Analysis of bioactive components in Ghost chili (Capsicum chinense) for antioxidant, genotoxic, and apoptotic effects in mice.

Apart from using traditionally in culinary preparations, chili peppers are also important constituents of herbal medicines. Although the bioactive components are studied mostly in the fruits of Capsicum annuum, no such study reports till date is available for Ghost chili (C. chinense) from North East India. Therefore, the aim of this study was to carry out an analysis of the bioactive constituents in the naturally occurring hottest chili Ghost chili (C. chinense), and evaluate its antioxidant, pro/anti-genotoxic, and apoptotic effects in in vitro and in vivo models. Three different antioxidant assays showed that lower doses of Ghost chili extract showed higher DNA protective and antioxidant activities. Furthermore, the administration for 7 alternate days into 6 week old Swiss albino mice showed that the lower doses (50 and 100 mg/kg bw) reduced DMBA induced genotoxicity beside significantly enhancing the activities of hepatic antioxidant enzymes, while higher dose (200 mg/kg bw) induced genotoxic effect in bone marrow cells. The administration of higher dose (200 mg/kg bw) also induced apoptosis and upregulation of Bax (pro) and downregulation of Bcl-2 (anti) apoptotic genes. Dose dependent increase of apoptosis was also observed in Hep G2 and Hep 3B liver cancer cell lines. Our findings in the present study suggest that low doses of C. chinense can exert cancer chemopreventive effects. The induction of apoptosis in both cancer cell lines and mouse bone marrow cells, and up-regulation of proapoptotic genes suggests that the higher dose of C. chinense can be used for targeted cancer therapy.

Coffee mitigates cyclophosphamide-induced genotoxic damage in Drosophila melanogaster germ cells.

In the present study, coffee (CF) was evaluated for its protective effects against genotoxic damage and oxidative stress induced by the chemotherapeutic drug, cyclophosphamide (CPH). The sex-linked recessive lethal (SLRL) test was employed to study the induction of mutations in the larvae as well as in all the successive germ cell stages of treated males. Control and treated third instar larvae were used to monitor the biomarkers of oxidative stress response such as glutathione content (GSH), glutathione-S-transferase (GST), catalase (CAT), superoxide dismutase (SOD) and lipid peroxidation (MDA content). Our results demonstrated that co-administration of CF (2%) with CPH (3 mM) has significantly reduced CPH-induced lethal mutations in the germ cells of larvae and adult flies. The reductions observed in mutation frequencies were: 75% in larvae and 62.4% in the adult. Significant enhancement in antioxidant enzymatic levels: CAT (46.6%) > SOD (43.0%) > GST (42.4%) > GSH (31.6%) and reduction in MDA levels (32.05%) in the pretreated third instar larvae demonstrated the antioxidant activity of CF against CPH-induced oxidative stress. The findings from the present study suggest that the Drosophila model is an ideal one for evaluating the antigenotoxic and antioxidant activity of complex mixtures like CF.

Intake of saffron reduces γ-radiation-induced genotoxicity and oxidative stress in mice.

Saffron (SAF), the dried stigmas of Crocus sativus, is commonly used for flavoring and coloring food. Studies on bioactivity of SAF have demonstrated its in vivo antioxidant activity. The aim of our study was to assess the impact of SAF intake on γ-radiation (RAD) induced (a) chromosomal damage, (b) oxidative stress in liver and brain, and (c) histopathological effects in the intestinal cells and male germ cells in mice. Freeze-dried aqueous extract of SAF was used for the experiments. Our preliminary cell-free DNA nicking assay using pBR322 DNA revealed protective effects of freeze-dried SAF extract against hydroxyl radical induced DNA damage. For the in vivo investigations, freeze-dried SAF extract in distilled water was administered by gavage (40 mg/kg b.w.) to male Swiss albino mice for six consecutive days. On the sixth day, the animals were exposed to RAD (1 or 2 Gy) and sacrificed 24 h later to collect bone marrow cells for assessing chromosomal damage by measuring micronucleated polychromatic erythrocytes (MnPCEs). Liver and brain samples from animals exposed to 2 Gy RAD were used for evaluating lipid peroxidation and activity of antioxidant enzymes. The testis and intestine were used for histopathological analysis. Our results demonstrated significant protective effects of SAF against RAD-induced genotoxic damage. SAF pretreatment reduced the level of lipid peroxidation with concomitant increase in glutathione content and activity of glutathione S-transferase, glutathione peroxidase, and catalase. The histopathological analysis showed minimal impact of SAF on RAD-induced damage in the intestinal cells and male germ cells.

Nano-Pelargonidin Protects Hyperglycemic-Induced L6 Cells against Mitochondrial Dysfunction.

Nano-encapsulation of several natural products has become an important tool in enhancing the bioavailability of some modern drugs against many diseases. Pelargonidin is an anthocyanidin found in many fruits and vegetables. Pelargonidin is loaded with poly-lactide-co-glycolic-acid, a non-toxic biodegradable polymer, to produce nano-pelargonidin. Size, morphology, zeta potential, and planar uniformity of formulated nano-pelargonidin were determined by atomic force microscopy and dynamic light scattering. The time required for cellular entry, folds of nano-pelargonidin, and drug encapsulation efficiency of poly-lactide-co-glycolic-acid were also ascertained. Relative functional efficacy of nano-pelargonidin and pelargonidin was evaluated by examining markers such as pyruvate kinase, glucokinase, calcium ion level, ATP/ADP ratio, mitochondrial membrane potential, cytosolic release of mitochondrial cytochrome-c, and structural analysis of mitochondrial DNA in controlled and experimental sets of alloxan-induced hyperglycemic L6 cells. Expressions of mitochondrial apoptotic proteins, such as bcl2 and caspase3, and glucose signalling cascades, such as GLUT4, IRS1, IRS2, and PI3, were analyzed. Nano-pelargonidin at a nearly 10-fold reduced dose significantly enhanced protection, presumably due to its smaller size, ability of faster entry, and drug delivery at target-specific sites. Thus, nano-pelargonidin can be used in formulating protective drugs for therapeutic management of mitochondrial dysfunction often encountered in diabetic conditions.

Impact of Anthocyanidins on Mitoxantrone-Induced Cytotoxicity and Genotoxicity: An In Vitro and In Vivo Analysis.

Hypothesis Anthocyanins possess well-known biological effects and suppress DNA damage induced by therapeutic topoisomerase poisons. Our study focusses on the modulatory effects of anthocyanidins-malvidin (MAL) and pelargonidin (PEL)-on topoisomerase II poison mitoxantrone (MXT)-induced cytotoxicity and genotoxicity in in vitro and in vivo conditions. Study design HepG2 cells were treated with MXT (1-10 µM), MAL (10-100 µM,) and PEL (5-640 µM) to determine cell viability. Further, experiments on cytotoxicity and apoptosis induction by single agents or combinations were performed. In vitro and in vivo antigenotoxic effect of MAL/PEL against MXT was evaluated in human lymphocytes and mouse bone marrow cells. Methods Cytotoxicity of test agents and apoptosis induction in HepG2 cells was assessed by MTT assay, trypan blue dye exclusion assay and Hoechst 33258 staining. Antigenotoxic effects of MAL/PEL against MXT were assessed in co-treated human lymphocytes and bone marrow from mice that received MXT intraperitoneally 30 minutes post MAL/PEL oral administration Results Dose-dependent cytotoxicity was observed with all 3 test agents in HepG2 cells. Highest test concentration of 100 µM MAL, 640 µM PEL, and 10 µM MXT decreased HepG2 cell viability by 80%, 30%, and 90%, respectively. The combination of 1 µM MXT + 80 µM MAL reduced cell viability better than single agents. MAL/PEL treatment significantly reduced MXT-induced genotoxicity in human lymphocytes and micronuclei formation in mice. Conclusion Combination of MAL/PEL with lower doses of MXT, especially MAL+MXT increases the cytotoxicity in cancer cells. In addition, MXT treatment with MAL/PEL reduced MXT-induced genotoxicity and protected normal cells during chemotherapy.

Protective effects of common anthocyanidins against genotoxic damage induced by chemotherapeutic drugs in mice.

Experiments were performed to assess in mice the inhibitory effects of the anthocyanidins cyanidin, delphinidin, malvidin, and pelargonidin on genotoxic damage induced by the anticancer drugs cyclophosphamide, procarbazine, and cisplatin. Each anthocyanidin was administered 30 min before injecting the drug, and genotoxicity was assessed by measuring micronucleated polychromatic erythrocytes in bone marrow cells. In addition, we monitored the effect of anthocyanidins on apoptosis induced by cyclophosphamide and procarbazine. The results showed significant protective effects of cyanidin, delphinidin, malvidin, and pelargonidin against DNA damage induced by cyclophosphamide. With delphinidin and malvidin, a biphasic dose-response was observed for protection against cyclophosphamide. Dose-related reduction of genotoxicity was observed with pelargonidin against procarbazine. However with cyanidin, the medium dose of 2 mg/kg showed maximum protection against procarbazine. Cyanidin and pelargonidin significantly reduced the chromosomal damage induced by cisplatin. Furthermore, pre-treatment with these anthocyanidins reduced the level of apoptosis induced by cyclophosphamide and procarbazine. In conclusion, this study shows that anthocyanidins can reduce the efficacy of anticancer drugs for inducing DNA damage and apoptosis.

Ascorbic acid and dietary polyphenol combinations protect against genotoxic damage induced in mice by endogenous nitrosation.

We investigated whether combinations of ascorbic acid (AA) plus dietary polyphenols can protect in vivo against genotoxic damage induced by endogenous nitrosation. A nitrosation reaction mixture consisting of methylurea (MU) plus sodium nitrite (SN), which can react to form N-nitroso-N-methylurea in the stomach, was administered orally to mice, together with AA and one of the dietary polyphenols ferulic acid (FA), gallic acid (GA), chlorogenic acid (CA), or epigallocatechin gallate (EGCG). Genotoxic damage in bone marrow cells was assessed by measuring micronucleated polychromatic erythrocytes (Mn PCEs) and metaphase chromosome aberrations. When compared to damage induced by MU plus SN alone, co-administration with AA, FA, GA, CA, or EGCG resulted in significant protective effects. Combinations of AA plus EGCG or AA plus CA showed a further protective effect. Reduction in the frequency of Mn PCEs to the control level was obtained following co-administration of a combination of AA, FA, GA, and CA with MU plus SN. A similar trend was observed for metaphase chromosome aberrations. Co-administration of AA, FA, GA, or CA with N-nitroso-N-methylurea (MNU) did not show any significant reduction in genotoxicity, indicating the absence of a protective effect against a preformed N-nitroso compound. Our work demonstrates the protective effects of the 'antinitrosating' combination of AA and dietary polyphenols FA, GA, or CA against genotoxic damage induced by an endogenously formed N-nitroso compound.

In vitro and in vivo evaluation of antioxidant and antigenotoxic potential of Punica granatum leaf extract.

CONTEXT: Several studies have reported the antioxidant activity and potential therapeutic properties of Punica granatum L. (Lythraceae) fruit. Medicinal properties have also been attributed to other parts of P. granatum tree, which are rich in bioactive phytochemicals. OBJECTIVE: To explore the phytochemical characteristics, in vitro and in vivo antioxidant and in vivo antigenotoxic potential of P. granatum leaf extract (PLE). MATERIALS AND METHODS: The in vitro antioxidant potential of PLE was assessed by DPPH (1,1-diphenyl-2-picrylhydrazyl), ferric reducing antioxidant power (FRAP). Inhibition of lipid peroxidation (LPO) and the total phenolic content of the samples were also determined. Thirty-six male Swiss albino mice were divided into six groups (six animals each). Group 1 (control) and group 2 mice received vehicle and genotoxin alone, respectively. Groups 3, 4 and 5 were pretreated with PLE (400, 600 and 800 mg/kg body weight, respectively) prior to the administration of genotoxin. Group 6 received highest test dose of PLE. DNA damage in the bone marrow cells, hepatic LPO and antioxidants were recorded. RESULTS: Phytochemical analysis of PLE showed the presence of flavonoids, phenols, phytosterols, tannins and carbohydrates. Aqueous PLE demonstrated free radical scavenging activity, reducing power and inhibition of LPO with the EC50 values of 10.25, 59.88 and 20.05, respectively. A significant protective effect was observed against cyclophosphamide induced DNA damage and inhibition of hepatic LPO with concomitant increase in reduced glutathione (GSH) glutathione S-transferase (GST), superoxide dismutase (SOD) and catalase (CAT) in mice pretreated with PLE. DISCUSSION AND CONCLUSION: PLE demonstrated a significant antioxidant and antigenotoxic potential and hence can be a potential natural source in health and medicine.

Analysis of in vitro chemoprevention of genotoxic damage by phytochemicals, as single agents or as combinations.

Cancer chemoprevention with low-dose combinations of bioactive phytochemicals instead of single agents has been suggested to induce less toxicity and improve efficacy. In this study, we selected four plant food-based phytochemicals, viz. chlorogenic acid (CLA), pelargonidin (PEL), resveratrol (RES) and epigallocatechin gallate (EGCG) to evaluate the in vitro chemoprevention of genotoxic damage in HL-60 cells. These agents were tested either individually or as a combination at two concentrations (with a 10-fold difference) against the genotoxins mitomycin C (MMC), diepoxybutane (DEB) and patulin (PAT). Our preliminary ferric reducing antioxidant power (FRAP) assay demonstrated additive effects when PEL, CLA, RES and EGCG were combined. Results of the cytokinesis-block micronucleus test showed significant protection against genotoxic damage induced by PAT, DEB and MMC when CLA, PEL, RES and EGCG were tested individually. This protective effect of the phytochemicals was not concentration-related. Both low- and high-concentration combinations of CLA, PEL, RES and EGCG showed significant reducing effects on the frequencies of micronuclei induced by PAT, DEB and MMC. However, the micronucleus test did not provide indications of additive or synergistic effects with this combination of phytochemicals. In conclusion, the chemo-preventive effects of PEL, CLA, RES and EGCG against genotoxic damage induced by MMC, DEB and PAT are indicative of a 'saturation effect' when higher concentrations and combinations of these phytochemicals are used.

Role of Syzygium cumini seed extract in the chemoprevention of in vivo genomic damage and oxidative stress.

ETHNOPHARMACOLOGICAL RELEVANCE: [corrected] The seeds of Syzygium cumini, Skeels (Jamun) are extensively used in India for treatment of diabetes and other ailments. AIM OF THE STUDY: The aim of this work was to assess the role of Jamun seed extract (JSE) as a chemoprotective agent against in vivo oxidative stress and genomic damage. MATERIALS AND METHODS: Experiments were carried out to evaluate in vitro protective effects of JSE against hydroxyl radical induced damage in pBR322 DNA, and in vivo genomic damage and oxidative stress in mice which received JSE orally for 5 days before exposure to genotoxic carcinogens urethane (URE) and 7,12-dimethyl benz(a)anthracene (DMBA). RESULTS: Aqueous and ethanolic extracts of JSE showed significant protective effects against hydroxyl radical induced strand breaks in pBR322 DNA. The in vivo experiments with aqueous JSE showed significant protective effects against chromosomal damage induced by the genotoxic carcinogens URE and DMBA. Biochemical assays registered significant inhibition of hepatic lipid peroxidation and increase in GSH level and activity of GST, SOD and CAT. CONCLUSION: Our findings suggest that JSE can possibly play an important role as a chemopreventive agent against oxidative stress and genomic damage.

Protective effect of the cruciferous vegetable mustard leaf (Brassica campestris) against in vivo chromosomal damage and oxidative stress induced by gamma-radiation and genotoxic chemicals.

We evaluated the possible protective effect of the popular Indian cruciferous vegetable mustard leaf (Brassica campestris) against chromosomal damage and oxidative stress induced by gamma-radiation, cyclophosphamide (CPH) and urethane (URE), in mice. In vivo bone marrow micronucleus test was performed to assess chromosomal damage, and oxidative stress was monitored by estimating the changes in lipid peroxidation and the status of glutathione (GSH) as well as redox cycle antioxidants. Pretreatment with 50-250 mg/kg body wt of mustard leaf extract (MLE) for seven days significantly reduced the frequencies of micronuclei induced by gamma-radiation, CPH and URE. The protective effect against chromosomal damage was associated with modulation of lipid peroxidation as well as an increase in GSH and the GSH-dependent enzyme glutathione S-transferase (GST). Mass spectral analysis showed the presence of glucosinolates in MLE used for the pretreatment of mice. These findings indicate that intake of the green leafy cruciferous vegetable mustard leaf can lead to protection against in vivo genotoxicity and oxidative stress.

Antigenotoxic effects of the phytoestrogen pelargonidin chloride and the polyphenol chlorogenic acid.

Pelargonidin (PEL), a common anthocyanidin with estrogenic activity, was tested in HL-60 cells for its genotoxicity and possible antigenotoxic effects against 4-nitroquinoline 1-oxide (NQO), a potent mutagen and carcinogen which induces oxidative stress. To take into account potential interactions between phytochemicals within normal human nutrition, we evaluated a combination of PEL with the nonestrogenic phytochemical chlorogenic acid (CLA), one of the most abundant polyphenols in the human diet. PEL (< or = 2 microM) and CLA (< or = 800 microM) were nongenotoxic in the micronucleus test. We observed significant antigenotoxic effects against NQO with both compounds, but no additive interaction of PEL and CLA. Comet assay results showed a nonsignificant reduction in NQO-induced DNA damage with both compounds and their combination. Flow cytometric analysis of oxidative stress revealed significant protection against NQO-induced oxidative stress by PEL, CLA, and their combination. Furthermore, PEL and CLA prevented the NQO-induced reduction in GSH level. This could be a mechanism for the observed reduction in genotoxicity. In conclusion, the phytoestrogen PEL revealed antioxidative and antigenotoxic properties in HL-60 cells, but no significant additive interaction with the abundant nutritional polyphenol CLA under the tested conditions.

Combinatorial antigenotoxic and anticarcinogenic effects of tomato and garlic through modulation of xenobiotic-metabolizing enzymes during hamster buccal pouch carcinogenesis.

OBJECTIVE: Combination chemoprevention by dietary agents is a promising approach toward cancer control. Many dietary agents are known to prevent experimental mutagenesis and carcinogenesis by modulating xenobiotic-metabolizing enzymes. The present study evaluated the combinatorial chemopreventive effects of tomato and garlic on hamster buccal pouch carcinogenesis induced by 7,12-dimethylbenz[a]anthracene (DMBA). METHODS: Hamsters were assigned to one of four groups. The right buccal pouches of animals in group 1 were painted with 0.5% DMBA three times a week. The right buccal pouches of animals in group 2 were painted with DMBA and received intragastric administration of a combined dose of tomato and garlic on days alternate to DMBA application. Animals in group 3 were given chemopreventive agents alone. Animals in group 4 served as controls. Levels of phase I and II enzymes and the frequency of bone marrow micronuclei were used as biomarkers of chemoprevention. RESULTS: All the hamsters painted with DMBA alone developed buccal pouch carcinomas that exhibited increased activities of xenobiotic-metabolizing enzymes associated with increased frequencies of bone marrow micronuclei. In the liver, an increase in phase I enzymes was accompanied by compromised phase II detoxification activities. Combined administration of tomato and garlic effectively suppressed the incidence and mean tumor burden of hamster buccal pouch carcinomas. In addition, tomato and garlic combination significantly decreased phase I enzymes and increased phase II enzyme activities in the pouch and liver with a decreased incidence of bone marrow micronuclei. CONCLUSION: From these results, we suggest that modulation of xenobiotic-metabolizing enzymes exerted by tomato and garlic combination plays a key role in mitigating the mutagenic and carcinogenic effects of DMBA.

Protective effects of ethanolic neem leaf extract on DMBA-induced genotoxicity and oxidative stress in mice.

We evaluated the effects of pretreatment with ethanolic neem leaf extract on 7,12-dimethylbenz[a]anthracene (DMBA)-induced genotoxicity and oxidative stress in male Swiss albino mice. The frequency of bone marrow micronuclei, the extent of hepatic lipid peroxidation and the status of antioxidants-reduced glutathione (GSH), glutathione peroxidase (GPx) and glutathione-S-transferase (GST) were used as intermediate biomarkers of chemoprotection. In DMBA-treated mice, the increases in micronuclei and lipid peroxides were accompanied by compromised antioxidant defenses. Pretreatment with ethanolic neem leaf extract (200 mg/kg body weight) significantly reduced DMBA-induced micronuclei and lipid peroxides and enhanced GSH-dependent antioxidant activities. The results of the present study suggest that ethanolic neem leaf extract exerts protective effects against DMBA-induced genotoxicity and oxidative stress by enhancing the antioxidant status.

Protective effect of tomato against N-methyl-N'-nitro-N-nitrosoguanidine-induced in vivo clastogenicity and oxidative stress.

OBJECTIVE: Chemoprevention by dietary constituents has emerged as a cost-effective approach to control the incidence of gastric cancer, the second most common malignancy worldwide, and a major cause of mortality in Chennai, India. Identification of dietary agents with chemopreventive potential requires prescreening in animal models. We evaluated the modifying effects of tomato paste against N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced genetic damage and oxidative stress in male Swiss mice. METHODS: Quantitation of bone marrow micronuclei, extent of lipid peroxidation, and the status of the antioxidants reduced glutathione, glutathione peroxidase, and glutathione S-transferase in the stomach and liver of Swiss albino mice were used to monitor the protective effects of tomato. Three different doses of tomato (0.5, 1, and 2 g/kg of body weight) were administered intragastrically for 5 d consecutively followed by intragastric intubation of MNNG 1.5 h after the final administration. RESULTS: Increased frequency of micronuclei and enhanced lipid peroxidation were accompanied by compromised antioxidant defenses in MNNG-treated rats. Pretreatment of tomato paste at the concentration of 1 and 2 g/kg of body weight significantly decreased micronuclei and lipid peroxidation and enhanced reduced glutathione-dependent antioxidant and detoxifying enzymes. CONCLUSION: Our results demonstrate that tomato protects against the clastogenic effects of MNNG by decreasing micronucleated polychromatic erythrocytes and lipid peroxidation and enhancing antioxidant status.

Inhibitory effects of aqueous crude extract of Saffron (Crocus sativus L.) on chemical-induced genotoxicity in mice.

Saffron (dried stigmas of Crocus sativus L.), was evaluated in the mouse bone marrow micronucleus test for its possible protective effects against chromosomal damage induced by cisplatin (CIS), mitomycin-C (MMC) and urethane (URE). Three doses of saffron (25, 50 and 100 mg/kg body weight) were orally administered to mice for five consecutive days prior to administration of genotoxins under investigation. From the results obtained, it was evident that the administration of 50 and 100 mg saffron/kg body weight could significantly inhibit the in vivo genotoxicity of these genotoxins. However, all the three doses of saffron were effective in exerting a protective effect against urethane.

Protective effects of saffron (Crocus sativus Linn.) on genotoxins-induced oxidative stress in Swiss albino mice.

The modifying effects of the aqueous extract of saffron (dried stigmas of Crocus sativus Linn.) on cisplatin (CIS), cyclophosphamide (CPH), mitomycin-C (MMC) and urethane (URE) induced alterations in lipid peroxidation and antioxidant status were investigated in Swiss albino mice. Three doses of saffron (20, 40 and 80 mg/kg body weight) were orally administered to mice for 5 consecutive days prior to administration of genotoxins. A significant reduction in the extent of lipid peroxidation with a concomitant increase in the liver enzymatic (SOD, CAT, GST, GPx) and non-enzymatic antioxidants (reduced glutathione) were observed in saffron pretreated animals compared with the genotoxins alone treated animals. However, the modulatory effects were not always dose dependent. Our data suggest that saffron may exert its chemopreventive effects by modulation of lipid peroxidation, antioxidants and detoxification systems.