Screening of the antioxidant and antibacterial effects of extracted essential oils from Thunbergia coccinea, Acacia polyacantha, Polygonum micrpcephallum, Abies spectabilis and Clerodendrum colebrookianum

During the previous few decades, it has been seen that there is a rapid emergence of pathogens resistant to multiple antibiotics. This has now become a global crisis. Some unexplored or less explored plants also provide some antibacterial, bactericidal and antioxidant properties. The antibacterial, bactericidal effects of extracted essential oils (EEOs) of Thunbergia coccinea, Acacia polyacantha, Polygonum micrpcephallum, Abies spectabilis and Clerodendrum colebrookianum was tested in comparison with standard antibiotics. The methods chosen were disc diffusion and deduction of minimum inhibitory concentration (MIC) by microbroth dilution assays of the EEOs against the bacterial strains.The antioxidant activity was found out utilizing DPPH free radical scavenging assay, MDA, Hydrogen peroxide radical inhibition assay and Superoxide radical inhibition assay (O 2 -). Some commonly used standard antibiotics (metronidazole, amoxicillin, clarithromycin, rifampicin, clindamycin and oxacillin,) were utilized to compare the EEO antibacterial action. Clerodendrum colebrookianum (85.17 ± 3.06 µg MDA/g extract) had a reasonable MDA. Acacia polyacantha in MIC had values of 3.86 ± 0.25 to 6.20 ± 0.16. Polygonum micrpcephallum had excessive H2O2 (48.27 ± 2.4 5%). The antibacterial actions determined by the paper disc‑diffusion technique of the EEO extracted from these plants showed that most had some antibacterial actions. Also, it was seen that the bactericidal action of the EEO extracted from E. alba was most potent against S. pyogenes (4.06 ± 0.15). The extract of the plant at varying concentrations (20, 40, 60, 80 and100 mg/mL) demonstrated noteworthy (P< 0.001) anthelmintic action in an effective change when the dose was adjusted. In conclusion, most of the tested plants contain a medicinal value, which can be utilized in the future to supplement artificial medicines and cure emerging diseases that create havoc for mankind.

Phytochemical analysis and investigation of Antimicrobial and Antioxidant potential of the Leaf Extracts of Putranjiva ruxburghi

Objectives: Herbs are in use as medicine worldwide from the time immemorial. Many drugs derived from plant, animal or minerals are in use as medicine till date. This is a study on leaf extract of Putranjiva roxburghii Wall. (Euphorbiaceae) involving pharmacognosy, phytochemistry, activity against microbes and oxidation to substantiate its use. The leaf extracts were collected by successive soxhlation using solvents like petroleum ether, chloroform, benzene, acetone & ethanol. Chemicals present in the crude leaf extracts and isolated constituents were analyzed for both quality and quantity followed by TLC, UV–Visible Spectrophotometric analysis, HPTLC, Phytochemical tests and TLC examination confirmed to contain flavonoids in acetone, chloroform, aqueous and ethanol extract. The activity of Petroleum-ether, chloroform, acetone, alcoholic and aqueous extracts against the microbes was assessed by cup plate method. Okada & Okada method was followed to evaluate the DPPH free radical for aging property. The reducing, total antioxidant and peroxide radical for aging action of the extracts were assessed. The study confirmed significant antimicrobial and antioxidant property of the Putranjiva roxburghii Wall leaves may be for the presence of constituents like flavonoids, saponins, Phytosterols, favoring its traditional usage as medicine.

An Anti-radical, Cytoprotective, Anti-proliferative, Anti-inflammatory and Toxicological Assessment of Metallo-porphyrins Isolated from Fresh Leaves of Spinacia oleracea L.

Aim: Synthetic lead molecules are associated with host of adverse effects while medicinal molecules isolated from natural sources are blessed with both safety as well as efficacy. The ancient doctrine of Ayurveda ardently advocates the therapeutic virtues contained in green leaves of Spinacia oleracea L. The principal constituent of the leaves is the class of metalloporphyrin chlorophyll, which is also the floral counterpart of faunal heme. Chlorophyll-a (Chl-a) and chlorophyll-b (Chl-b) are the cardinal members of the chlorophyll family. Study design: Herein, we have explored the anti-radical, cytoprotective, anti-inflammatory and anti-proliferative efficacy of Chl-a and Chl-b in reference to standard drug and crude extract of Spinacia leaves. The current study is aimed to establish, naturally mined metaloporphyrins as safe and efficacious replacement of synthetic leads that are associated with a wide range of toxicological issues. Methodology: Using a combination of Silica Gel-G column chromatography and preparative thin layer chromatography, the two principal green metallo-porphyrins (Chl-a and Chl-b) were sequentially extracted and isolated from crude extract of Spinacia oleracea L leaves. Antiradical efficacy, of the isolated green porphyrins was quantified by DPPH and Hydrogen peroxide radical scavenging assay. Cytoprotective efficacy was evaluated using ex-vivo hemolysis assay and anti-inflammatory potency was attested employing carrageenan induced paw edema bioassay. To enumerate on the anti-proliferative potency, MTT assay was employed, while toxicology of the isolates was evaluated employing OECD 420 acute toxicity guidelines. Findings: The study confirmed that isolated green porphyrins Chl-a and Chl-b as well as crude extract all exerts significant anti-radical, cytoprotective, anti-inflammatory and anti-proliferative efficacy however while potency of Chl-a was at par with that of reference standard and superior to the crude extract, Chl-b clocked in a value inferior to both. Furthermore, acute toxicity study indicated that even at p.o. dose of 2000mg/Kg b.w, no toxicity was manifested in either of the metalloporpyrin treated groups thus ascertaining the safe nature of the naturally mined metalloporphyrin entities. Also naturally mined Chl-a is not only a safer alternative to synthetic medicine but it is more potent and safe than its parent extract popularly used in herbal medicine. Conclusion: The results of the study indicates that Chl-a having a more profound structural resemblance to heme than Chl-b can be further modulated as a cost-effective and safe anti-radical alternative to synthetic leads in inhibiting inflammation and untoward cell proliferative while extending cyto-protection from pathological ROS generated in diseased states.

In vitro Antioxidant Potential of Abrus precatorius L. and Asystasia gangetica (L.) T. Anderson

Antioxidant substances are an important part of human life as it plays a key role in nutraceuticals and also help to defend free radicals present in our body. The present study highlights the need to find potent natural antioxidants from medicinal plants. Different extracts of Abrus precatorius L. (Fabaceae) and Asystasia gangetica (L.) T. Anderson (Acanthaceae) were successively prepared using petroleum ether, benzene, chloroform, and ethanol. It was evaluated for antioxidant activities using various assays. Maximum extractable total phenolics and flavonoids were recorded in varied extracts of A. precatorius and A. gangetica. The extracts also showed efficient phosphomolybdenum reduction, reducing power activity, nitric oxide, and hydrogen peroxide radical scavenging properties. It is very clear from the results that the studied plants A. precatorius and A. gangetica have remarkable medicinal uses with extraordinary potential for pharmaceuticals. Further detailed studies will pave the way to promote natural drugs for health benefits.

In vitro antioxidant potential and antimicrobial activity of leaves and stem extracts of Anogeissus pendula Edgew.

Anogeissus pendula Edgew. is commonly used in the conventional Indian medicinal system and is reported to contain phenolic compounds which have antioxidant and antimicrobial potential. The goal of our study is to look at the antioxidant function and antibacterial activity of A. pendula leaf and stem extracts. The total phenolic content (TPC), total flavonoid content (TFC) and total tannin content (TTC) were determined using a spectrophotometric technique (TTC). In vitro techniques such as 1, 1-diphenyl-2-picrylhydrazyl (DPPH), hydrogen peroxide radical scavenging tests (H2O2) and Ferric reducing antioxidant power (FRAP) assay were used in the study. The disc diffusion technique was used to assess antibacterial activity and the minimum inhibitory concentration (MIC) was investigated against four bacterial strains. The TTC of leaf and stem methanol extract was considerably higher which ranged from 15.07 ± 0.506 to 38.77 ± 1.253 mg gallic acid equivalent (GAE) /g in leaves and 19.83 ± 0.084 to 28.56 ± 0.437 mg GAE/g in the stem. The content of flavonoid in the leaf and stem methanol extract varied from 12.53 ± 0.603 to 37.28 ± 0.466 mg rutin equivalent (RE) /g in leaves and 10.01 ± 0.177 to 37.28 ± 0.466 mg RE/g in stems. Hydroalcoholic extract of leaf and stem showed the highest tannin content and ranged from 23.73 ± 0.091 to 34.08 ± 0.261 mg tannic acid equivalent (TAE) /g. In order of efficacy (IC50) of the plant extracts, the effective inhibitor was the methanol extract of leaf and stem in the DPPH and H2O2 assay. FRAP value was higher in the hydroalcoholic extract of both leaf and stem. Antimicrobial activity tests revealed that all extracts limit the development of diverse microbial strains such as Escherichia coli, Bacillus subtilis, Pseudomonas putida and Streptococcus aureus with a mean zone of inhibition ranging from 0 to 15.67 mm. The MIC of A. pendula leaf and stem solvent extracts against bacterial strains ranged from 0.195 to 50 mg/ml. The findings revealed that A. pendula has a variety of phytochemicals with substantial antioxidant and antibacterial properties, confirming its usage in traditional medicine.

A proposed enzyme-linked hydrogen peroxide radical assay system (HORAS)

Reactive Oxygen Species (ROS) are chemically reactive molecules that contain oxygen. ROS are naturally generated as a byproduct during mitochondrial oxidative metabolism as well as by cellular responses to a variety of inflammatory stimuli. Intracellularly formed ROS plays an important role in maintaining homeostasis and in cell signaling but, ROS are challenging to quantify. Phagocytic cells such as macrophages may produce H2O2 during the action of bacterial engulfment. Here UV-Vis versus LC-ESI-MS detection methods for an enzyme-linked, cellular assay of H2O2 production in cultured macrophages are compared. In the presence of Horseradish Peroxidase (HRP), Amplex Red (AR) reacts with H2O2 in a 1:1 stoichiometry to produce the red-fluorescent oxidation product resorufin that can be measured by UV/Vis at an absorbance of 570 nm or by LC-ESI-MS at 214 m/z [M+H]+. RAW 264.7 macrophages were stimulated by microscopic foreign particles, with the addition of 0.1mM of Amplex Red substrate and 10 ng/mL of HRP to the cellular media to enzymatically detect H2O2 production. The oxidation product resorufin can be detected by the colorimetric method as low as 50 pmol while liquid chromatography with electrospray ionization and mass spectrometry (LC-ESI-MS) was able to detect as little as 0.2 pmol in vitro. Thus, it was possible to measure low levels of H2O2 released by cells using an enzyme coupled cellular assay with LC-ESI-MS.

A proposed enzyme-linked hydrogen peroxide radical assay system (HORAS)

Reactive Oxygen Species (ROS) are chemically reactive molecules that contain oxygen. ROS are naturally generated as a byproduct during mitochondrial oxidative metabolism as well as by cellular responses to a variety of inflammatory stimuli. Intracellularly formed ROS plays an important role in maintaining homeostasis and in cell signaling but, ROS are challenging to quantify. Phagocytic cells such as macrophages may produce H2O2 during the action of bacterial engulfment. Here UV-Vis versus LC-ESI-MS detection methods for an enzyme-linked, cellular assay of H2O2 production in cultured macrophages are compared. In the presence of Horseradish Peroxidase (HRP), Amplex Red (AR) reacts with H2O2 in a 1:1 stoichiometry to produce the red-fluorescent oxidation product resorufin that can be measured by UV/Vis at an absorbance of 570 nm or by LC-ESI-MS at 214 m/z [M+H]+. RAW 264.7 macrophages were stimulated by microscopic foreign particles, with the addition of 0.1mM of Amplex Red substrate and 10 ng/mL of HRP to the cellular media to enzymatically detect H2O2 production. The oxidation product resorufin can be detected by the colorimetric method as low as 50 pmol while liquid chromatography with electrospray ionization and mass spectrometry (LC-ESI-MS) was able to detect as little as 0.2 pmol in vitro. Thus, it was possible to measure low levels of H2O2 released by cells using an enzyme coupled cellular assay with LC-ESI-MS.

Pharmacodynamic findings for the usefulness of Luffa cylindrica (L.) leaves in atherosclerosis therapy with supporting antioxidant potential

Background Luffa cylindrica (L.) is a commonly used vegetable in different parts of Asia. Its fruits are generally used as a vegetable, but pharmacological activities of the leaves were unrevealed. The study evaluated the antihyperlipidemic activity and in vitro antioxidant potential of methanolic extract of Luffa cylindrica (L.) leaves (MELCL). The antihyperlipidemic potential was investigated in Triton X-100-induced hyperlipidemic rats. Animals were pre-treated with Triton X-100 (400 mg/kg). The Triton X-100-treated animals were then treated with MELCL at the doses of 100 and 200 mg/kg using 5% CMC, as a vehicle, per oral (p.o) for 7 days. Antioxidant activity was studied by examining the DPPH and hydrogen peroxide radical scavenging potential of the extract. Results The plasma sample of rats was analyzed, and it was found that MELCL shows significant (p < 0.05) antihyperlipidemic activity at 200 mg/kg of MELCL. Serum analysis showed a marked reduction in the level of multiple biochemicals like total cholesterol (TC) (85.48 ± 3.230 mg/dl), triglycerides (TG) (74.62 ± 8.764 mg/dl), low-density lipoproteins (LDL) (31.97 ± 3.475 mg/dl), very low-density lipoproteins (VLDL) (14.92 ± 1.635 mg/dl), and an increase in the level of high-density lipoproteins (HDL) (40.58 ± 1.625 mg/dl). MELCL also showed significant scavenging of DPPH radical (46.66 ± 0.002%) at concentration and hydrogen peroxide radical (47.55 ± 0.001%) at 100 μg/ml. Conclusion Quantitative results of the study showed that MELCL has considerable antihyperlipidemic and antioxidant potential and could be the option for the treatment of atherosclerosis.

Formulation and Antioxidant Activity Evaluation of Spondias mombin - Abelmoschus esculentus Mucilage Emulsion

Plant gums and mucilage have wide applications in pharmaceutical formulations as emulsifying agents. But these are quite costly, therefore there is always the need for cheaper alternatives. In this study, Okra mucilage from Abelmoschus esculentus (L.) Moench pods was used to formulate Spondias mombin L. leaf extract emulsion. The emulsion was assessed for the effect of the mucilage on antioxidant activity of the extract using four standard in vitro assays. Phytochemical screenings of the mucilage as well as physico-chemical and microscopic characterization of the emulsion were also carried out. The mucilage contained carbohydrates, reducing sugars, terpenoids and unsaturated lactones. The pH of the resulting emulsion was 6.88. The size of the oil globules were small with the average size of 4. 8μm. The extract had superior 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, hydrogen peroxide radical scavenging and ferric ion reducing capacity compared to the emulsion (p ≤ 0.05). In addition, the emulsion had a better metal chelating activity compared to the extract (p ≤ 0.05). Okra mucilage may have reduced the antioxidant activity of the extract suggesting its incongruity in this emulsion formulation. Bangladesh Pharmaceutical Journal 24(1): 17-25, 2021