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Piper nigrum is a widely used plant in traditional remedies and known for its numerous biological properties. However, fraction-based antioxidant activity and their antimutagenic potential are not yet fully investigated. Different extracts of the seeds P. nigrum were obtained by sequential extraction in different solvents. All extracts were evaluated for antibacterial and antioxidant activities using different methods. The most active fraction was analyzed for antimutagenic activity using the Ames Salmonella test. The antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) was found to be more prominent compared to ESβL producing Klebsiella pneumoniae isolates. The MIC values were found to be lower against MRSA than K. pneumoniae. The extract showing highest antioxidant activity (methanol extract) was further tested for antimutagenic activity both against direct and indirect-acting mutagens. A varying level of antimutagenic activity was shown by methanol extract at highest tested concentration (200 µg/plate). Alkaloids, phenols, and flavonoids were detected as major class of compounds in methanol extract. Gas chromatography-mass spectrometry (GC–MS) analysis showed the presence of various phytocompounds. Based on molecular docking of two major active phytocompounds (piperine and copaene), they were found to interact at the minor groove of DNA. Molecular dynamics (MD) simulation revealed that both the ligands were quite stable with DNA under physiological conditions. The ability of phytocompounds to interact with DNA might be reducing the interaction of mutagens and could be one of the possible mechanism of anti-mutagenic activity of P. nigrum extract. This study highlights the antioxidant and antimutagenic potential of Piper nigrum. The role of phytocompounds present in the bioactive extract is needed to be explored further for herbal drug research.

Piper nigrum is a widely used plant in traditional remedies and known for its numerous biological properties. However, fraction-based antioxidant activity and their antimutagenic potential are not yet fully investigated. Different extracts of the seeds P. nigrum were obtained by sequential extraction in different solvents. All extracts were evaluated for antibacterial and antioxidant activities using different methods. The most active fraction was analyzed for antimutagenic activity using the Ames Salmonella test. The antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) was found to be more prominent compared to ESβL producing Klebsiella pneumoniae isolates. The MIC values were found to be lower against MRSA than K. pneumoniae. The extract showing highest antioxidant activity (methanol extract) was further tested for antimutagenic activity both against direct and indirect-acting mutagens. A varying level of antimutagenic activity was shown by methanol extract at highest tested concentration (200 µg/plate). Alkaloids, phenols, and flavonoids were detected as major class of compounds in methanol extract. Gas chromatography-mass spectrometry (GC–MS) analysis showed the presence of various phytocompounds. Based on molecular docking of two major active phytocompounds (piperine and copaene), they were found to interact at the minor groove of DNA. Molecular dynamics (MD) simulation revealed that both the ligands were quite stable with DNA under physiological conditions. The ability of phytocompounds to interact with DNA might be reducing the interaction of mutagens and could be one of the possible mechanism of anti-mutagenic activity of P. nigrum extract. This study highlights the antioxidant and antimutagenic potential of Piper nigrum. The role of phytocompounds present in the bioactive extract is needed to be explored further for herbal drug research.

// Sujita Khanal 1 , Brian S. Shumway 2 , Maryam Zahin 3 , Rebecca A. Redman 3, 4 , John D. Strickley 3, 4 , Patrick J. Trainor 3 , Shesh N. Rai 3 , Shin-je Ghim 3 , Alfred Bennett Jenson 3 and Joongho Joh 3, 4, 5 1 Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA, USA 2 Department of Surgical and Hospital Dentistry, University of Louisville School of Dentistry, Louisville, KY, USA 3 James Graham Brown Cancer Center, University of Louisville, Louisville, KY, USA 4 Department of Medicine, School of Medicine, University of Louisville, Louisville, KY, USA 5 Center for Predictive Medicine, University of Louisville, Louisville, KY, USA Correspondence to: Joongho Joh, email: joongho.joh@louisville.edu Keywords: human papillomavirus (HPV); HPV integration; HPV methylation; high-grade oral epithelial dysplasia (hgOED); head and neck cancer Received: April 10, 2018 Accepted: May 25, 2018 Published: July 13, 2018 ABSTRACT This study evaluated the integration and methlyation of human papillomavirus type 16 (HPV16) in head and neck squamous cell carcinoma (HNSCC) and its oral precursor, high-grade oral epithelial dysplasia (hgOED). Archival samples of HPV16-positive hgOED ( N = 19) and HNSCC ( N = 15) were evaluated, along with three HNSCC (UMSCC-1, -47 and -104) and two cervical cancer (SiHa and CaSki) cell lines. HgOED cases were stratified into three groups with increasing degrees of cytologic changes (mitosis, karyorrhexis and apoptosis). The viral load was higher and the E2/E6 ratio lower (indicating a greater tendency toward viral integration) in group 3 than in groups 1 or 2 ( p = 0.002, 0.03). Methylation was not observed in hgOED cases and occurred variably in only three HNSCC cases (26.67%, 60.0% and 93.3%). In HNSCC cell lines, lower E7 expression correlated with higher levels of methylation. HgOED with increased cytologic change, now termed HPV-associated oral epithelial dysplasia (HPV-OED), exhibited an increased viral load and a tendency toward DNA integration, suggesting a potentially increased risk for malignant transformation. More detailed characterization and clinical follow-up of HPV-OED patients is needed to determine whether HPV-OED is a true precursor to HPV-associated HNSCC and to clarify the involvement of HPV in HNSCC carcinogenesis.

Previous studies of naturally occurring mouse papillomavirus (PV) MmuPV1-induced tumors in B6.Cg-Foxn1nu/nu mice suggest that T cell deficiency is necessary and sufficient for the development of such tumors. To confirm this, MmuPV1-induced tumors were transplanted from T cell-deficient mice into immunocompetent congenic mice. Consequently, the tumors regressed and eventually disappeared. The elimination of MmuPV1-infected skin/tumors in immunocompetent mice was consistent with the induction of antitumor T cell immunity. This was confirmed by adoptive cell experiments using hyperimmune splenocytes collected from graft-recipient mice. In the present study, such splenocytes were injected into T cell-deficient mice infected with MmuPV1, and they eliminated both early-stage and fully formed tumors. We clearly show that anti-tumor T cell immunity activated during tumor regression in immunocompetent mice effectively eliminates tumors developing in T cell-deficient congenic mice. The results corroborate the notion that PV-induced tumors are strongly linked to the immune status of the host, and that PV antigens are major anti-tumor antigens. Successful anti-PV T cell responses should, therefore, lead to effective anti-tumor immune therapy in human PV-infected patients.

Fruits, vegetables and medicinal herbs rich in phenolics antioxidants contribute toward reduced risk of age-related diseases and cancer. In this study, Psidium guajava leaf extract was fractionated in various organic solvents viz. petroleum ether, benzene, ethyl acetate, ethanl and methanol and tested for their antioxidant and antimutagenic properties. Methanolic fraction showed maximum antioxidant activity comparable to ascorbic acid and butylated hydroxyl toluene (BHT) as tested by DPPH free radical scavenging, phosphomolybdenum, FRAP (Fe3 + reducing power) and CUPRAC (cupric ions (Cu2+) reducing ability) assays. The fraction was analyzed for antimutagenic activities against sodium azide (NaN3), methylmethane sulfonate (MMS), 2-aminofluorene (2AF) and benzo(a)pyrene (BP) in Ames Salmonella tester strains. The methanol extracted fraction at 80 μg/ml concentration inhibited above 70% mutagenicity. Further, phytochemical analysis of methanol fraction that was found to be most active revealed the pr...

Antioxidant properties of methanol extracts of 20 plants were studied by four different assays over a range of concentrations (12.5–400 µg mL−1), and compared with standard antioxidants’ ascorbic acid and butylated hydroxytoluene. Ascorbic acid–equivalent antioxidant activity of extracts by the phosphomolybdenum method ranged from 392.8 to 2942.7 µmoles g−1 at 400 µg mL−1, whereas DPPH radical scavenging was between 30.9% and 95.2%; extracts also showed strong activity by FRAP and CUPRAC assays. The extracts demonstrated dose-dependent activity and strong correlation between antioxidant activities and total phenolics. Total phenolic content was the highest in Syzygium aromaticum (279 mg GAE g−1) and the lowest in Sesame indicum (28.16 mg GAE g−1).

Spices and herbs are recognized as sources of natural antioxidants and thus play an important role in the chemoprevention of diseases and aging. Piper cubeba is one among them and known for its medicinal properties for decades. Various biological activities are associated with its extract and phytocompounds. However, the anti-mutagenic activity of antioxidant rich extract is less explored. In this study, we performed the fraction-based antioxidant activity of P. cubeba using four different assays and evaluated the anti-mutagenic activity of most potent antioxidant fraction using Salmonella typhimurium tester strains against four mutagens (methyl methanesulfonate [MMS], sodium azide [SA], benzo(a)pyrene, and 2-aminoflourene) respectively. Among all tested fractions at 25–200 µg/ml, ethanolic extract revealed highest antioxidant activity and significant anti-mutagenicity against both direct and indirect acting mutagens at least one tester strain. Phytochemical analysis by gas chromatography–mass spectrometry (GC/MS) revealed the presence of various phytocompounds including copaene, isocaryophyllene, α-cubebene, etc. Molecular docking studies on DNA binding interactions of GC/MS detected phytocompounds highlight the possible mode of binding. In summary, these in vitro studies have provided the scientific basis for validation of using this plant in the traditional system of medicine and highlighted the need for exploring the role of various compounds for therapeutic efficacy. On the other hand, synergistic interaction among phytocompounds is to be explored to optimize or standardize the extracts for the exploitation in modern phytomedicine.

Background Current vaccines against Human Papillomavirus (HPV) are highly effective and based on recombinant virus-like particles (VLPs) of the major capsid protein L1. Since these vaccines are HPV type-specific and expensive for global implementation, an alternative, broader-spectrum immunogen would be the N-terminus of the minor capsid protein L2 that induces low titered broadly cross-neutralizing antibodies. Here we analyzed the reactivity of different synthetic L2 peptides containing N-terminus amino acids 17–36 in order to test their antigenicity. Methods Different synthetic peptides were designed to target the 17–36 amino acid sequences, present in highly antigenic amino-terminus of L2 protein. Six different peptides including Cys22–Cys28 disulfide bonded cyclized L2 peptide were examined for their antigenicity against mouse monoclonal antibody RG-1 and rabbit polyclonal antisera to HPV L2 by enzyme-linked immunosorbent assay (ELISA). Results Here we report that the cyclized form of synthetic L2 peptide, which is formed through Cys22–Cys28 disulfide bridges, has the highest reactivity to antibodies than other synthetic L2 peptides. Conclusion A cyclized L2 peptide has potential to be an excellent candidate to formulate a low-cost, broadly protective pan-oncogenic HPV vaccine.

The Florida manatee (Trichechus manatus latirotris) is considered a threatened aquatic mammal in United States coastal waters. Over the past decade, the appearance of papillomavirus-induced lesions and viral papillomatosis in manatees has been a concern for those involved in the management and rehabilitation of this species. To date, three manatee papillomaviruses (PVs) have been identified in Florida manatees, one forming cutaneous lesions (TmPV1) and two forming genital lesions (TmPV3 and TmPV4). In this study, we identified DNA sequences with the potential to form G-quadruplex structures in all three PVs. G-quadruplex structures (G4) are guanine-rich nucleic acid sequences capable of forming secondary structures in DNA and RNA. In humans, G4 are known to regulate molecular processes such as transcription and translation. Although G4 have been identified in several viral genomes, including human PVs, no attempt has been made to identify G4 in animal PVs. We found that sequences capable of forming G4 were present on both DNA strands and across coding and non-coding regions on all PVs. The vast majority of the identified sequences would allow the formation of non-canonical structures with only two G-tetrads. The formation of one such structure was supported through biophysical analysis. Computational analysis demonstrated enrichment of G4 sequences on the reverse strand in the E2/E4 region on all manatee PVs and on the forward strand in the E2/E4 region on one genital PV. Several G4 sequences occurred at similar regional locations on all PVs, most notably on the reverse strand in the E2 region. In other cases, G4 were identified at similar regional locations only on PVs forming genital lesions. On all PVs, G4 sequences were located near putative E2 binding sites in the non-coding region. Together, these findings suggest that G4 are likely regulatory elements in manatee PVs.Author summaryG-quadruplex structures (G4) are found in the DNA and RNA of many species and are known to regulate the expression of genes and the synthesis of proteins, among other important molecular processes. Recently, these structures have been identified in several viruses, including the human papillomavirus (PV). As regulatory structures, G4 are of great interest to researchers as drug targets for viral control. In this paper, we identify the first G4 sequences in three PVs infecting a non-human animal, the Florida manatee. Through computational and biophysical analysis, we find that a greater variety of sequence patterns may underlie the formation of these structures than previously identified. The sequences are found in all protein coding regions of the virus and near sites for viral replication in non-coding regions. Furthermore, the distribution of these sequences across the PV genomes supports the notion that sequences are conserved across PV types, suggesting they are under selective pressure. This paper extends previous research on G4 in human PVs with additional evidence for their role as regulators. The G4 sequences we identified also provide potential regulatory targets for researchers interested in controlling this virus in the Florida manatee, a threatened aquatic mammal.

The Florida manatee (Trichechus manatus latirotris) is a threatened aquatic mammal in United States coastal waters. Over the past decade, the appearance of papillomavirus-induced lesions and viral papillomatosis in manatees has been a concern for those involved in the management and rehabilitation of this species. To date, three manatee papillomaviruses (TmPVs) have been identified in Florida manatees, one forming cutaneous lesions (TmPV1) and two forming genital lesions (TmPV3 and TmPV4). We identified DNA sequences with the potential to form G-quadruplex structures (G4) across the three genomes. G4 were located on both DNA strands and across coding and non-coding regions on all TmPVs, offering multiple targets for viral control. Although G4 have been identified in several viral genomes, including human PVs, most research has focused on canonical structures comprised of three G-tetrads. In contrast, the vast majority of sequences we identified would allow the formation of non-canonical structures with only two G-tetrads. Our biophysical analysis confirmed the formation of G4 with parallel topology in three such sequences from the E2 region. Two of the structures appear comprised of multiple stacked two G-tetrad structures, perhaps serving to increase structural stability. Computational analysis demonstrated enrichment of G4 sequences on all TmPVs on the reverse strand in the E2/E4 region and on both strands in the L2 region. Several G4 sequences occurred at similar regional locations on all PVs, most notably on the reverse strand in the E2 region. In other cases, G4 were identified at similar regional locations only on PVs forming genital lesions. On all TmPVs, G4 sequences were located in the non-coding region near putative E2 binding sites. Together, these findings suggest that G4 are possible regulatory elements in TmPVs.

We isolated two new manatee papillomavirus (PV) types, TmPV3 and TmPV4, from a Florida manatee (Trichechus manatus latirostris). Two PV types were previously isolated from this species. TmPV1 is widely dispersed amongst manatees and a close-to-root PV; not much is known about TmPV2. The genomes of TmPV3 and TmPV4 were 7622 and 7771 bp in size, respectively. Both PVs had a genomic organization characteristic of all PVs, with one non-coding region and seven ORFs, including the E7 ORF that is absent in other cetacean PVs. Although these PVs were isolated from separate genital lesions of the same manatee, an enlarged E2/E4 ORF was found only in the TmPV4 genome. The full genome and L1 sequence similarities between TmPV3 and TmPV4 were 63.2 and 70.3 %, respectively. These genomes shared only 49.1 and 50.2 % similarity with TmPV1. The pairwise alignment of L1 nucleotide sequences indicated that the two new PVs nested in a monophyletic group of the genus Rhopapillomavirus, together with the cutaneotropic TmPV1 and TmPV2.

Punicalagin (PC) is an ellagitannin found in the fruit peel ofPunica granatum. We have demonstrated antioxidant and antigenotoxic properties ofPunica granatumand showed that PC and ellagic acid (EA) are its major constituents. In this study, we demonstrate the antimutagenic potential, inhibition of BP-induced DNA damage, and antiproliferative activity of PC and EA. Incubation of BP with rat liver microsomes, appropriate cofactors, and DNA in the presence of vehicle or PC and EA showed significant inhibition of the resultant DNA adducts, with essentially complete inhibition (97%) at 40 μM by PC and 77% inhibition by EA. Antimutagenicity was tested by Ames test. PC and EA dose-dependently and markedly antagonized the effect of tested mutagens, sodium azide, methyl methanesulfonate, benzo[a]pyrene, and 2-aminoflourine, with maximum inhibition of mutagenicity up to 90 percent. Almost all the doses tested (50–500 μM) exhibited significant antimutagenicity. A profound antiproliferative effect on human lung cancer cells was also shown with PC and EA. Together, our data show that PC and EA are pomegranate bioactives responsible for inhibition of BP-induced DNA adducts and strong antimutagenic, antiproliferative activities. However, these compounds are to be evaluated in suitable animal model to assess their therapeutic efficacy against cancer.

Medicinal plants and plant-derived bioactive compounds are well known for their contribution to primary health care as a source novel drug discovery for various ailments. Emergence and spread of microbial drug resistance due to various mechanisms has impacted the efficacy of almost all old and new antibacterial drugs. The biofilm mode of microbial growth has significantly increased the survival strategies and resistance levels of microbes to drugs, making the treatment of infections more difficult. Currently, efforts are going on to develop novel strategies including targeting biofilms to treat infections. Various natural products are known to inhibit biofilm formation or preformed biofilms. In recent years medicinal plants and phytocompounds were reported with promising antibiofilm activity in vitro from different parts of the world. In this chapter we have reviewed the current literature on antibiofilm agents derived from medicinal plants and/or plant-derived compounds. Plant extracts and phytocompounds of various classes have been found effective against bacterial or fungal biofilms, with some compounds showing activity against both. Such compounds are expected to be effective against mixed biofilms. Interestingly, certain quorum-sensing inhibiting plant extracts or compounds can also inhibit biofilms made by bacteria such as Pseudomonas aeruginosa. The majority of the antibiofilm phytocompounds identified so far have been tested in vitro; however, only a few compounds have been reported effective under in vivo condition. This could be due to the lack of access of the investigators to suitable animal models for different diseases to assess the therapeutic efficacy of these antibiofilm agents. The results of this chapter indicated that the phytocompounds may be effective alone or in combination with antibiotics, as in the treatment of systemic infection. However, further investigation on their mode of action and in vivo efficacy are prerequisites to obtain broad-spectrum antifungal agents of clinical value.

It is well known that the intake of antioxidants with increased consumption of fruits and vegetables and medicinal herbs contributes towards reduced risk of certain diseases including cancers. This study aims to evaluate the broad-spectrum antioxidant and antimutagenic activities as well as to elucidate phytochemical profile of an Indian medicinal plantMurraya koenigii(curry) leaves. Leaves of the plant were successively fractionated in various organic solvents. Benzene fraction demonstrated the highest phenolic content followed by petroleum ether. The benzene fraction showed maximum antioxidant activity in all tested assays, namely, phosphomolybdenum, 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical, ferric reducing antioxidant power (FRAP) and cupric reducing antioxidant capacity (CUPRAC) assays. Based on the promising broad-spectrum antioxidant activity, benzene fraction was further evaluated for antimutagenic activity and showed a dose-dependent antimutagenic response in AmesSalmonellamutagenicity assay. It inhibited 72–86% mutagenicity induced by sodium azide, methyl methanesulfonate, benzo(a)pyrene, and 2-aminoflourene at the maximum tested concentration (100 μg/mL) inSalmonella typhimuriumtester strains. At least 21 compounds were detected by GC/MS. The findings clearly demonstrated that phenolic-rich benzene fraction has promising broad-spectrum antioxidant and antimutagenic property and needs further evaluation to exploit its therapeutic potential.

Cervical cancer is the most common malignancy among women particularly in developing countries, with human papillomavirus (HPV) 16 causing 50% of invasive cervical cancers. A plant-based HPV vaccine is an alternative to the currently available virus-like particle (VLP) vaccines, and would be much less expensive. We optimized methods to express HPV16 L1 protein and purify VLPs from tobacco (Nicotiana benthamiana) leaves transfected with the magnICON deconstructed viral vector expression system. L1 proteins were extracted from agro-infiltrated leaves using a series of pH and salt mediated buffers. Expression levels of L1 proteins and VLPs were verified by immunoblot and ELISA, which confirmed the presence of sequential and conformational epitopes, respectively. Among three constructs tested (16L1d22, TPL1d22, and TPL1F), TPL1F, containing a full-length L1 and chloroplast transit peptide, was best. Extraction of HPV16 L1 from leaf tissue was most efficient (> 2.5% of total soluble protein) with a low-salt phosphate buffer. VLPs were purified using both cesium chloride (CsCl) density gradient and size exclusion chromatography. Electron microscopy studies confirmed the presence of assembled forms of HPV16 L1 VLPs. Collectively; our results indicated that chloroplast-targeted transient expression in tobacco plants is promising for the production of a cheap, efficacious HPV16 L1 VLP vaccine. Studies are underway to develop plant VLPs for the production of a cervical cancer vaccine.

Multidrug resistance (MDR) in microorganisms is a cause of major concern for clinicians and pharmaceutical industries. Continuous development of new antimicrobial drugs with multiple targets and potentials is expected to efficiently combat MDR in these microorganisms. In a continued exploration of new antimicrobial drug leads, 11 marine natural products, semisynthetic, or related synthetic analogs (1-11) and two tobacco cembranoids (12 and 13) were screened for their antimicrobial, antioxidant, and antimutagenic activities. Eight compounds showed varying levels of both antibacterial and antifungal activities. Compounds such as 17-O-methyllatrunculin-A, verongiaquinol, (1S,2E,4R,6R,7E,11E)-2,7,11-cembratriene-4,6-diol), and manzamine-A showed a broad spectrum of activity, inhibiting six of seven tested bacteria with zone of inhibition diameter from 9 to 30 mm. Four of these active compounds also showed antifungal activity. The findings of the in vitro time-kill assay of the most active compound, verongiaquinol, against Staphylococcus aureus indicated its subinhibitory effect at the level lower than the minimal inhibitory concentration (MIC) values (i.e., 2 and 4 µg/mL). At the MIC (8 µg/mL), bacterial cells were completely killed within 18 hours of incubation. DPPH free radical scavenging activity was demonstrated by five compounds in the range of 89.65-36.19% decolorization. Further, four compounds evaluated for their antimutagenic activity against the directly acting mutagens, methyl methanesulfonate and sodium azide, in Salmonella typhimurium strains, interestingly, showed no sign of mutagenicity. Verongiaquinol and manzamine A, in fact, reduced the mutagenicity by 50-75% at a dose of 5 µg/plate in different test strains. Our study seems to provide some novel antimicrobial leads with strong antioxidant potential and the associated ability of antimutagenicity.

Discovery of quorum sensing (QS) system to coordinate virulence and biofilm formation in bacterial pathogens has triggered search for safe, stable and non-toxic anti-QS compounds from natural products. Ethanolic extracts of 24 Indian medicinal plants were tested by agar well and disc diffusion assay for anti-QS activity using Chromobacterium violaceum (CV12472 and CVO26) reporter strains. AHL from C. violaceum CV31532 was isolated and partially purified for its use in CVO26 based bioassay. Effect on swarming-motility of Pseudomonas aeruginosa (PAO1) was also recorded at sub-MIC concentrations of extracts. Of the 24 medicinal plants screened Hemidesmus indicus (L.) Schult (root), Holarrhena antidysenterica (Roth) A.DC. (bark), Mangifera indica L. (seed) Punica granatum L. (pericarp) and Psoralea corylifolia L. (seed) demonstrated varying level of inhibition of violacein production in the reporter strains. Moreover, a significant reduction in swarms was recorded over control. The inhibition of violacein production and swarming motility may be due to direct or indirect interference on QS by active constituents or the interactive effect of different phytocompounds present in the extracts. These plant extracts may be selected for activity guided fractionation to identify and characterize the active principle.

Bacteria use the language of low-molecular-weight ligands to assess their population densities in a process called quorum sensing (QS). Different types of quorum sensing pathways are present in Gram-negative and Gram-positive bacteria. Signal molecules most commonly used in Gram-negative bacteria are acyl homoserine lactones. In recent years, a substantial amount of literature and data have been available on bacterial QS. Recently, interest in modulation of quorum sensing with different approaches has increased among scientific communities. In this chapter, we provide an updated overview on bacterial QS, assays and methods for detecting signal molecules, and various approaches to inhibit AHL-based quorum sensing. Significance of QS interference by prokaryotic and eukaryotic organisms in relation to plant health and the environment is discussed here.

Over the past few decades, scientific research has indicated a credible basis for some of the traditional ethnomedicinal uses of pomegranate. This study aims to evaluate the broad spectrum antioxidant and antimutagenic activities of peel extracts of pomegranate. The sequentially extracted Punica granatum peel fractions were tested for their antioxidant activity by DPPH free radical scavenging, phosphomolybdenum, FRAP (Fe 3+ reducing power) and CUPRAC (cupric ions (Cu 2+ ) reducing ability) assays. The methanol fraction showed highest antioxidant activity by all the four in vitro assays comparable to ascorbic acid and butylated hydroxy toluene (BHT) followed by activity in ethanol, acetone, and ethyl acetate fractions. Based on the promising antioxidant activities, the methanol fraction was evaluated for antimutagenic activity by Ames Salmonella /microsome assay against sodium azide (NaN 3 ), methyl methane sulphonate (MMS), 2-aminofluorene (2-AF) and benzo(a)pyrene (B(a)P) induced mutagenicity in Salmonella typhimurium (TA97a, TA98, TA100 and TA102) tester strains. The methanol fraction showed no sign of mutagenicity at tested concentration of 10–80 μg/mL. This fraction showed antimutagenic activity against NaN 3 and MMS with percent inhibition of mutagenicity ranging from 66.76% to 91.86% in a concentration-dependent manner. Similar trend of inhibition of mutagenicity (81.2–88.58%) against indirect mutagens (2-AF and B(a)P) was also recorded. Phytochemical analysis by HPLC, LC–MS and total phenolic content revealed high content of ellagitannins which might be responsible for promising antioxidant and antimutagenic activities of P. granatum peel extract. Further, contribution of bioactive compounds detected in this study is to be explored to understand the exact mechanism of action as well as their therapeutic efficacy.

Effect of growing seedling, seeds and seedlings extracts from seven leguminous plants (Pisum sativum, Vigna radiata, Vigna mungo, Cajanus cajan, Lentil culinaris, Cicer arietinum and Trigonella foenum graecum) were screened for their ability to influence quorum sensing controlled pigment production in Chromobacterium violaceum indicator strains (CV12472 and CVO26). Germinating seedling and seedling extracts of only P. sativum (pea) showed inhibition of violacein production. Interestingly, the T. foenum graecum (fenugreek) seed extracts enhances the pigment production. Quorum sensing regulated swarming motility in Pseudomonas aerugionsa PAO1 was reduced by pea seedling extract while enhanced by the fenugreek seed extracts. These findings suggest that plant metabolites of some legumes interact actively with bacterial quorum sensing and could modulate its associated functions.

The epidemiological data suggest that the incidence and prevalence of serious mycoses continues to be a public health problem. The increased use of antifungal agents has resulted in the development of resistance to these drugs. The spread of multidrug-resistant strains of fungus and the reduced number of drugs available make it necessary to discover new classes of antifungals from natural products including medicinal plants. Historically, herbs and spices have enjoyed a rich tradition of use for their medicinal properties and provide unlimited opportunities for new drug leads because of the huge chemical diversity. Assays of bioactive compounds have been reported with good antifungal properties in vitro or in vivo. It is almost impossible to discuss the various characteristics of these plants such as mode of action and extraction of active compounds in a single review. Therefore, we have focussed here mainly on the antifungal plant extracts, their use against pathogeinc and drug resistant fungi. The various classes of compounds such as phenolics, terpenoids, saponins, and alkaloids, etc., are discussed in detail. The new emerging classes of antifungal proteins and peptides are also reviewed briefly. In this chapter, we also describe the technical aspects related to the methodology for screening and identification of antifungal compounds. The technical aspects regarding the use of reliable methodology of extraction, screening, bioautography, and identification of pure compounds from crude extracts and fractions are also discussed here.

Invasive fungal infections are a major problem in immunocompromised patients. This has necessitated an increased interest in the development of new antifungals to treat these life-threatening infections. However, our means of combating fungal infections are still lagging behind those for bacterial infections, due to toxicity and the lower clinical efficacy of available antifungals against some invasive fungal infection. Thus, more efforts are needed in antifungal drug discovery, as well as in developing effective ways of minimizing toxicity and improving delivery of available antifungal drugs. One approach is the effective use of newer antifungal agents in combination therapy against invasive aspergillosis, cryptococcosis and candidiasis. On the other hand, identifying and validating new antifungal drug targets is a prerequisite for new antifungal drug discovery. These new targets might be discovered by both conventional but improved assays and genomic approaches. Also, targeting virulence is expected to be a new paradigm for antifungals. In this chapter, an attempt is made to describe recent progress in combinational therapy and new approaches to antifungal drug discovery.

The ajowain (Carum copticum (L.)) is a popular spice and traditionally used in Indian system of medicine. Considering the importance of natural products in modern phytomedicine, the antioxidant and antimutagenic activities of C. copticum fruits extract and its fractions were evaluated. The methanol fraction showed highest antioxidant activity by phosphomolybdenum (2087.7 micromol) and DPPH assay (90.2%) followed by other fractions comparable to ascorbic acid and BHT. Based on antioxidant activity, methanol fraction was evaluated for antimutagenic potential against direct acting mutagens sodium azide (NaN(3)) and methyl methane sulphonate (MMS) and indirect acting mutagens 2-aminofluorene (2-AF) and benzo(a)pyrene (B(a)P), using Salmonella typhimurium (TA97a, TA98, TA100, and TA102) tester strains. The methanolic fraction showed no sign of mutagenicity at tested concentrations (25-100 microg/plate). Antimutagenic activity was recorded with inhibition of mutagenicity ranging from 10.8% to 83.1% in a concentration dependent manner. The phytochemical analysis by IR, HPLC, GC-MS, and total phenolic assay revealed a high content of phenolic terpenoids. Further, characterization of active principle is needed to understand the mechanism of action and therapeutic efficacy in vivo.

Aims: To evaluate quorum sensing (QS) inhibitory activity of plant essential oils using strains of Chromobacterium violaceum (CV12472 and CVO26) and Pseudomonas aeruginosa (PAO1). Methods and Results: Inhibition of QS-controlled violacein production in C. violaceum was assayed using disc diffusion and agar well diffusion method. Of the 21 essential oils, four oils showed varying levels of anti-QS activity. Syzygium aromaticum (Clove) oil showed promising anti-QS activity on both wild and mutant strains with zones of pigment inhibition 19 and 17 mm, respectively, followed by activity in cinnamon, lavender and peppermint oils. The effect of clove oil on the extent of violacein production was estimated photometrically and found to be concentration dependent. At sub-MICs of clove oil, 78·4% reduction in violacein production over control and up to 78% reduction in swarming motility in PAO1 over control were recorded. Gas chromatography–mass spectrometry analysis of clove oil indicated presence of many phytocompounds. Eugenol, the major constituent of clove oil could not exhibit anti-QS activity. Conclusions: Presence of anti-QS activity in clove oil and other essential oils has indicated new anti-infective activity. The identification of anti-QS phytoconstituents is needed to assess the mechanism of action against both C. violaceum and Ps. aeruginosa. Significance and Impact of the study: Essential oils having new antipathogenic drugs principle because of its anti-QS activity might be important in reducing virulence and pathogenicity of drug-resistant bacteria in vivo.

A rapid and efficient one-pot condensation reaction of long-chain alkyl and alkenyl acid hydrazides and nitriles was carried out to afford 3,5-disubstituted-1H-1,2,4-triazoles. The compounds 5a–o were screened for in-vitro antibacterial activity against the representative panel of two Gram-positive and two Gram-negative bacteria. All the synthesized compounds were also tested for their inhibitory action against five strains of fungi. The various compounds show potent inhibitory action against test organisms. The compounds 5a–o were characterized on the basis of elemental analysis and spectral data.

Methanolic extracts of Acorus calamus (Rhizome), Hemidesmus indicus (Stem), Holarrhena antidysenterica (Bark) and Plumbago zeylanica (Root), were tested for their antimutagenic potential. These extracts, at tested concentrations, showed no sign of mutagenicity to Salmonella typhimurium tester strains. The extracts of the plants exhibited varying level of antimutagenicity. At a dose of 100 microg/plate, the extracts exhibited the inhibition of His+ revertants from 18.51% to 82.66% against direct acting mutagens, methyl methanesulphonate (MMS) and sodium azide (NaN3) induced mutagenicity in Salmonella tester strains TA 97a, TA 100, TA 102 and TA 104. However, at lower concentrations (25 and 50 mcirog/plate) of the plant extracts, a decrease in antimutagenic activity was recorded. Dose dependent antimutagenic activity of the extracts is also evident from linear regression analysis of the data. The over all antimutagenic potential of above four extracts was found to be in order of A. calamusH. indicusH. antidysentericaP. zeylanica. Further, total phenolic content of these extracts did not correlate with its antimutagenic activity in A. calamus and P. zeylanica.

A rapid and efficient one-pot condensation reaction of long-chain alkyl and alkenyl acid hydrazides and nitriles was carried out to afford 3,5-disubstituted-1H-1,2,4-triazoles. The compounds 5a–o were screened for in-vitro antibacterial activity against the representative panel of two Gram-positive and two Gram-negative bacteria. All the synthesized compounds were also tested for their inhibitory action against five strains of fungi. The various compounds show potent inhibitory action against test organisms. The compounds 5a–o were characterized on the basis of elemental analysis and spectral data.

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