-8 mM range. The discussion refrains from safety comments with no projection of the BA level that could be GRAS.

Sulfonamidyl derivatives of sigmacidin: Protein-protein interaction inhibitors targeting bacterial RNA polymerase and sigma factor interaction exhibiting antimicrobial activity against antibiotic-resistant bacteria.

RNA polymerase is an essential enzyme involved in bacterial transcription, playing a crucial role in RNA synthesis. However, it requires the association with sigma factors to initiate this process. In our previous work, we utilized a structure-based drug discovery approach to create benzoyl and benzyl benzoic acid compounds. These compounds were designed based on the amino acid residues within the key binding site of sigma factors, which are crucial for their interaction with RNA polymerase. By inhibiting bacterial transcription, these compounds exhibited notable antimicrobial activity, and we coined them as sigmacidins to highlight their resemblance to sigma factors and the benzoic acid structure. In this study, we further modified the compound scaffolds and developed a series of sulfonamidyl benzoic acid derivatives. These derivatives displayed potent antimicrobial activity, with minimum inhibitory concentrations (MICs) as low as 1 µg/mL, demonstrating their efficacy against bacteria. Furthermore, these compounds demonstrated low cytotoxicity, indicating their potential as safe antimicrobial agents. To ascertain their mechanism of action in interfering with bacterial transcription, we conducted biochemical and cellular assays. Overall, this study showcases the effectiveness of sulfonamidyl benzoic acid derivatives as antimicrobial agents by targeting protein-protein interactions involving RNA polymerase and sigma factors. Their strong antimicrobial activity and low cytotoxicity implicate their potential in combating antibiotic-resistant bacteria.

Synergy between polypyrrol and benzoic acid against antibiotic-resistant Salmonella spp.

AIMS: The purpose was to characterize Salmonella Heidelberg (SH) and Minnesota (SM) isolates in terms of their resistance and persistence profile and to assess the antimicrobial effect of benzoic acid (BA) and polypyrrole (PPy). METHODS AND RESULTS: The 20 isolates from broiler litter drag swabs were submitted to antibiogram and efflux pump expression. The minimum inhibitory/bactericidal concentration (MIC/MBC) of the compounds, synergistic activity, time kill, biofilm production, presence of related genes, and molecular docking between compounds and bacterial target sites were evaluated. All isolates showed multidrug resistance (MDR) and BA and PPy showed mean MIC (1750 and 342 µg ml-1) and MBC (3167 and 1000 µg ml-1), respectively. None of the isolates expressed an efflux pump. The compounds showed synergism against an SH isolate and reduced the count by 3 logs in the presence of the compounds after 4 h. Most isolates (16/20) produced weak to moderate biofilm and 17 showed genes related to biofilm. The compounds interacted with two essential proteins, 3,4-dihydroxy-2-butanone 4-phosphate synthase proteins and ferritin-like domain-containing protein, in bacterial metabolism at different target sites. CONCLUSIONS: It can be concluded that BA and PPy showed activity on SH and SM, MDR, and biofilm producers, with a potential synergistic effect.

Benzoic acid derivatives as potent antibiofilm agents against Klebsiella pneumoniae biofilm.

Klebsiella pneumoniae is responsible for a significant proportion of human urinary tract infections, and its biofilm is a major virulence. One potential approach to controlling biofilm-associated infections is targeting the adhesin MrkD1P to disrupt biofilm formation. We employed Schrodinger's Maestro tool with the OPLS 2005 force field to dock compounds with the target protein. Two benzoic acid derivatives, 3-hydroxy benzoic acid and 2,5-dihydroxybenzoic acid, had strong binding free energies (-55.57 and -18.68 kcal/mol) and were the most potent compounds. The in-vitro experiments were conducted to validate the in-silico results. The results showed that both compounds effectively inhibited biofilm formation at low concentrations (4 and 8 mg/mL, respectively) and had antibiofilm activity, restricting cell attachment. Both compounds demonstrated a strong biofilm inhibitory effect, with 97% and 89% reduction in biofilm by 3-hydroxy benzoic acid and 2,5-dihydroxybenzoic acid, respectively. These findings suggest that natural compounds can be a potential source of new drugs to combat biofilm-associated infections. The study highlights the potential of targeting adhesin MrkD1P as an effective approach to controlling biofilm-associated infections caused by K. pneumoniae. The results may have implications for the development of new therapies for biofilm-associated infections and pave the way for future research in this area.

Evaluating the potential of natural product combinations with sorbic acid for improving preservative action against food-spoilage yeasts.

Fungal control methods commonly involve the use of antifungals or preservatives, which can raise concerns about broader effects of these stressors on non-target organisms, spread of resistance and regulatory hurdles. Consequently, control methods enabling lower usage of such stressors are highly sought, for example chemical combinations that synergistically inhibit target-organisms. Here, we investigated how well such a principle extends to improving efficacy of an existing but tightly controlled food preservative, sorbic acid. A screen of ∼200 natural products for synergistic fungal inhibition in combinations with sorbic acid, in either 2% or 0.1% (w/v) glucose to simulate high or reduced-sugar foods, did not reveal reproducible synergies in either of the spoilage yeast species Saccharomyces cerevisiae or Zygosaccharomyces bailii. Potentially promising screen candidates (e.g. lactone parthenolide, ethyl maltol) or a small additional panel of rationally-selected compounds (e.g. benzoic acid) all gave Fractional Inhibitory Concentration Indices (FICI) ≥ 0.5 in combinations with sorbic acid, corroborating absence of synergy in either glucose condition (although FICI values did differ between the glucose conditions). Synergies were not achieved either in a tripartite combination with screen candidates or in a soft-drink formulation as matrix. In previous work with other stressors synergy 'hits' have been comparatively frequent, suggesting that sorbic acid could be unusually resistant to forming synergies with other potential inhibitors and this may relate to the weak acid's known multifactorial inhibitory-actions on cells. The study highlights a challenge in developing appropriate natural product or other chemical combinations applicable to food and beverage preservation.

Inhibitory Effects of 3-Cyclopropylmethoxy-4-(difluoromethoxy) Benzoic Acid on TGF-ß1-Induced Epithelial-Mesenchymal Transformation of In Vitro and Bleomycin-Induced Pulmonary Fibrosis In Vivo.

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease characterized by lung inflammation and excessive deposition of extracellular matrix components. Transforming growth factor-ß1 (TGF-ß1) induced epithelial-mesenchymal transformation of type 2 lung epithelial cells leads to excessive extracellular matrix deposition, which plays an important role in fibrosis. Our objective was to evaluate the effects of 3-cyclopropylmethoxy-4-(difluoromethoxy) benzoic acid (DGM) on pulmonary fibrosis and aimed to determine whether EMT plays a key role in the pathogenesis of pulmonary fibrosis and whether EMT can be used as a therapeutic target for DGM therapy to reduce IPF. Firstly, stimulation of in vitro cultured A549 cells to construct EMTs with TGF-ß1. DGM treatment inhibited the expression of proteins such as α-SMA, vimentin, and collagen Ⅰ and increased the expression of E-cadherin. Accordingly, Smad2/3 phosphorylation levels were significantly reduced by DGM treatment. Secondly, models of tracheal instillation of bleomycin and DGM were used to treat rats to demonstrate their therapeutic effects, such as improving lung function, reducing lung inflammation and fibrosis, reducing collagen deposition, and reducing the expression of E-cadherin. In conclusion, DGM attenuates TGF-ß1-induced EMT in A549 cells and bleomycin-induced pulmonary fibrosis in rats.

Comparative effects of benzoic acid and sodium benzoate in diets for nursery pigs on growth performance and acidification of digesta and urine.

The objective of this study was to evaluate the comparative effects of benzoic acid and sodium benzoate in feeds on digesta pH, urinary pH, and growth performance for nursery pigs. A total of 432 pigs (6.9 ±â€…0.9 kg BW) were assigned to eight treatments (6 pigs per pen, replication = 9) in a randomized complete block design with initial body weight (BW) as a block and fed for 41 d in three phases (7/17/17 d, respectively). Treatments were 1) a basal diet (NC), 2) NC + 0.25% bacitracin methylene disalicylate (antibiotic; bacitracin: 250 g/t feed; PC), 3) NC + 0.25% benzoic acid, 4) NC + 0.35% benzoic acid, 5) NC + 0.50% benzoic acid, 6) NC + 0.30% sodium benzoate, 7) NC + 0.40% sodium benzoate, and 8) NC + 0.60% sodium benzoate. Growth performance and fecal scores were measured for each phase. One gilt representing the median BW of each pen was euthanized to collect digesta from the stomach, proximal jejunum, distal jejunum, and cecum, and urine. The PC tended to improve average daily gain (ADG) in phase 1 (P = 0.052) and phase 2 (P = 0.093) as well as average daily feed intake (ADFI) in phase 2 (P = 0.052). Overall, increasing supplemental benzoic acid tended to have a quadratic effect on ADG (P = 0.094), but no difference in ADFI was observed. Increasing supplemental sodium benzoate showed a quadratic effect (P < 0.05) on ADG and linearly increased (P < 0.05) ADFI. Urinary pH linearly decreased (P < 0.05) with increasing supplemental benzoic acid, but was not affected by supplemental sodium benzoate. Increasing supplemental benzoic acid or sodium benzoate linearly increased (P < 0.05) benzoic acid content in digesta of the stomach. Increasing supplemental benzoic acid or sodium benzoate also linearly increased (P < 0.05) urinary hippuric acid. However, the PC did not decrease urinary pH or increase urinary benzoic acid and hippuric acid. With slope-ratio assay using ADG and urinary hippuric acid as dependent variables and benzoic acid intake as an independent variable, the relative bioavailability of benzoic acid compared to sodium benzoate was not different. In conclusion, supplementation of benzoic acid and sodium benzoate could improve the growth performance of nursery pigs. The relative bioavailability of sodium benzoate to benzoic acid of nursery pigs did not differ based on BW gain and urinary hippuric acid.

Newly weaned pigs are exposed to various challenges during the postweaning period, resulting in retarded growth performance. Dietary antibiotics have been used to reduce the negative impacts of weaning stress. However, use of antibiotics in feeds has been phased out in response to concerns associated with microbial resistance. In this study, dietary benzoic acid was supplemented to promote growth performance and increase urinary hippuric acid of nursery pigs. The sodium benzoate may show similar effects with benzoic acid on growth performance and urinary hippuric acid, as sodium benzoate can be highly converted to benzoic acid via the action of gastric acid in stomach. Thus, this study aimed to investigate the effects of increasing benzoic acid and sodium benzoate supplementation on growth performance and acidification of digesta and urine, and to investigate the comparative effects of benzoic acid and sodium benzoate supplemented in diets for nursery pigs. Dietary benzoic acid and sodium benzoate improved body weight gain and increased urinary hippuric acid of nursery pigs. Both sodium benzoate and benzoic acid had similar effects when fed to nursery pigs for their body weight gain and metabolism. Benzoic acid, however, had a stronger effect acidifying urine compared with sodium benzoate.

Effect of benzoic acid, Enterococcus faecium, and essential oil complex on intestinal microbiota of laying hens under coccidia and Clostridium perfringens challenge.

The objective of this study was to investigate whether dietary supplementation with benzoic acid, Enterococcus faecium, and essential oil complex (BEC) could help laying hens recover from coccidia and Clostridium perfringens type A challenge. A total of 60 (35-wk-old) Lohmann-laying hens were randomly assigned to 3 experimental groups (10 replicates with 2 hens per replicate): I) control group (CON), II) challenge group (CC), and III) BEC group (2,000 mg/kg BEC). The total experimental period was 8 wk. The results shown that the challenge layers had lower egg-laying rate and average daily feed intake (ADFI) (P < 0.05), and addition of BEC after challenge increased egg-laying rate (P < 0.05). The content of propionic acid (PA) and butyric acid (BA) in short-chain fatty acid (SCFA) was significantly decreased by challenge (P < 0.05). CC and BEC groups had lower villus height to crypt depth ratio (V/C) and higher pathological scores in duodenum (P < 0.05), whereas the BEC group had lower pathological scores in jejunum when compared with the CC group (P < 0.05). The challenge increased the concentration of proinflammatory cytokines (IL-1ß and IL-6) (P < 0.05). An increase in the abundance of Bacteroidoes (genus), Bacteroidaceae (family), Bacteroidoes sp. Marseille P3166 (species), Bacteroidoes caecicola (species) was observed in the CC group, whereas the BEC group had higher abundance of Bacteroides caecigallinarum (species). The genera Faecalibacterium and Asterolplasma were positively correlated with egg-laying rate (r = 0.57, 0.60; P < 0.01); and the genera Bacteroides and Romboutsia were negatively correlated with egg-laying rate (r = -0.58, -0.74; P < 0.01). The genera Bacteroides, Lactobacillus, and Rombutzia were positively correlated with jejunal mucosa proinflammatory factor IL-1ß level (r = 0.61, 0.60, 0.59; P < 0.01), which were negatively correlated with genera Rikenbacteriaceae RC9, Faecalibacterium, and Olsenlla (r = -0.56, -0.57, -0.61; P < 0.01). There genera UCG.005 was positively correlated with proinflammatory factor IL-6 level in jejunal mucosa (r = 0.58; P < 0.01), which was negatively correlated with Rikenbacteriaceae RC9 (r = -0.62; P < 0.01). The experiment results revealed that the addition of BEC to the diet restored the production performance of the laying hens. In addition, supplementation of 2,000 mg/kg BEC modulated gut health by reducing gut damage scores and modulating microbial composition, thereby promoting recovery of laying hens after coccidia and Clostridium perfringens challenge.

Towards the mechanism of spermatotoxicity of p-tert-butyl-alpha-methylhydrocinnamic aldehyde: inhibition of late stage ex-vivo spermatogenesis in rat seminiferous tubule cultures by para-tert-butyl- benzoic acid.

Molecules metabolized to para-tert-butyl-benzoic acid (p-TBBA) affect male reproduction in rats through effects on spermatogenesis. This toxicity is specific to p-TBBA and not observed in meta-substituted analogues. The underlying mode of action was evaluated by comparing effects of p-TBBA and the position isomer m-TBBA (2-50 µM) in an ex vivo 3D primary seminiferous tubule cell culture system from juvenile Sprague Dawley rats (Bio-AlteR®). Treated cultures were evaluated for CoA-conjugate formation, cytotoxicity, blood-testis barrier functionality and different germ cell populations to assess effects on spermatogenesis. In addition, an evaluation of the metabolome of treated cultures was performed by using MxP® Broad Profiling via a LC-MS/MS and GC-MS platform. Para-TBBA decreased germ cell populations of late stages of spermatogenesis and led to the formation of CoA-conjugates in the ex vivo tissue. In addition, p-TBBA had a pronounced effect on the metabolome by affecting lipid balance and other CoA-dependent pathways contributing to energy production and the redox system. Meta-TBBA did not affect germ cell populations and no m-TBBA related CoA-conjugates were detectable. The metabolic profile of m-TBBA treated cells was comparable to vehicle control treated cultures, indicating that formation of CoA-conjugates, inhibition of spermatogenesis, and effects on the metabolome are mechanistically linked events. Thus, for this specific chemical group an adverse outcome pathway can be postulated, including the formation of benzoic acid metabolites, accumulation of CoA-conjugates to a certain threshold and CoA depletion, which affects the metabolic and lipid profile and leads to tissue specific effects with impaired functionalities such as spermatogenesis.

Development of Novel Pyridine-Thiazole Hybrid Molecules as Potential Anticancer Agents.

Novel pyridine-thiazole hybrid molecules were synthesized and subjected to physico-chemical characterization and screening of their cytotoxic action towards a panel of cell lines derived from different types of tumors (carcinomas of colon, breast, and lung, glioblastoma and leukemia), and normal human keratinocytes, for comparison. High antiproliferative activity of the 3-(2-fluorophenyl)-1-[4-methyl-2-(pyridin-2-ylamino)-thiazol-5-yl]-propenone 3 and 4-(2-{1-(2-fluorophenyl)-3-[4-methyl-2-(pyridin-2-ylamino)-thiazol-5-yl]-3-oxopropylsulfanyl}-acetylamino)-benzoic acid ethyl ester 4 was revealed. The IC50 of the compound 3 in HL-60 cells of the acute human promyelocytic leukemia was 0.57 µM, while in the pseudo-normal human cell lines, the IC50 of this compound was >50 µM, which suggests that the compounds 3 and 4 might be perspective anticancer agents. The detected selectivity of the derivatives 3 and 4 for cancer cell lines inspired us to study the mechanisms of their cytotoxic action. It was shown that preincubation of tumor cells with Fluzaparib (inhibitor of PARP1) reduced the cytotoxic activity of the derivatives 3 and 4 by more than twice. The ability of these compounds to affect DNA nativity and cause changes in nucleus morphology allows for the suggestion that the mechanism of action of the novel pyridine-thiazole derivatives might be related to inducing the genetic instability in tumor cells.

The effect of benzoic acid with or without a direct-fed microbial on the nutrient metabolism and gas emissions of growing pigs.

Twenty-four gilts (PIC 337 × 1050, PIC Genus, Hendersonville, TN) with an initial body weight (BW) of 33.09 ± 1.33 kg were used to investigate the effects of benzoic acid (BA) and a Bacillus-based direct-fed microbial (DFM) on the nutrient metabolism and manure gas emissions of growing pigs. Pigs were blocked by BW, placed into metabolism stalls, and randomly assigned to one of four dietary treatments: basal control (PC), low nitrogen (NC), PC plus 0.3% BA (PC+BA; VevoVitall, DSM Nutritional Products), and PC plus 0.3% BA and 0.025% DFM (PC+BA+DFM; PureGro, DSM Nutritional Products). Pigs were fed a common diet from day 0 to 14, and the experimental diets were fed in two phases (day 14 to 28 and day 28 to 53). The experiment consisted of four collection periods, with each period subdivided into two subperiods to collect samples for gas emissions and nutrient balance. Firstly, manure samples were collected for 72 h. Twice daily, urine and feces were weighed, and urine pH was measured. After each period, manure was subsampled and taken to the lab to measure gas emissions. Secondly, urine and feces were quantitatively collected for 96 h to allow for measurement of nutrient digestibility (ATTD) and retention. Data were analyzed as repeated measures in SAS 9.4 (SAS Inst., Cary, NC) with fixed effects of treatment, collection period, and block. Pig was the experimental unit, and results were considered significant at P ≤ 0.05 and a tendency at 0.05 < P ≤ 0.10. Pigs fed PC+BA had the greatest ADG compared to pigs fed NC (P = 0.016), with intermediate ADG for pigs fed PC or PC+BA+DFM (P ≥ 0.148). The ATTD of dry matter, gross energy, P, and N did not differ between treatments (P ≥ 0.093). However, the ATTD of Ca was reduced in pigs fed PC+BA+DFM compared to pigs fed PC+BA (P = 0.012). Pigs fed PC+BA or NC excreted less urinary N compared to PC and PC+BA+DFM (P ≤ 0.034), which contributed to greater nitrogen retention in PC+BA compared to PC (P = 0.016). Furthermore, decreased manure pH from pigs fed PC+BA or NC resulted in lower ammonia (NH3) emissions compared to pigs fed PC+BA+DFM or PC. There was no effect of dietary treatment on manure hydrogen sulfide, methane, or carbon dioxide emissions. In conclusion, supplementing 0.3% BA improved N retention and reduced manure pH and NH3 emissions, similar to feeding pigs low N, but improved the ADG of pigs when compared to feeding a low N diet.

Diet formulation as a strategy to improve economic and nutritional efficiency may be combined with nonnutritive feed additives, such as organic acids, specifically benzoic acid, and direct-fed microbials, to further improve nutrient utilization in pigs. Therefore, the objective of this trial was to investigate the effect of supplementing benzoic acid with or without a direct-fed microbial on the nutrient metabolism and emissions of ammonia, hydrogen sulfide, carbon dioxide, and methane from the manure of growing pigs. Feeding a diet containing 0.3% benzoic acid did not affect nutrient digestibility but reduced urinary nitrogen excretion, which resulted in improved nitrogen retention compared to the basal diet. Furthermore, benzoic acid reduced urine and manure pH, contributing to reduced manure ammonia emissions. However, supplementing the direct-fed microbial alongside benzoic acid attenuated these effects.

5-Nitro-2-(3-phenylpropylamino) Benzoic Acid Inhibits the Proliferation and Migration of Lens Epithelial Cells by Blocking CaMKII Signaling.

Posterior capsule opacification (PCO) is a post-surgery complication of cataract surgery, and lens epithelial cells (LECs) are involved in its development. A suppressive effect on LECs is exerted by the non specific chloride channel inhibitor 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) exerts. Herein, the growth and migration inhibitory effects of NPPB on LECs were assessed, and the mechanism underlying the effects were investigated by focusing on Ca2+/CaMKII signaling. LECs were treated with different concentrations of NPPB, and the changes in cell viability, cell-cycle distribution, anchorage-dependent growth, migration, Ca2+ level, and CaMKII expression were evaluated. NPPB inhibited LECs' proliferation and induced G1 cell-cycle arrest in the cells. Regarding LECs' mobility, NPPB suppressed the cells' anchorage-dependent growth ability and inhibited their migration. Changes in cell phenotypes were associated with an increased intracellular Ca2+ level and down-regulation of CaMKII. Together these results confirmed the inhibitory effect of NPPB on the proliferation and migration of LECs, and the effect was shown to be associated with the induced level of Ca2+ and the inhibition of CaMKII signaling transduction.

Larvicidal, antioxidant and biotoxicity assessment of (2-(((2-ethyl-2 methylhexyl)oxy)carbonyl)benzoic acid isolated from Bacillus pumilus against Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus.

Mosquitoes are a vector for many dreadful diseases known for their public health concern. The continued use of synthetic insecticides against vector control has led to serious environmental impacts, human health problems, and the development of insect resistance. Hence, alternative mosquito control methods are needed to protect the environment and human health. In the present study, the bioefficacy of (2-(((2-ethyl-2 methylhexyl)oxy)carbonyl) benzoic acid isolated from Bacillus pumilus were tested against Aedes aegypti, Culex quinquefasciatus and Anopheles stephensi. The isolated bioactive compound was characterized through thin layer chromatography (TLC), UV-visible spectroscopy (UV), Fourier-transform infrared spectroscopy, nuclear magnetic resonance spectroscopy, and gas chromatography-mass spectrometry analysis. The pure compound caused a high percent mortality rate in a dose-dependent manner, the obtained values were 96, 82, 69, 50 and 34%; 86, 72, 56, 43, and 44%; 100, 90, 83, 70 and 56% against Ae. aegypti, Cx. quinquefasciatus, and An. stephensi respectively. The effective lethal concentration values (LC50) were 13.65, 14.90 and 9.64 ppm against Ae. aegypti, Cx. quinquefasciatus, An. Stephensi, respectively. The effect of (2-(((2-ethyl-2 methylhexyl)oxy)carbonyl) benzoic acid significantly increased the superoxide dismutase, catalase, α, ß esterase and Glutathione-S-transferase level after 24 h of the treatment period. The comet assay confirmed that isolated compound causes DNA damage in all tested insects. Histopathological examinations of treated larvae showed shrunken body posture, damaged epithelial cells and microvillus as compared to control organisms. The biosafety of the isolated compound was assessed against G. affinis and did not produce mortality which confirmed that the activity of the isolated compound is species specific. The current study concludes that the critical success factors of new insecticidal agent development are based on the eco-compatibility and alternative tools for the pesticide producing industry.

Iridoid Glycosides Isolated from Bellardia trixago Identified as Inhibitors of Orobanche cumana Radicle Growth.

Orobanche cumana is an obligate holoparasitic plant with noxious effects in sunflower crops. Bellardia trixago is a facultative hemiparasitic plant that infects ruderal plants without noxious significance in agriculture and is known to produce a wide spectrum of bioactive metabolites. The objective of this study was to evaluate the allelopathic effects of B. trixago on the growth of O. cumana seedlings. Three different extracts using solvents of increasing polarity (n-hexane, dichloromethane and ethyl acetate) were prepared from the flowers, aerial green organs and roots of two populations, a white-flowered and a yellow-flowered population of B. trixago, both collected in southern Spain. Each extract was studied using allelopathic screenings on O. cumana which resulted in the identification of allelopathic activity of the ethyl acetate extracts against Orobanche radicles. Five iridoid glycosides were isolated together with benzoic acid from the ethyl acetate extract of aerial green organs by bio-guided purification. These compounds were identified as bartsioside, melampyroside, mussaenoside, gardoside methyl ester and aucubin. Among them, melampyroside was found to be the most abundant constituent in the extract (44.3% w/w), as well as the most phytotoxic iridoid on O. cumana radicle, showing a 72.6% inhibition of radicle growth. This activity of melampyroside was significantly high when compared with the inhibitory activity of benzoic acid (25.9%), a phenolic acid with known allelopathic activity against weeds. The ecotoxicological profile of melampyroside was evaluated using organisms representing different trophic levels of the aquatic and terrestrial ecosystems, namely producers (green freshwater algae Raphidocelis subcapitata and macrophyte Lepidium sativum), consumers (water flea Daphnia magna and nematode Caenorhabditis elegans) and decomposers (bacterium Aliivibrio fischeri). The ecotoxicity of melampyroside differed significantly depending on the test organism showing the highest toxicity to daphnia, nematodes and bacteria, and a lower toxicity to algae and macrophytes. The findings of the present study may provide useful information for the generation of green alternatives to synthetic herbicides for the control of O. cumana.

Exploring the potential of benzoic acid derived from the endophytic fungus strain Neurospora crassa SSN01 as a promising antimicrobial agent in wound healing.

Endophytic fungi are known to produce bioactive compounds with the potential to be used as promising drugs to treat a wide range of diseases. To the best of our knowledge, the use of bioactive metabolites derived from endophytic fungi, particularly against multidrug resistant (MDR) pathogens inhabiting burn wounds, has been emphasized for the first time. Therefore, the purpose of this study is to investigate the potential of purified benzoic acid (BA) derived from Neurospora crassa, an endophytic fungus isolated from Lycium shawii, as a promising and alternative safe antimicrobial candidate in wound healing. As a result, benzoic acid, a safe and nontoxic compound, may be a promising candidate for combating clinical MDR pathogens of burn wound infections. In this study, Neurospora crassa strain SSN01 (MW856826) was successfully identified for the first time as a new BA-producing endophytic fungus isolated from Lycium shawii. The concentration of BA in the ethyl acetate extract reached 244 mg/mL. Purified BA had a detrimental effect on the MDR strains tested, and the MDR Staphylococcus aureus strain SA-17 was clearly more susceptible to BA as compared to the other tested MDR bacterial and fungal strains. Toxicological studies on experimental animals were conducted to evaluate the toxicity of BA and a suitable dose regimen for future human use. Oral administration of BA at the highest concentration of 300 µg/kg body weight resulted in nontoxic signs and no mortality. In vivo histopathological examination revealed that BA, as a nontoxic and safe compound, could be a promising candidate for wound healing, combating MDR pathogens of burn wound infections.

Structure-Activity Relationships and Antileukemia Effects of the Tricyclic Benzoic Acid FTO Inhibitors.

The N6-methyladenosine (m6A) demethylase FTO is overexpressed in acute myeloid leukemia (AML) cells and promotes leukemogenesis. We previously developed tricyclic benzoic acid FB23 as a highly potent FTO inhibitor in vitro. However, it showed a moderate antiproliferative effect on AML cells. In this work, we performed a structure-activity relationship study of tricyclic benzoic acids as FTO inhibitors. The analog 13a exhibited excellent inhibitory effects on FTO similar to that of FB23 in vitro. In contrast to FB23, 13a exerted a strong antiproliferative effect on AML cells. Like FTO knock down, 13a upregulated ASB2 and RARA expression and increased the protein abundance while it downregulated MYC expression and decreased MYC protein abundance. These genes are key FTO targets in AML cells. Finally, 13a treatment improved the survival rate of MONOMAC6-transplanted NSG mice. Collectively, our data suggest that targeting FTO with tricyclic benzoic acid inhibitors may be a potential strategy for treating AML.

Differential effects of TRPM4 channel inhibitors on Guinea pig urinary bladder smooth muscle excitability and contractility: Novel 4-chloro-2-[2-(2-chloro-phenoxy)-acetylamino]-benzoic acid (CBA) versus classical 9-phenanthrol.

Non-selective cation channels in urinary bladder smooth muscle (UBSM) are thought to mediate increases in cellular excitability and contractility. For transient receptor potential melastatin type-4 (TRPM4) channels, the evidence primarily relies on the inhibitor 9-phenanthrol, which exhibits pharmacological limitations. Recently, 4-chloro-2-[2-(2-chloro-phenoxy)-acetylamino]-benzoic acid (CBA) has been discovered as a novel TRPM4 channel blocker. We examined how, in comparison to 9-phenanthrol, CBA affects the excitability of freshly isolated guinea pig UBSM cells and the contractility of UBSM strips. Additionally, non-selective TRPM4 channel inhibitor flufenamic acid (FFA) and potentiator BTP2 (also known as YM-58483) were studied in UBSM cells. Unlike robust inhibition for 9-phenanthrol already known, CBA (up to 100 µM) displayed either no or a very weak reduction (<20%) in spontaneous phasic, 20 mM KCl-induced, and electrical field stimulated contractions. For 300 µM CBA, reductions were higher except for an increase in the frequency of KCl-induced contractions. In UBSM cells, examined under amphotericin B-perforated patch-clamp, CBA (30 µM) did not affect the membrane potential (I = 0) or voltage step-induced whole-cell cation currents, sensitive to 9-phenanthrol. The currents were not inhibited by FFA (100 µM), increased by BTP2 (10 µM), nor enhanced under a strongly depolarizing holding voltage of -16 or + 6 mV (vs. -74 mV). None of the three compounds affected the cell capacitance, unlike 9-phenanthrol. In summary, the novel inhibitor CBA and nonselective FFA did not mimic the inhibitory properties of 9-phenanthrol on UBSM function. These results suggest that TRPM4 channels, although expressed in UBSM, play a distinct role rather than direct regulation of excitability and contractility.

Antidiabetic and neuroprotective effects of a novel repaglinide analog.

Repaglinide (RPG) is an oral insulin secretagogue used in the treatment of diabetes. In this study, a new RPG analog was synthesized. Its antidiabetic and neuroprotective effects on dorsal root ganglions (DRG) in streptozotocin (STZ)-induced diabetic rats were examined compared to RPG. To assess the effects of 2-methoxy-4-(2-((3-methyl-1-(2-(piperidin-1-yl)phenyl)butyl)amino)-2-oxoethoxy)benzoic acid (OXR), the impact of OXR on oxidative stress biomarkers, motor function, and the expression of the glutamate dehydrogenase 1 (GLUD1), SLC2A2/glucose transporter 2 (GLUT2), and glucokinase (GCK) genes in STZ-induced diabetic rats were assessed. DRGs were examined histologically using hemotoxylin and eosin staining. Molecular docking was used to investigate the interactions between OXR and the binding site of RPG, the ATP-sensitive potassium (KATP) channel. Following 5 weeks of treatment, OXR significantly increased the level of total antioxidant power, decreased reactive oxygen species, and lipid peroxidation in the DRGs of diabetic rats. OXR restored STZ-induced pathophysiological damages in DRG tissues. Administration of OXR improved motor function of rats with diabetic neuropathy. Administration of 0.5 mg/kg OXR reduced blood glucose while promoting insulin, mainly through upregulation of messenger RNA expression of GLUD1, GLUT2, and GCK in the pancreas. Molecular docking revealed a favorable binding mode of OXR to the KATP channel. In conclusion, OXR has neuroprotective effects in diabetic rats by lowering oxidative stress, lowering blood glucose, and stimulating insulin secretion. We report that 0.5 mg/kg OXR administration was the most effective concentration of the compound in this study. OXR may be a promising target for further research on neuroprotective antidiabetic molecules.