A Capsicum annuum L. seed extract exerts anti-neuroexcitotoxicity in HT22 hippocampal neurons.

The hippocampal memory deficit stands out as a primary symptom in neurodegenerative diseases, including Alzheimer's disease. While numerous therapeutic candidates have been proposed, they primarily serve to delay disease progression. Given the irreversible brain atrophy or injury associated with these conditions, current research efforts are concentrated on preventive medicine strategies. Herein, we investigated whether the extracts of Capsicum annuum L. seeds (CSE) and Capsicum annuum L. pulp (CPE) have preventive properties against glutamate-induced neuroexcitotoxicity (one of the main causes of Alzheimer's disease) in HT22 hippocampal neuronal cells. Pretreatment with CSE demonstrated significant anti-neuroexcitotoxic activity, whereas CPE did not exhibit such effects. Specifically, CSE pretreatment dose-dependently inhibited the elevation of excitotoxic elements (intracellular calcium influx and reactive oxygen species; ROS) and apoptotic elements (p53 and cleaved caspase-3). In addition, the glutamate-induced alterations of neuronal activity indicators (brain-derived neurotrophic factor; BDNF and cAMP response element-binding protein phosphorylation; CREB) were significantly attenuated by CSE treatment. We also found that luteolin is the main bioactive compound corresponding to the anti-neuroexcitotoxic effects of CSE. Our results strongly suggest that Capsicum annuum L. seeds (but not its pulp) could be candidates for neuro-protective resources especially under conditions of neuroexcitotoxicity. Its underlying mechanisms may involve the amelioration of ROS-mediated cell death and BDNF-related neuronal inactivity and luteolin would be an active compound.

Impact of induced mutation-derived genetic variability, genotype and varieties for quantitative and qualitative traits in Mentha species.

PURPOSE: The genus Mentha spp. is an aromatic herb from the family 'Lamiaceae'. It is extensively predominant in temperate and sub-temperate regions of the world. The essential oil of this species is enriched with broad aroma constituents extensively utilized in food, beverages, flavor, cosmetics, perfumery, and pharmaceutical enterprises. With the global menthol market size estimated to be worth USD 765 million in 2022, India (accompanied by China and Brazil) is the world's primary manufacturer, consumer, and exporter of Mentha oil. Despite prominent global demand, the crucial bottleneck in mint cultivation is the need for more superior commercial cultivars. Predominant vegetative propagation mode with difficulties in manual emasculation, differential blooming times, sterile/sub-sterile hybrids, and low seed viability are the primary containment in creating genetic variability by classical breeding approaches. Therefore, genetic complications encountered in conventional breeding have led the breeders to apply mutation breeding as an alternative crop improvement approach in Mentha spp. These attempts at mutation breeding have produced some distinctive mutants as genetic pools for plant breeding programs, and some novel mutant mint cultivars have been made available for commercial cultivation. CONCLUSIONS: The prime strategy in mutation-based breeding has proven an adept means of encouraging the expression of recessive genes and producing new genetic variations. The present review comprises a significant contribution of mutation breeding approaches in the development of mutant mint species and its effects on physiological variation, photosynthetic pigment, essential oil content and composition, phytochemical-mediated defense response, pathogen resistivity, and differential expression of genes related to terpenoid biogenesis. Development and diversification have led to the release of varieties, namely Todd's Mitcham, Murray Mitcham, Pranjal, Tushar, and Kukrail in M. piperita L., Mukta, and Pratik in M. cardiaca Baker, Neera in M. spicata L., Kiran in M. citrata Ehrh., and Rose mint in M. arvensis L. that have revolutionized and uplifted mint cultivation leading to economic gain by the farmers and entrepreneurs.

Ten weeks of Capsicum annuum L. extract supplementation did not change adipose tissue-derived hormones, appetite, body composition, and muscle strength when combined with resistance training in healthy untrained men: A clinical trial study.

Capsiate (CAP) is a nonpungent capsaicin analog (Capsicum annuum L. extract) that has been studied as a potential antiobesity agent. However, the interaction between chronic CAP supplementation and resistance training is not clear. The purpose of this study was to examine the changes in adipose tissue-derived hormones, body composition, appetite, and muscle strength after 10 weeks of resistance training, combined with chronic CAP supplementation in healthy untrained men. We hypothesized that CAP could induce higher benefits when combined with resistance training after 10 weeks of intervention compared to resistance training alone. Twenty-four young men (age, 22.0 ± 2.9) were randomized to either capsiate supplementation (CAP = 12 mg/day) or placebo (PL), and both groups were assigned to resistance training. Body composition, leptin and adiponectin concentrations, subjective ratings of appetite, energy intake, and exercise performance were assessed at before and after 10 weeks of progressive resistance training. There was a significant increase in body mass (P < .001), fat-free mass (CAP: 58.0 ± 7.1 vs. post, 59.7 ± 7.1 kg; PL: pre, 58.4 ± 7.3 vs. post, 59.8 ± 7.1 kg; P < .001), resting metabolic rate (CAP: pre, 1782.9 ± 160.6 vs. post, 1796.3 ± 162.0 kcal; PL: pre, 1733.0 ± 148.9 vs. post, 1750.5 ± 149.8 kcal; P < .001), maximal strength at 45 leg press (P < .001) and bench press (P < .001) in both groups, but no significant (P > .05) supplementation by training period interaction nor fat mass was observed. For subjective ratings of appetite, energy intake, leptin, and adiponectin, no significant effect of supplementation by training period interaction was observed (P > .05). In conclusion, 10 weeks of resistance training increased total body weight, muscle mass, and maximum strength in healthy untrained men; however, CAP supplementation (12 mg, 7 days per week) failed to change adipose tissue-derived hormones, appetite, body composition and muscle strength in this population. Registered under Brazilian Registry of Clinical Trials (RBR-8cz9kfq).

A contrast of emerging contaminants rac- and l-menthol toxicities to Microcystis aeruginosa through biochemical, physiological, and morphological investigations.

Fragrances rac- and l-menthol extracted from peppermint are widely used and considered as emerging contaminants recently, which are persistent in the environment. Menthol has always been considered as a safe chemical for humans, but its potential adverse ecological effects on aquatic organisms and the toxic mechanisms have not yet been fully understood. The present study aims to investigate the physiological response of Microcystis aeruginosa after exposure to the two menthol isomers, and to explore the toxic mechanisms and ecological risks of these two chemicals. Results showed that rac-menthol exhibited a hormesis effect on the cell growth, chlorophyll a and protein contents; while l-menthol showed an inhibition effect. Adenosine triphosphate (ATP) content increased significantly at day 3 and then decreased markedly at day 6 after exposure to the two chemicals. Compared with rac-menthol, l-menthol can cause damage to the antioxidant system and plasmalemma more severely, promote the production and release of microcystins-LR (MC-LR) more dramatically, upregulate the expression of MC-transportation-related gene mcyH, and induce higher apoptosis rates. Overall results revealed that the toxic effects of l-menthol on cyanobacteria were significantly greater than those of rac-menthol. The significant increase in the malondialdehyde (MDA) content and the ultrastructural characteristics of the cells indicated that the plasma membranes were damaged. Thus, further attention should be paid to the scientific use, ecological and environmental risk assessment of chiral menthol. This study will also provide a scientific basis for future water quality criteria establishment on emerging contaminants such as fragrances.

Menthol causes mitochondrial Ca2+-influx, affects structure-function relationship and cools mitochondria.

Menthol is a small bioactive compound able to cause several physiological changes and has multiple molecular targets. Therefore, cellular response against menthol is complex, and still poorly understood. In this work, we used a human osteosarcoma cell line (Saos-2) and analysed the effect of menthol, especially in terms of cellular, subcellular and molecular aspects. We demonstrate that menthol causes increased mitochondrial Ca2+ in a complex manner, which is mainly contributed by intracellular sources, including ER. Menthol also changes the Ca2+-load of individual mitochondrial particles in different conditions. Menthol increases ER-mito contact points, causes mitochondrial morphological changes, and increases mitochondrial ATP, cardiolipin, mitochondrial ROS and reduces mitochondrial membrane potential (ΔΨm). Menthol also prevents the mitochondrial quality damaged by sub-lethal and lethal doses of CCCP. In addition, menthol lowers the mitochondrial temperature within cell and also serves as a cooling agent for the isolated mitochondria in a cell free system too. Notably, menthol-induced reduction of mitochondrial temperature is observed in diverse types of cells, including neuronal, immune and cancer cells. As the higher mitochondrial temperature is a hallmark of several inflammatory, metabolic, disease and age-related disorders, we propose that menthol can serve as an active anti-aging compound against all these disorders. These findings may have relevance in case of several pharmacological and clinical applications of menthol. SIGNIFICANCE STATEMENT: Menthol is a plant-derived bioactive compound that is widely used for several physiological, behavioural, addictive, and medicinal purposes. It is a well-established "cooling and analgesic agent". However, the exact cellular and sub-cellular responses of menthol is poorly understood. In this work, we have characterized the effects of menthol on mitochondrial metabolism. Menthol regulates mitochondrial Ca2+, ATP, superoxides, cardiolipin, membrane-potential, and ER-mito contact sites. Moreover, the cooling agent menthol also cools down mitochondria and protects mitochondrial damage by certain toxins. These findings may promote use of menthol as a useful supplementary agent for anti-aging, anti-cancer, anti-inflammatory purposes where higher mitochondrial temperature is prevalent.

Effects of endogenous capsaicin stress and fermentation time on the microbial succession and flavor compounds of chili paste (a Chinese fermented chili pepper).

Chili paste, is a popular traditional product derived from chili pepper, and its fermentation system is affected by the variable concentration of capsaicin, which originates from the peppers. In the present study, the effects of capsaicin and fermentation time on the microbial community and flavor compounds of chili paste were investigated. After capsaicin supplementation, the total acid was significantly decreased (p < 0.05) along with lower total bacteria, especially lactic acid bacteria. Lactiplantibacillus, Lactobacillus, Weissella, Issatchenkia, Trichoderma, and Pichia were the shared and predominant genera; whereas, the Bacteroides and Kazachstania abundance was significantly increased due to the selection effect of capsaicin over time. Additionally, alterations of the microbial interaction networks and their metabolic preferences led to less lactic acid content with greater accumulation of ethyl nonanoate, methyl nonanoate, etc. This study will provide a perspective for selecting chili pepper varieties and improving the quality of fermented chili paste.

Transient Receptor Potential (TRP) based polypharmacological combination stimulates energy expending phenotype to reverse HFD-induced obesity in mice.

BACKGROUND & AIM: Obesity is a worldwide epidemic leading to decreased quality of life, higher medical expenses and significant morbidity. Enhancing energy expenditure and substrate utilization in adipose tissues through dietary constituents and polypharmacological approaches is gaining importance for the prevention and therapeutics of obesity. An important factor in this regard is Transient Receptor Potential (TRP) channel modulation and resultant activation of "brite" phenotype. Various dietary TRP channel agonists like capsaicin (TRPV1), cinnamaldehyde (TRPA1), and menthol (TRPM8) have shown anti-obesity effects, individually and in combination. We aimed to determine the therapeutic potential of such combination of sub-effective doses of these agents against diet-induced obesity, and explore the involved cellular processes. KEY FINDINGS: The combination of sub-effective doses of capsaicin, cinnamaldehyde and menthol induced "brite" phenotype in differentiating 3T3-L1 cells and subcutaneous white adipose tissue of HFD-fed obese mice. The intervention prevented adipose tissue hypertrophy and weight gain, enhanced the thermogenic potential, mitochondrial biogenesis and overall activation of brown adipose tissue. These changes observed in vitro as well as in vivo, were linked to increased phosphorylation of kinases, AMPK and ERK. In the liver, the combination treatment enhanced insulin sensitivity, improved gluconeogenic potential and lipolysis, prevented fatty acid accumulation and enhanced glucose utilization. SIGNIFICANCE: We report on the discovery of therapeutic potential of TRP-based dietary triagonist combination against HFD-induced abnormalities in metabolic tissues. Our findings indicate that a common central mechanism may affect multiple peripheral tissues. This study opens up avenues of development of therapeutic functional foods for obesity.

Convalescent action of menthol against T-2 mycotoxin-induced toxicity: An in vitro study with HaCaT cells.

Only T-2 mycotoxin is emitted as an aerosol and is the most toxic fungal secondary metabolite among mycotoxins. In its clinical condition, the skin is severely irritated and painful due to lesions and alimentary toxic aleukia. Herein, we have assessed various bioactive molecules, viz. kaempferol, menthol, curcumin, and quercetin, against T-2-induced toxicity in HaCaT cells. Menthol offered exceptional protection, protecting 92% of HaCaT cells after exposure to 300 nM T-2 and reducing LDH leakage by up to 42%. Its pre-treatment provided considerable protection against T-2 toxicity, as evidenced by the assessment of mitochondrial membrane potential. Propidium iodide staining revealed a cell cycle halt at the G1, S, and M phases and a significant increase in the sub-G1 percentage in T-2-challenged cells, indicating cell death. However, pre-treatment with menthol promoted cell cycle progression in cells exposed to T-2. Immunoblotting results demonstrated that menthol resulted in a discernible down-regulation of i-NOS expression in T-2-challenged HaCaT cells.

Beneficial Effects of Fermentation of Red Chili Pepper Using Lactococcus lactis subs. Cremoris RPG-HL-0136 in High-Fat Diet-Induced Obese Mice.

Red chili pepper is a beneficial natural spicy food that has antiobesity and antitype II diabetes effects, but it is not conducive to in-depth research as a dietary strategy to treat obesity. This study aims to investigate the beneficial effects of red chili pepper, fermented with a novel Lactococcus lactis subs. cremoris RPG-HL-0136. LC-MS/MS analysis is conducted to detect the content of capsaicin and dihydrocapsaicin, and no significant difference is observed between the nonfermented red chili pepper (NFP) (W/W) and the prepared L. lactis subs. cremoris RPG-HL-0136-fermented chili mixture (LFP). After establishing a high-fat diet-induced obese type II diabetic mouse model, the effects on weight gain, weight loss of liver and testicular fat, total cholesterol, triglyceride, fasting glucose, insulin, and homeostatic model assessment for insulin resistance in LFP were evaluated to be better than those in NFP following 10 weeks of interventions. All animal experiments were approved by the Institutional Animal Care and Use Committee of Xinxiang medical university. NFP and LFP could increase the expression of transient receptor potential vanilloid subfamily 1, peroxisome proliferator-activated receptor-alpha and caspase-2 in the high-fat mice. Compared with unfermented red chili pepper, the fermented red chili pepper complex significantly reduced LPS, tumor necrosis factor-alpha, and interleukin-6 in serum (P < .05). Intake of LFP significantly increased the expression of claudin-1 and occludin in the colon of the high-fat mice (P < .05), and there was no damage to the stomach and colon. This study provides scientific evidence that red chili pepper, fermented with L. lactis subs. cremoris RPG-HL-0136, may be beneficial for future treatment of obesity and accompanying diabetes. (IACUC.No.XYLL-20200019).

Promoting Activity of Terpenes on Skin Permeation of Famotidine.

Famotidine (FMT) is a competitive histamine-2 (H2) receptor antagonist that inhibits gastric acid secretion for the treatment of Gastroesophageal reflux disease. To study the promoting effect and mechanism of terpenes, including l-menthol, borneol, and geraniol, as chemical enhancers, FMT was used as a model drug. Attenuated total reflectance-Fourier transform IR spectroscopy (ATR-FTIR) and differential scanning calorimetry (DSC) were used to explore the effects of terpenes on the skin. Hairless mouse skin was mounted on Franz-type diffusion cell, and skin permeation experiment of FMT hydrogel was carried out. The results suggested that the thermodynamic activity influenced the permeability of the drug, and the main mechanism of terpenes to enhance skin permeation of the drug was based on increasing the fluidity of the intercellular lipids. Moreover, it was revealed that l-menthol simultaneously relaxed the packing structure and lamellar structure, whereas geraniol had a great influence on the lamellar structure only. Collectively, all terpenes had a promoting effect on skin permeation of FMT, indicating their potential as chemical enhancers to change the microstructure of stratum corneum and improve the permeation of FMT through the skin, and it has great potential to be used in transdermal formulations of FMT.

Glandular trichome specificity of menthol biosynthesis pathway gene promoters from Mentha × piperita.

MAIN CONCLUSION: Several cis-elements including Myb-binding motifs together confer glandular trichome specificity as revealed from heterologous expression and analysis of menthol biosynthesis pathway gene promoters. Glandular Trichomes (GTs) are result of division of epidermal cells that produce diverse metabolites. Species of mint family are important for their essential oil containing many high-value terpenoids, biosynthesized and stored in these GTs. Hence, GTs constitute attractive targets for metabolic engineering and GT-specific promoters are important. In this investigation, the upstream regions of the Mentha × piperita menthol biosynthetic pathway genes (-)-limonene synthase, (-)-P450 limonene-3- hydroxylase, (-)-trans-isopiperitenol dehydrogenase, (-)-Isopiperitenone reductase, ( +)-Pulegone reductase, (-)-Menthone reductase/ (-)-Menthol dehydrogenase and a branched pathway gene ( +)-menthofuran synthase were isolated and characterized. These fragments, fused to ß-glucuronidase (GUS) reporter gene of pBI101 binary vector, are able to drive high level gene expression in transgenic tobacco trichomes with strong signals in GTs, except for (-)-Isopiperitenone reductase. The GT-enriched tissue from transformed plants were analysed for GUS enzyme activity and RNA expression which correlates the GUS staining. To characterize the cis-elements responsible for GT-specific expression, a series of 5' deletion constructs for MpPLS and MpPMFS were cloned and analysed in stable transgenic tobacco lines. The specificity of trichome expression was located to -  797 to-  598 bp sequence for (-)-limonene synthase and-  629 to -   530 bp for ( +)-menthofuran synthase promoters containing specific Myb-binding motifs in addition to other unique motifs described for developmental regulation without any defined pattern. All other pathway promoters also recruits specific but different Myb factors as indicated by this analysis.

Phospholipid and menthol based nanovesicle impregnated transdermal patch for nutraceutical delivery to diminish folate and iron deficiency.

Adequate micronutrient availability is particularly important in women, children and infants. Micronutrient deficiencies are the major cause of maternal and neonatal morbidity. To overcome this, WHO recommends the use of folic acid and iron supplements for reducing anaemia and improving the health of the mother and infants. Oral intake of supplements for nutritional deficiencies are associated with gastric irritation, nausea, constipation and non-patient compliance due to associated taste. In case of absorption deficiency nutrients administered orally pass-through digestive tract unabsorbed. In the present study, we propose transdermal delivery of nutraceuticals to avoid the limitations associated with oral intake. Transdermal delivery has limited use because of the closely packed barrier of the stratum corneum that limits the permeability of molecules across skin. Here, we have used biomimetic nanovesicles impregnated in transdermal patches for delivery of folic acid and iron. Nanovesicles are prepared using an abundant component of cell membrane, phosphatidyl choline and a permeation enhancer. Further these nanovesicles are impregnated onto polyacrylate based transdermal patch.In vitrostudies have shown the ability of nanovesicles to fluidise skin lipids and penetrate into deeper skin.In vivoapplication of transdermal patches gradually increased the systemic concentration of nutraceuticals. Post application of the patch, five-fold increase in plasma folic acid concentration and 1.5-fold increase in plasma iron concertation was achieved in 6 h. Developed nanovesicles were compatible with keratinocytes and fibroblasts as testedin vitroand have the potential to enhance the cellular uptake of molecules. Skin irritation studies on human volunteers have confirmed the safety of nutraceutical loaded nanovesicles. Thus, the developed nutraceutical loaded transdermal patches provide a potential, easy to use platform for micronutrient delivery in infants and mothers.

Disruption impact of citronella and menthol insecticides on adults behavior and hemocytes morphology in the red palm weevil Rhynchophorus ferrugineus "Oliver" (Coleoptera: Curculionidae).

This study was conducted to evaluate some terpenes effect on the behavior and immune function of hemocytes in adults of the red palm weevil Rhynchophorus ferrugineus. Six individual different terpenes these are: (±)-menthol, B-citronellol, ( + )-3-carene, (R)- ( + )- limonene, citronella oil and orange terpenes. The results revealed significant differences between the terpenes used on the olfactory response on this insect, in that half of the compounds were very attractive while the other half were repellant to them. This behavior study results with olfactometer citronella oil exhibited an 80% attraction response rate for both sexes, while menthol exhibited a 60% attraction response rate for females and 100% for males. By contrast, menthol had a more significant effect on adults than citronella, lethal concentration at 50 scale (LC50) values of 1.03, 0.89, and 0.9 mg, and LC95 values of 5.09, 2.01, and 1.59 mg, after 24, 48 and 72 h, respectively. For citronella oil, the LC50 values were 2.09, 1.76, and 1.70 mg after 24, 48, and 72 h, and the LC95 values were 5.5, 3.7, and 1.5 mg after 24, 48 and 72h, were noted. In the present study, the effects of citronella and methanol insecticides were observed on six types of hemocytes namely prohemocytes, granulocytes, plasmatocytes, oenocytes, coagulocytes and spherulocytes. Both citronella oil and menthol had a histopathological effect on the hemocytes of the adult red palm weevil, specifically, on the cell membrane, cytoplasm, and nucleus. The findings also revealed that the vacuoles in some hemocytes, specifically, the prohemocytes, plasmatocytes, and granulocytes were more sensitive than those in other hemocytes, which remained unaffected by the treatment.The effects of citronella and menthol on RPW immunity were demonstrated in this study, and this information may be applied to their usage in integrated pest control at sub-lethal dosages.

Identification of Fungal Limonene-3-Hydroxylase for Biotechnological Menthol Production.

More than 30,000 tons of menthol are produced every year as a flavor and fragrance compound or as a medical component. So far, only extraction from plant material and chemical synthesis are possible. An alternative approach for menthol production could be a biotechnological-chemical process with ideally only two conversion steps, starting from (+)-limonene, which is a side product of the citrus processing industry. The first step requires a limonene-3-hydroxylase (L3H) activity that specifically catalyzes hydroxylation of limonene at carbon atom 3. Several protein engineering strategies have already attempted to create limonene-3-hydroxylases from bacterial cytochrome P450 monooxygenases (CYPs, or P450s), which can be efficiently expressed in bacterial hosts. However, their regiospecificity is rather low compared to that of the highly selective L3H enzymes from the biosynthetic pathway for menthol in Mentha species. The only naturally occurring limonene-3-hydroxylase activity identified in microorganisms so far was reported for a strain of the black yeast-like fungus Hormonema sp. in South Africa. We have discovered additional fungi that can catalyze the intended reaction and identified potential CYP-encoding genes within the genome sequence of one of the strains. Using heterologous gene expression and biotransformation experiments in yeasts, we were able to identify limonene-3-hydroxylases from Aureobasidium pullulans and Hormonema carpetanum Further characterization of the A. pullulans enzyme demonstrated its high stereospecificity and regioselectivity, its potential for limonene-based menthol production, and its additional ability to convert α- and ß-pinene to verbenol and pinocarveol, respectively.IMPORTANCE (-)-Menthol is an important flavor and fragrance compound and furthermore has medicinal uses. To realize a two-step synthesis starting from renewable (+)-limonene, a regioselective limonene-3-hydroxylase enzyme is necessary. We identified enzymes from two different fungi which catalyze this hydroxylation reaction and represent an important module for the development of a biotechnological process for (-)-menthol production from renewable (+)-limonene.

Menthol facilitates excitatory and inhibitory synaptic transmission in rat medullary dorsal horn neurons.

Menthol, which acts as an agonist for transient receptor potential melastatin 8 (TRPM8), has complex effects on nociceptive transmission, including pain relief and hyperalgesia. Here, we addressed the effects of menthol on spontaneous excitatory and inhibitory postsynaptic currents (sEPSCs and sIPSCs, respectively) in medullary dorsal horn neurons, using a whole-cell patch-clamp technique. Menthol significantly increased sEPSC frequency, in a concentration-dependent manner, without affecting current amplitudes. The menthol-induced increase in sEPSC frequency could be completely blocked by AMTB, a TRPM8 antagonist, but was not blocked by HC-030031, a transient receptor potential ankyrin 1 (TRPA1) antagonist. Menthol still increased sEPSC frequency in the presence of Cd2+, a general voltage-gated Ca2+ channel blocker, suggesting that voltage-gated Ca2+ channels are not involved in the menthol-induced increase in sEPSC frequency. However, menthol failed to increase sEPSC frequency in the absence of extracellular Ca2+, suggesting that TRPM8 on primary afferent terminals is Ca2+ permeable. On the other hand, menthol also increased sIPSC frequency, without affecting current amplitudes. The menthol-induced increase in sIPSC frequency could be completely blocked by either AMTB or CNQX, an AMPA/KA receptor antagonist, suggesting that the indirect increase in excitability of inhibitory interneurons may lead to the facilitation of spontaneous GABA and/or glycine release. The present results suggested that menthol exerts analgesic effects, via the enhancement of inhibitory synaptic transmission, through central feed-forward neural circuits within the medullary dorsal horn region.

The impact of menthol essential oil against inflammation, immunosuppression, and histopathological alterations induced by chlorpyrifos in Nile tilapia.

Chlorpyrifos (CPF) is one of the predominant water pollutants associated with inflammation and immunodepression in aquatic animals. In this study, menthol oil (MNT) impacted the immunity, antioxidative, and anti-inflammatory responses against CPF toxicity in Nile tilapia. Fish fed two diets with or without MNT and placed in four groups (control, CPF, MNT, and CPF/MNT). After 30 days, fish fed MNT displayed higher growth performance and lower FCR than CPF-intoxicated fish without feeding MNT (P < 0.05). The survival rate of fish was reduced in the CPF group without MNT feeding (P < 0.05). Blood Hb, PCV, RBCs, and WBCs were decreased in fish by CPF toxicity, while the highest Hb, PCV, RBCs, and WBCs were observed in fish fed MNT followed by those fed the control without CPF toxicity (P < 0.05). Fish fed MNT had the highest total protein, albumin, and globulin, as well as the lowest urea, bilirubin, and creatinine after 15 and 30 days. However, fish under CPF toxicity had the most inferior total protein, albumin, and globulin, as well as the highest urea, bilirubin, and creatinine among the groups (P < 0.05). The enzyme activities of ALP and ALT displayed low levels by MNT with or without CPF exposure than fish fed without MNT with or without CPF exposure after 15 and 30 days (P < 0.05). The lysozyme and phagocytic activities displayed reduced levels by CPF without MNT feeding after 15 and 30 days, while increased activities were noticed by MNT feeding without CPF toxicity followed by fish fed MNT with CPF toxicity (P < 0.05). The transcription of CAT and GPX genes displayed upregulated levels in tilapia fed MNT and exposed to CPF (P < 0.05). Also, CPF toxicity increased the transcription of the IFN-γ gene but decreased the IL-8 and IL-1ß genes. The transcription of HSP70 displayed lower levels (P < 0.05) by CPF without supplementing MNT than fish fed MNT and exposed to CPF. Histopathological analysis revealed that inflammation existed in the liver, gills, and intestine of tilapia due to CPF toxicity while MNT protected tissues from inflammation. To conclude, MNT activated the immunity, antioxidative, and anti-inflammatory responses of Nile tilapia under CPF toxicity.

Role of l- and d-Menthol in the Glucuronidation and Detoxification of the Major Lung Carcinogen, NNAL.

Menthol, which creates mint flavor and scent, is often added to tobacco in both menthol and nonmenthol cigarettes. A potent tobacco carcinogen, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), is extensively metabolized to its equally carcinogenic metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) as (R)- or (S)-NNAL enantiomers. NNAL is detoxified by UDP-glucuronosyltransferase (UGT) enzymes, with glucuronidation occurring on either NNAL's pyridine ring nitrogen (NNAL-N-Gluc) or the chiral alcohol [(R)- or (S)-NNAL-O-Gluc]. To characterize a potential effect by menthol on NNAL glucuronidation, in vitro menthol glucuronidation assays and menthol inhibition of NNAL-Gluc formation assays were performed. Additionally, NNAL and menthol glucuronides (MG) were measured in the urine of smokers (n = 100) from the Southern Community Cohort Study. UGTs 1A9, 1A10, 2A1, 2A2, 2A3, 2B4, 2B7, and 2B17 all exhibited glucuronidating activity against both l- and d-menthol. In human liver microsomes, both l- and d-menthol inhibited the formation of each NNAL-Gluc, with a stereospecific difference observed between the formation of (R)-NNAL-O-Gluc and (S)-NNAL-O-Gluc in the presence of d-menthol but not l-menthol. With the exception of three nonmenthol cigarette smokers, urinary MG was detected in all menthol and nonmenthol smokers, with l-MG comprising >98% of total urinary MG. Levels of urinary NNAL-N-Gluc were significantly (P < 0.05) lower among subjects with high levels of total urinary MG; no significant changes in free NNAL were observed. These data suggest that the presence of menthol could lead to increases in alternative, activating metabolic pathways of NNAL in tobacco target tissues, increasing the opportunity for NNAL to damage DNA and lead to the development of tobacco-related cancers. SIGNIFICANCE STATEMENT: High levels of the major menthol metabolite, menthol-glucuronide, was observed in the urine of smokers of either menthol or nonmenthol cigarettes. The fact that a significant inverse correlation was observed between the levels of urinary menthol-glucuronide and NNAL-N-glucuronide, a major detoxification metabolite of the tobacco carcinogen, NNK, suggests that menthol may inhibit clearance of this important tobacco carcinogen.

Drosophila menthol sensitivity and the Precambrian origins of transient receptor potential-dependent chemosensation.

Transient receptor potential (TRP) cation channels are highly conserved, polymodal sensors which respond to a wide variety of stimuli. Perhaps most notably, TRP channels serve critical functions in nociception and pain. A growing body of evidence suggests that transient receptor potential melastatin (TRPM) and transient receptor potential ankyrin (TRPA) thermal and electrophile sensitivities predate the protostome-deuterostome split (greater than 550 Ma). However, TRPM and TRPA channels are also thought to detect modified terpenes (e.g. menthol). Although terpenoids like menthol are thought to be aversive and/or harmful to insects, mechanistic sensitivity studies have been largely restricted to chordates. Furthermore, it is unknown if TRP-menthol sensing is as ancient as thermal and/or electrophile sensitivity. Combining genetic, optical, electrophysiological, behavioural and phylogenetic approaches, we tested the hypothesis that insect TRP channels play a conserved role in menthol sensing. We found that topical application of menthol to Drosophila melanogaster larvae elicits a Trpm- and TrpA1-dependent nocifensive rolling behaviour, which requires activation of Class IV nociceptor neurons. Further, in characterizing the evolution of TRP channels, we put forth the hypotheses that three previously undescribed TRPM channel clades (basal, αTRPM and ßTRPM), as well as TRPs with residues critical for menthol sensing, were present in ancestral bilaterians. This article is part of the Theo Murphy meeting issue 'Evolution of mechanisms and behaviour important for pain'.

Omeprazole alleviates water stress in peppermint and modulates the expression of menthol biosynthesis genes.

Water stress is a worldwide agricultural challenge that limits crop growth and quality. Chemical compounds that promote tolerance to water stress, such as omeprazole showed recently promising results. The present study investigates the effect of weekly drenching applications of 0, 10, 50, 100, or 200 µM omeprazole on Mentha piperita (peppermint) subjected to water stress by watering at 100%, 70%, and 50% of container substrate capacity for 7 weeks in an experiment that spanned two seasons. Peppermint that received higher doses of omeprazole showed increased plant height, leaf number, leaf area, and dry weight under normal and water stress conditions. The amounts of chlorophyll and proline in the leaves as well as gas exchange increased in omeprazole-treated plants relative to the control plants. Omeprazole treatment also resulted in increased activity of the enzymes catalase and ascorbate peroxidase, reduced accumulation of the reactive oxygen species hydrogen peroxide, increase in the essential oil ratio, and improvement in essential oil composition. Omeprazole-treated plants showed higher ratios of menthol and menthone composition relative to the control plants. The changes in essential oil composition were associated with increased expression of genes associated with the menthol biosynthesis pathway. These findings indicate that omeprazole can ameliorate water stress in peppermint by increasing vegetative and root growth; increasing chlorophyll amount, photosynthetic rate, and gas exchange; reducing water loss by boosting leaf water potential and relative water content; increasing proline content; and modulating the gene expression of secondary metabolites.

Evolution of the human cold/menthol receptor, TRPM8.

Genes showing versatile functions or subjected to fast expansion and contraction during the adaptation of species to specific ecological conditions, like sensory receptors for odors, pheromones and tastes, are characterized by a great plasticity through evolution. One of the most fascinating sensory receptors in the family of TRP channels, the cold and menthol receptor TRPM8, has received significant attention in the literature. Recent studies have reported the existence of TRPM8 channel isoforms encoded by alternative mRNAs transcribed from alternative promoters and processed by alternative splicing. Since the first draft of the human genome was accomplished in 2000, alternative transcription, alternative splicing and alternative translation have appeared as major sources of gene product diversity and are thought to participate in the generation of complexity in higher organisms. In this study, we investigate whether alternative transcription has been a driving force in the evolution of the human forms of the cold receptor TRPM8. We identified 33 TRPM8 alternative mRNAs (24 new sequences) and their associated protein isoforms in human tissues. Using comparative genomics, we described the evolution of the human TRPM8 sequences in eight ancestors since the origin of Amniota, and estimated in which ancestors the new TRPM8 variants originated. In order to validate the estimated origins of this receptor, we performed experimental validations of predicted exons in mouse tissues. Our results suggest a first diversification event of the cold receptor in the Boreoeutheria ancestor, and a subsequent divergence at the origin of Simiiformes.