Antioxidant compounds from microbial sources: A review.

Free radicals are one or more unpaired electrons containing reactive molecules, which can damage nucleic acids, proteins, carbohydrates, and lipids, leading to several diseases including early aging, cancer and atherosclerosis. Antioxidants can scavenge these free radicals to prevent cellular damage by ultimately reducing the oxidative stress and thus have a beneficial effect on human health. Epidemiological studies have already revealed that higher intake of antioxidants as food supplements results in reduced risk of many diseases. Exploring natural antioxidants and its role in human health & nutrition is an emerging field. Several biological sources like medicinal plants, vegetables, spices and fruits have been evaluated as sources of potentially safe natural antioxidants. Beside plants, microorganisms are the potential source of novel bioactive compounds to be used in medical, agricultural, and industrial sectors. As compared to plants, microbes can be grown under controlled conditions at a faster rate, which make them a potential source of natural bioactive molecules for food and nutraceutical applications. This review summarizes the potential of different microorganisms including actinomycetes, bacteria, blue green algae, fungi, lichens and mushrooms to be explored as the source of such bioactive compounds.

Stimuli Responsive In Situ Gelling Systems Loaded with PLGA Nanoparticles of Moxifloxacin Hydrochloride for Effective Treatment of Periodontitis.

The objectives of the present research work were systematic development of novel in situ gel formulation containing nanoparticles for localised delivery of moxifloxacin against bacterial periodontitis. PLGA nanoparticles were prepared and optimised in a systematic manner. Factor screening was performed with the help of half-factorial design to identify the influential factors, while response surface optimisation of the nanoparticles was conducted using central composite design. The optimum nanoparticle formulation was chosen on the basis of lower particle size, higher drug entrapment and controlled drug release characteristics up to 1 week time period, while the optimum in situ gel was selected on the basis of faster gelling and higher viscosity and gel strength properties for improved retention in the periodontium. In vivo histopathological studies and in vivo gamma scintigraphy studies revealed the extended release, superior efficacy and enhanced retention of nanoparticle-loaded in situ gelling system. Results obtained from in vivo histopathological studies after 1 week treatment with in situ gel formulation containing nanoparticles of moxifloxacin were found to be better than with 3 weeks treatment of marketed gel formulation. Overall, the studies ratify successful development of an effective site-specific drug delivery system with enhanced biopharmaceutical attributes for the periodontitis treatment.

Effect of rumen-protected nutrients on feed intake, body weights, milk yield, and composition in Murrah buffaloes during early lactation.

The effect of rumen-protected nutrients (bypass fat, BPF; bypass protein, BPP; or their combination, BPPF) was investigated in Murrah buffaloes during the early stage of lactation. Forty Murrah buffaloes (BW 531.92 ± 10.85 kg) just after parturition were randomly distributed into four groups according to parity and milk production. Buffaloes individually fed ration from day 0 to 90 postpartum according to feeding group and nutrient requirement. Control and BPF fed groups received a concentrate mixture, CM1 with 25% rumen-protected protein (using barley, wheat bran, and mustard oil cake), BPP and BPPF groups received a second concentrate mixture, CM2 with 40% rumen-protected protein (using barley, de-oiled rice bran, and cottonseed cake). Bypass fat fed groups (BPF and BPPF) additionally were supplemented with 15 g BPF (Ca salt of long-chain fatty acids) per kg milk yield in their respective concentrate mixtures. Dry matter intake, body weights, body condition score, and total milk yield were similar between groups (P > 0.05). Fat-corrected milk (FCM) production was improved (14.5%, P > 0.05) in groups fed BPP and BPPF, while significant (19.45%, P < 0.05) improvement was observed in BPF-fed group. Overall mean values of milk fat, solid not fat, protein, lactose, and total solids were found to be high (P < 0.05) in treatment groups as compared with control values. It may be concluded that supplementation with BPP or BPF either alone or in combination positively influences the quality of milk produced in Murrah buffaloes during early lactation and BPF additionally had improvement on the quantitative trait of milk as well.

Role of Aqueous Extract of the Wood Ear Mushroom, Auricularia polytricha (Agaricomycetes), in Avoidance of Haloperidol-lnduced Catalepsy via Oxidative Stress in Rats.

Haloperidol-induced catalepsy is an animal model of a psychotic disorder that may be associated with neurodegeneration and free radical damage. Auricularia polytricha is effective in both prevention and treatment of numerous types of neurological disorders. In the present study, anticataleptic activity of aqueous extract of A polytricha (AEAP) at different doses (400 and 600 mg/kg, respectively, p.o.) was studied using haloperidol-induced (1 mg/ kg, i.p.) catalepsy in rats. Repeated treatment with haloperidol (1 mg/kg, i.p.) on each other day for 15 days (days 5, 10, and 15) significantly induced catalepsy in rats. The effect of AEAP at different doses (400 and 600 mg/kg, p.o.) on levels of superoxide dismutase, catalase, and glutathione reductase as well as inhibition of lipid peroxidation in the forebrain region was assessed. After 15 days of treatment, AEAP (400 and 600 mg/kg) significantly inhibited haloperidol-induced catalepsy. Treatment with AEAP (400 and 600 mg/kg) exhibited significant elevation in the levels of superoxide dismutase, catalase, and glutathione reductase as well as lipid peroxidation in the forebrain region compared to the haloperidol-treated group. The study concludes that AEAP (400 and 600 mg/kg) significantly protects animals against haloperidol-induced catalepsy.

Aqueous Extract of Wood Ear Mushroom, Auricularia polytricha (Agaricomycetes), Demonstrated Antiepileptic Activity against Seizure Induced by Maximal Electroshock and Isoniazid in Experimental Animals.

Auricularia polytricha is a popular mushroom found all over the world. This article describes a study of the antiepileptic effect of A. polytricha, a mushroom that is used traditionally for treating asthma, rheumatism, tumors, cough, fever, and epilepsy, and for its antimicrobial effect. We carried out toxicity studies to identify a standard dose of A. polytricha aqueous extract; maximal electroshock (MES)- and isoniazid (INH)-induced seizures in albino mice were used to screen for the extract's antiepileptic activity. Per Organisation for Economic Co-operation and Development Guideline 423, up to 2000 mg/kg body weight of extract was toxic. Animals were treated with aqueous extract at doses of 200, 400, and 600 mg/kg body weight. Phenytoin was used as the reference anticonvulsant drug for comparison. The investigation found a significant interruption in INH-induced clonic seizure. During MES, we found a reduction in the period of hind leg extensor phase; mice exhibited a significant decrease in the duration of hind limb extension after being treated with 400 and 600 mg/kg doses of A. polytricha. Comparable results were obtained in the INH group, as the extract seemed to delay the onset of a clonic seizure. The aqueous extract of A. polytricha showed antiepileptic action against MES- and INH-induced epilepsy in the mice. This extract, however, requires additional study in order to completely explain its active ingredients and their mechanisms of action.

Preparation, characterization and evaluation of the zinc titanate and silver nitrate incorporated wipes for topical chemical and biological decontamination.

Chemical and biological warfare agents have detrimental effects on biological systems. These agents are rapidly absorbed through skin and hence warrant immediate decontamination. Zinc titanate (ZnTiO3), is a well-proven moiety used for neutralizing chemical warfare agents (CWA) and silver is widely used as an antibacterial agent. Spacer fabric sheet, due to its ability to hold large amount of agents, was hydrothermally treated with silver nitrate (AgNO3) and incorporated with nano ZnTiO3 to prepare decontamination (deconwipes). Prepared deconwipe was characterized using X-ray diffraction (XRD) and Scanning Electron Microscopy (SEM). Uniform deposition of nano ZnTiO3 and AgNO3 in the wipe was demonstrated by SEM and XRD. In-vivo dermal decontamination efficacy of the prepared deconwipe was evaluated against nerve agent simulant (diethylchlorophosphate; DCP) and sulfur mustard simulant (2­chloroethyl ethyl sulfide; CEES) on rat model using SEM, Flow Cytometry, Comet Acetylcholinesterase (AChE) inhibition assay and Organ Histopathology. In-vitro antibacterial and antifungal efficacy of the prepared deconwipe was evaluated using growth inhibitory efficacy techniques followed by SEM analysis. SEM and histopathology at dermal level revealed the decontamination efficacy of the prepared deconwipe. Nearly 90% decrease in AChE inhibition was observed in decontaminated rat in place of rate model as compared to the DCP contaminated rats. No significant attenuation was observed in DCP and CEES-induced cell cycle distribution and DNA damage analysis in decontaminated group. Furthermore, >95% inhibition of bacterial and fungal growth, proved its antibacterial and antifungal activity respectively. Thus, prepared deconwipes exhibit promising skin decontamination property against Chemical and biological contaminants.

Aphrodisiac Activity of an Aqueous Extract of Wood Ear Mushroom, Auricularia polytricha (Heterobasidiomycetes), in Male Rats.

Auricularia polytricha is a popular mushroom found all over the world. In this study we considered the effect of an aqueous extract of A. polytricha (AEAP) on restoring sexual performance parameters to normal, evaluated by considering observations of sexual behavior. At 0, 6, 12, 18, and 24 days, the following parameters of sexual performance were identified before and throughout the observations: mount latency, intromission latency, ejaculation latency, mounting frequency, intromission frequency, ejaculation frequency, and postejaculatory interval. Treatment of rats under stress with AEAP showed promising effects on overcoming stress-induced sexual dysfunction, on sexual performance, and on accessory sexual organs and body weight. Mounting latency, intromission latency, ejaculation latency, and postejaculatory interval parameters were significantly decreased by AEAP, whereas mounting frequency, intromission frequency, and ejaculation frequency were significantly increased by AEAP. These properties were identified in sexually dynamic and indolent male rats. We conclude that AEAP has a potent aphrodisiac activity.

Immunomodulatory activities of curcumin-stabilized silver nanoparticles: Efficacy as an antiretroviral therapeutic.

We synthesized and characterized curcumin-stabilized silver nanoparticles (Cur-AgNP) and found them to be 45 nm by dynamic light scattering with a maximum absorbance at 406 nm. We evaluated Cur-AgNP for immunomodulatory activities and their potential as an antiretroviral agent. The antiretroviral effects of Cur-AgNP were determined in ACH-2 cells latently infected with human immunodeficiency virus (HIV)-1. ACH-2 cells, 200,000/ml, were treated with Cur-AgNP for 24-48 h. Expression of HIV-1 LTR and p24, the pro-inflammatory cytokines, IL-1ß, TNF-α, and NF-κB was quantitated. Treatment of ACH-2 cells latently infected with HIV-1 with Cur-AgNP produced no toxic effects but significantly inhibited the expression of HIV-1 LTR (-73%, P < 0.01) and p24 (-57%, P < 0.05), IL-1ßα (-61%, P < 0.01), TNF-αα (-54%, P < 0.05), IL-6 (-68%, P < 0.01), and NF-κB (-79%, P < 0.0001) as compared to untreated controls. Thus, Cur-AgNP have therapeutic potential as direct antiretroviral agents, as well as having immunomodulatory activities inhibiting the expression of pro-inflammatory mediators induced by infection with HIV-1. Experimental controls, such as curcumin alone, and conventional silver nanoparticles capped with citric acid, produced no similar biological effects. We conclude that treatment of HIV-1 infected cells with Cur-AgNP significantly reduced replication of HIV by inhibition of NF-κB nuclear translocation and the downstream expression of the pro-inflammatory cytokines IL-1ß, TNF-α, and IL-6. Subsequent in vivo studies with Cur-AgNP using a humanized mouse model of HIV infection are underway.

Zingiber officinale: Its antibacterial activity on Pseudomonas aeruginosa and mode of action evaluated by flow cytometry.

Biofilm formation, low membrane permeability and efflux activity developed by Pseudomonas aeruginosa, play an important role in the mechanism of infection and antimicrobial resistance. In the present study we evaluate the antibacterial effect of Zingiber officinale against multi-drug resistant strain of P. aeruginosa. The study explores antibacterial efficacy and time-kill study concomitantly the effect of herbal extract on bacterial cell physiology with the use of flow cytometry and inhibition of biofilm formation. Z. officinale was found to inhibit the growth of P. aeruginosa, significantly. A major decline in the Colony Forming Units was observed with 3 log10 at 12 h of treatment. Also it is found to affect the membrane integrity of the pathogen, as 70.06 ± 2.009% cells were found to stain with Propidium iodide. In case of efflux activity 86.9 ± 2.08% cells were found in Ethidium bromide positive region. Biofilm formation inhibition ability was found in the range of 68.13 ± 4.11% to 84.86 ± 2.02%. Z.officinale is effective for killing Multi-Drug Resistant P. aeruginosa clinical isolate by affecting the cellular physiology and inhibiting the biofilm formation.

Protective effect of Berberis aristata against peritonitis induced by carbapenem-resistant Escherichia coli in a mammalian model.

OBJECTIVES: Berberis aristata is known to contain a variety of phenolic compounds contributing to its holistic capability of mitigating bacterial multidrug resistance. METHODS: B. aristata stem bark extract was prepared and was characterised using liquid chromatography-mass spectrometry (LC-MS). The antimicrobial efficacy of the extract against carbapenem-resistant Escherichia coli was assessed in vivo in an animal model using Sprague Dawley rats. Microbial counts in blood and urine, physical health status, haematological and biochemical analysis of blood, and histopathology of the kidney were assessed as the study endpoints. RESULTS: An aquo-alcoholic extract of B. aristata (PTRC-2111-A) was found to effectively manage peritonitis induced by carbapenem-resistant E. coli in a rat model at a single post-exposure prophylactic dose of 0.5mg/kg body weight (BW). The extract was also found to show a no observed adverse effect level (NOAEL) up to a dose of 2000mg/kg BW. Physical, immunological, haematological, biochemical and histopathological aberrations were found to be restored to normal in the herbal-treated group at a dose of 0.5mg/kg BW. CONCLUSIONS: The antimicrobial and hepatorenal protective ability of PTRC-2111-A could be attributed to the presence of isoquinoline alkaloids.

Effect of aquo-alchoholic extract of Glycyrrhiza glabra against Pseudomonas aeruginosa in Mice Lung Infection Model.

The prevalence of lung infection caused by Pseudomonas aeruginosa strains that are classified as multi-drug resistant has increased considerably and is mainly attributed to relative insufficiency of potent chemotherapeutic modalities. The present study was conducted to evaluate the antimicrobial activity of aquo-alcoholic extract of Glycyrrhiza glabra against the P. aeruginosa causing lung infection in Swiss albino mice. The study involves evaluation of lethal dose of P. aeruginosa in Swiss albino mice and analysis of disease manifestation that includes bacteremia, hypothermia, reduction in body weight and other parameters for 48h of infection. Physical manifestations of infected mice showed a significant decline in body temperature that is 29±0.57°C (at 48th h) from 38.81±0.33°C (0h) and 30% weight loss was observed at the end of the study. Further the efficacy of G. glabra extract against lung infection induced with the calculated lethal dose was evaluated by employing bacteremia, histopathology and radiological analysis. Bacterial burden showed that 2.30±0.02 Log10CFU/mL at day 7, a significant decline in the bacterial load as compared to day 1 when the bacterial burden was found to be 3.32±0.1 Log10CFU/mL. Histopathological results showed more diffuse and patchy accumulation of inflammatory cells within the alveolar space also the infiltrates were noted in all the lung section of infected mice. In treated animal group improved lung histology was seen with the exudates were less seen in D1 dose (20mg/kg) and disappeared in D2 dose (80mg/kg). The study clearly declares that the G. glabra extract is effective against lung infection caused by P. aeruginosa at dose of 80mg/kg. The LCMS results revealed that the extract contains Glycyrrhizin, Stigmasterol and Ergosterol, Licochalcone and Glabridin. The current study expected to further exploit the biomedical properties of this extract in the preparation of a potent regimen against such threatening pathogen.

Pharmacological evaluation of aqueous extract of syzigium cumini for its antihyperglycemic and antidyslipidemic properties in diabetic rats fed a high cholesterol diet-Role of PPARγ and PPARα.

In India syzygium cumini (Myrtaceae) is commonly used traditional medicine to treat diabetes. The present study was undertaken to assess an investigation of antihyperglycemic and antidyslipidemic properties of aqueous extract of Syzigium Cumini (SC) in diabetic rats fed a high cholesterol diet. The aqueous extract of SC was screened for antihyperglycemic and antidyslipidemic activity by streptozotocin induced diabetes in rats. Animals were treated with 100, 200 and 400mg/kg body weight of aqueous extract of SC. Metformin were used as reference antihyperglycemic drugs for comparison. Administration of aqueous extract of SC or metformin for 21days resulted in a significant (P<0.05) reduction in serum glucose, insulin and Homeostasis model assessment of insulin resistance (HOMA-IR) compared with diabetic controls. Treatment with 100mg/kg/day aqueous extract of SC did not result in a significant reduction in serum insulin levels, but 200mg/kg/day and 400mg/kg/day, aqueous extract of SC and metformin showed significant reductions 17.89%, 19.60% and 24.40%, respectively. Furthermore, administration of 100, 200 and 400mg/kg/day, aqueous extract of SC and metformin resulted in significant decrease in insulin resistance of 19.20%, 41.59%, 51.55% and 68.68%, respectively. In high fat diet- streptozotocin (HFD - STZ) treated rats ß-cells function (HOMA-B) were markedly reduced (5.8-fold), however observed a significant (P<0.01) improvement of ß-cell function with aqueous extract of SC (400mg/kg/day) and metformin. The aqueous extract of SC seeds exhibits significant insulin-sensitizing, antidyslipidemic, antioxidant, anti-inflammatory and ß-cell salvaging activity in HFD-STZ-induced type 2 diabetic rats via overexpression of PPARγ and PPARα activity, affirming its potential to be used in the prevention and treatment of type 2 diabetes mellitus (T2DM). Further isolation and characterization of active components in SC seed extract are needed to explore the other possible mechanisms and pathways that are involved in its anti-diabetic effect.

Intravaginal Delivery of Polyphenon 60 and Curcumin Nanoemulsion Gel.

Polyphenon 60 (P60) and curcumin (CUR) were loaded in a single nanoemulsion system and their combined antibacterial action was studied against uropathogenic Escherichia coli. To enhance availability at target organs and to inhibit enzymatic degradation in gastro intestinal tract, vaginal route of administration was explored. P60 + CUR nanoemulsion (NE) was formulated by ultra-sonication and optimized using Box-Behnken design. Optimized NE showed Z-average of 211.2 nm, polydispersity index of 0.343, and zeta potential of -32.7 mV. Optimized P60+ CUR NE was characterized by stability testing and transmission electron microscopy, and it was observed that NE was stable at 4°C for 30 days and monodisperse in nature with particle size of 195-205 nm. P60+ CUR NE was further formulated as gel and characterized by viscosity, growth curve analysis, and in vitro permeation studies. In vitro drug permeation studies in simulated vaginal media showed maximum permeation (84 ± 0.21%) of curcumin within 5 h and (91 ± 0.16%) of P60 within 8 h. Both the drugs maintained sustained permeation for 12 h. To investigate the transport via intravaginal route, gamma scintigraphy and biodistribution study of P60 + CUR NBG was performed on Sprague-Dawley rats using 99mtechnetium pertechnetate for radiolabeling to P60 molecule. Following intravaginal administration, P60 + CUR NBG dispersed in the kidney and urinary bladder with (3.07 ± 0.15) and (3.35 ± 0.45) percentage per gram after 3 h for P60 and CUR, respectively, and remained active for 12 h. Scintigraphy images suggested that the P60 + CUR NBG given by intravaginal route led to effective distribution of actives in urinary tract, and this observation was in agreement with the biodistribution results.

Augmenting the potency of third-line antibiotics with Berberis aristata: In vitro synergistic activity against carbapenem-resistant Escherichia coli.

The aim of this study was to analyse the in vitro synergistic antibacterial potential of an aquoethanolic extract of the stem bark of Berberis aristata (PTRC-2111-A) with third-line antibiotics against carbapenem-resistant Escherichia coli. PTRC-2111-A was prepared and was characterised using phytochemical- and bioactivity-based fingerprinting. Fourier transform infrared spectroscopy (FTIR) and liquid chromatography-mass spectrometry (LC-MS) analyses were performed, and superoxide and hydroxyl scavenging activities were assessed in conjunction with in vitro antimicrobial efficacy testing against the test micro-organism. Analysis of drug combinations of PTRC-2111-A and third-line antibiotics was performed using CompuSyn software. PTRC-2111-A from B. aristata was found to have seven common functional groups in comparison with the pre-identified marker compound quercetin, and phytochemical quantitation analysis revealed the presence of 25.44% alkaloids. Moreover, PTRC-2111-A was found to contain isoquinoline alkaloids, namely berbamine, berberine, reticuline, jatrorrhizine, palmatine and piperazine, as elucidated in the LC-MS analysis. Analysis of combinations of PTRC -2111-A and antibiotics revealed synergistic behaviour [fractional inhibitory concentration index (FICI)<1] with colistin, tigecycline and amoxicillin/clavulanate potassium (Augmentin(®)), whereas antagonism (FICI>1) was seen with ertapenem and meropenem.

Alternative to antibiotics against Pseudomonas aeruginosa: Effects of Glycyrrhiza glabra on membrane permeability and inhibition of efflux activity and biofilm formation in Pseudomonas aeruginosa and its in vitro time-kill activity.

The multi-drug resistance offered by Pseudomonas aeruginosa to antibiotics can be attributed towards its propensity to develop biofilm, modification in cell membrane and to efflux antibacterial drugs. The present study explored the activity of Glycyrrhiza glabra and one of its pure compounds, glycyrrhizic acid against P. aeruginosa and their mechanism of action in terms of the effect on membrane permeability, efflux activity, and biofilm formation were determined. Minimum inhibitory concentrations were determined by using broth dilution technique. The minimum bactericidal concentrations were assessed on agar plate. The MIC of the extract and glycyrrhizic acid was found to be 200 and 100 µg ml(-1), respectively. The MBC was found to be 800 and 400 µg ml(-1) in the case of extract and glycyrrhizic acid, respectively. Time -dependent killing efficacy was also estimated. Flowcytometric analysis with staining methods was used to determine the effect of extract and glycyrrhizic acid at 2 × MIC on different physiological parameters and compared it with the standard (antibiotic). The growth of P. aeruginosa was significantly inhibited by extract and the pure compound. The herbal extract and the glycyrrhic acid were also found to effective in targeting the physiological parameters of the bacteria that involve cell membrane permeabilization, efflux activity, and biofilm formation. This study reports the antipseudomonal action of Glycyrrhiza glabra and one of its compound and provides insight into their mode of action.

In vitro bactericidal activity of promising nutraceuticals for targeting multidrug resistant Pseudomonas aeruginosa.

OBJECTIVE: We evaluated the bactericidal activity of nutraceuticals against multidrug resistant Pseudomonas aeruginosa. The nutritionally valued herbs were screened on the basis of a matrix modeling approach and molecular docking based validation analysis. METHODS: The database of 38 herbs developed earlier using fuzzy logic based scoring analysis was subjected to molecular docking based validation. The molecular docking (Hex 6.12) analyses of predominant phytoligands (∼10 per herb) against exoenzyme S of P. aeruginosa filtered potent herbs were selected. The preauthenticated bacterial inoculum (10(8) CFU/mL) was added to the sterile nutrient broth impregnated with standardized aqueous-alcoholic herbal extracts (1-1600 µg/mL). After overnight incubation at 37°C, antibacterial activity was evaluated in terms of minimum inhibitory and minimum bactericidal concentrations. RESULTS: Five herbs were selected on the basis of fuzzy set scoring, an herbal informatics model, and validation analysis based on energy of docking (i.e., Evalue of 380) phytoligands with maximum scoring obtained by Glycyrrhiza glabra. Among the 5 nutraceuticals, G. glabra showed maximum bactericidal activity significantly (P < 0.05) higher than Amikacin, a standard antibiotic, which was in consonance with in silico bioprospection. Zingiber officinale, despite a low Evalue, showed considerably higher inhibition attributed to its higher flavonoid content as compared to other herbs. CONCLUSION: G. glabra (licorice), a flavoring agent; Z. officinale (ginger), a condiment; and Mentha piperita (mint), a fragrance component, showed significant therapeutic potential against multidrug resistant strains of P. aeruginosa.

In vitro bactericidal activity of Berberis aristata extract against clinical isolates of carbapenem-resistant Escherichia coli.

BACKGROUND: Berberis aristata is known to contain a variety of phenolic compounds, flavonoids such as quercetin attributing towards its holistic capability of mitigating multidrug resistance. METHODS: B. aristata stem bark extract was prepared and characterized using phytochemical and bioactivity-based fingerprinting. Anti-oxidant and anti-lipid peroxidation profiling was also done in conjunction with in vitro anti-microbial efficacy testing against the test microorganism i. e., New Delhi Metallo-ß-lactamase-1 (NDM-1) Escherichia coli. RESULTS: Aquo-alcoholic (1:1) extract of B. aristata (PTRC-2111-A), containing 3.0±0.02 µg of QUERCETIN/mg of dried extract, exhibited [flavonoid/polyphenol: F/P (quercetin %) ~ 0.16(0.06 %)]. The bioactivity fingerprint profile of PTRC-2111-A included IC50 ratio [DPPH/NOS]=0.064 as functional standardized value having IC50 (DPPH Scavenging)=16±0.5 µg/mL and IC50 (Nitric Oxide Scavenging)=250±0.5 µg/mL respectively. The reducing ability and anti-lipid peroxidation equivalent (extract: standard) of PTRC-2111-A with respect to standard was estimated to be 3.44 (ascorbic acid) and 0.78 (quercetin) respectively. In vitro anti-microbial activity evaluated against sts-09 multidrug-resistant strain of carbapenem-resistant E. coli was found to be 25 µg/mL. CONCLUSIONS: B. aristata was found to contain a number of phytoconstituents, which acts in a synergistic manner to provide significant bactericidal potential against carbapenem-resistant E. coli.

Assessment of aquo-ethanolic extract of Camellia sinensis against Carbapenem Resistant Escherichia coli: In Vivo Trials in a Murine Model.

The prevalence of Carbapenem Resistant Escherichia coli (CRE) has increased considerably during the last decade, which can be ascribed to relative scarcity of effective non toxic antimicrobial agents. The present study was conducted to evaluate the antimicrobial activity of aquo-ethanolic (1:1) extract of leaves of Camellia sinensis (PTRC-31911-A) against Carbapenem Resistant Escherichia coli at preclinical level using peritonitis infection model in Sprague Dawley rats. Efficacy analysis of PTRC-31911-A involved enumeration of CRE colonies in blood and urine samples of test animals for a period of 5 days from infection. A reduction in microbial count of biological fluids was considered as the primary endpoint of the selected murine model. Physical, biochemical, hematological and histological indices of toxicity were employed as secondary relative indicators of the induced disease. Physical manifestations of infected rats included significantly high body temperature (TempInfected=103.18°F, ∼5% increase) and noteworthy reduction in weight (WeightInfected=126.83g, ∼15% decrease) as compared to control. Significant (P<0.05) increase in total white blood cells, eosinophil and monocyte counts as well as a significant decrease (P<0.05) in erythrocytes count, hematocrit volume, red blood cell distribution width and hemoglobin concentration were observed in the infected group as compared to the control group. Furthermore, noteworthy increase in liver and kidney function test parameters were observed in case of infected groups. All the hematological and biochemical parameters were found to be within optimum range in case of treatment group, indicating restoration of homeostasis. Histopathological studies also presented symptoms of hemorrhage and glomerular damage with structural distortion in glomerular capillary loops of infected groups, which were later recovered in treated groups, indicating the nephro-protective potential of PTRC-31911-A. The study clearly points out that Camellia sinensis extract (PTRC-31911-A; single dose of 5mg/Kg bwt; oral,+24h) is highly effective against Carbapenem Resistant Escherichia coli owing mainly to the presence of flavonoids and polyphenolic compounds, identified by LCMS. Ongoing studies are expected to further unravel the mechanism of action and bioactivity determinants of this broad spectrum plant extract.

Anti-hemolytic, hemagglutination inhibition and bacterial membrane disruptive properties of selected herbal extracts attenuate virulence of Carbapenem Resistant Escherichia coli.

Expression of a multitude of virulence factors by multi-drug resistant microbial strains, e.g., Carbapenem Resistant Escherichia coli (Family: Enterobacteriaceae; Class: Gammaproteobacteria), is responsible for resistance against beta-lactam antibiotics. Hemolysin production and induction of hemagglutination by bacterial surface receptors inflicts direct cytotoxicity by destroying host phagocytic and epithelial cells. We have previously reported that Berberis aristata, Camellia sinensis, Cyperus rotundus Holarrhena antidysenterica and Andrographis paniculata are promising herbal leads for targeting Carbapenem resistant Escherichia coli. These herbal leads were analyzed for their anti-hemolytic potential by employing spectrophotometric assay of hemoglobin liberation. Anti-hemagglutination potential of the extracts was assessed by employing qualitative assay of visible RBC aggregate formation. Camellia sinensis (PTRC-31911-A) exhibited anti-hemolytic potential of 73.97 ± 0.03%, followed by Holarrhena antidysenterica (PTRC-8111-A) i.e., 68.32 ± 0.05%, Berberis aristata (PTRC-2111-A) i.e., 60.26 ± 0.05% and Cyperus rotundus (PTRC-31811-A) i.e., 53.76 ± 0.03%. Comprehensive, visual analysis of hemagglutination inhibition revealed that only Berberis aristata (PTRC-2111-A) and Camellia sinensis (PTRC-31911-A) exhibited anti-hemagglutination activity. However, Andrographis paniculata (PTRC-11611-A) exhibited none of the inhibitory activities. Furthermore, the pair wise correlation analysis of the tested activities with quantitative phytochemical descriptors revealed that an increased content of alkaloid; flavonoids; polyphenols, and decreased content of saponins supported both the activities. Additionally, flow cytometry revealed that cell membrane structures of CRE were damaged by extracts of Berberis aristata (PTRC-2111-A) and Camellia sinensis (PTRC-31911-A) at their respective Minimum Inhibitory Concentrations, thereby confirming noteworthy antibacterial potential of both these extracts targeting bacterial membrane; hemolysin and bacterial hemagglutination.

Attenuation of adhesion, quorum sensing and biofilm mediated virulence of carbapenem resistant Escherichia coli by selected natural plant products.

The multi-drug resistance offered by Carbapenem Resistant Escherichia coli (Family: Enterobacteriaceae; Class: Gammaproteobacteria) against third line antibiotics can be attributed towards its ability to develop biofilm. Such process involves adhesion and quorum-sensing induced colonization leading to biomass development. The present study explored the anti-adhesion, anti-quorum sensing and anti-biofilm potential of 05 pre-standardized potent herbals. Berberis aristata (PTRC-2111-A) exhibited maximum potential in all these activities i.e. 91.3% ± 0.05% (Anti-adhesion), 96.06% ± 0.05% (Anti-Quorum sensing) and 51.3% ± 0.07% (Anti-Biofilm formation) respectively. Camellia sinensis (PTRC-31911-A) showed both anti-adhesion (84.1% ± 0.03%) and anti-quorum sensing (90.0%) potential while Holarrhena antidysenterica (PTRC-8111-A) showed only anti-quorum sensing potential as compared to standards/antibiotics. These findings were in line with the molecular docking analysis of phytoligands against Lux S and Pilin receptors. Furthermore, the pairwise correlation analysis of the tested activities with qualitative, quantitative and bioactivity functional descriptors revealed that an increased content of alkaloid, moderate content of flavonoids and decreased content of tannins supported all the three activities. In addition, nitric oxide and superoxide scavenging activity were found to be correlated with anti-quorum sensing activity. The findings indicated clearly that B. aristata (Family: Berberidaceae) and C. sinensis (Family: Theaceae) were potent herbal leads with significant therapeutic potential which further needs to be explored at pre-clinical level in the future.