Clinical presentation and management of methanol poisoning outbreaks in Riyadh, Saudi Arabia: a retrospective analysis.

BACKGROUND: Acute methanol intoxication, whether unintentional or deliberate, necessitates prompt intervention to prevent severe morbidity and mortality. Homemade alcoholic beverages are a frequent source of such poisoning. This retrospective analysis examined two outbreaks of methanol intoxication in Saudi Arabia. It investigated the clinical presentation, implemented management strategies, and any lasting complications (sequelae) associated with these cases. The aim was to assess the potential impact of different treatment modalities and the timeliness of their initiation on patient outcomes. METHODS: This was a retrospective case series of methanol poisoning cases which presented to the adult emergency department (ED) at King Abdulaziz Medical City (KAMC) in Riyadh, Saudi Arabia. There were two separate outbreaks in the city, the first one was from September 1 to September 10, 2020 and the second one was from May 14 to May 20, 2021. Electronic charts were reviewed, and data were extracted to previously prepared data extraction sheets. RESULT: From the 22 patients who arrived in the ED alive, the most common complaints were nausea or vomiting followed by altered level of consciousness. About 9% from the patient were hypotensive, 36% were tachycardic, 41% were tachypneic and 4% were having SpO2 < 94%. Brain CT was abnormal in 6 patients. Vision impairment was the most common sequalae of methanol poisoning (7 out of 12 patients who were assessed by ophthalmologist, 58%). When the patients were divided based on severity (mild, moderate, severe), nausea or vomiting and loss of consciousness were the most common complaints among the moderate group while loss of consciousness predominated in the severe group. Two patients presented with low blood pressure and were in the sever group. The severe group had a mean Glasgow Coma Scale (GCS) of 8. Most of the patients in the severity groups underwent the same management apart from those who died or deposited. Eight patients in the severe group had to be intubated. CONCLUSION: This study demonstrates the multifaceted clinical presentation of methanol poisoning, culminating in a 17.4% mortality rate. Notably, our findings emphasize the critical role of prompt diagnosis and swift initiation of combined fomepizole therapy and hemodialysis in mitigating mortality and minimizing the potential for chronic visual sequelae associated with methanol poisoning.

Nephroprotective and Anti-Diabetic Potential of Beta vulgaris L. Root (Beetroot) Methanolic Extract in a Rat Model of Type 2 Diabetes Mellitus.

Background and Objectives: Diabetes mellitus is a chronic metabolic disease associated with several complications, including that of kidney disease. Plant-based dietary products have shown promise in mitigating these effects to improve kidney function and prevent tissue damage. This study assessed the possible favorable effects of beetroot extract (BE) in improving kidney function and preventing tissue damage in diabetic rats. Materials and Methods: Type 2 diabetes mellitus (T2DM) was induced using a low dose of streptozotocin (STZ). Both control and rats with pre-established T2DM were divided into six groups (each consisting of eight rats). All treatments were given by gavage and continued for 12 weeks. Fasting blood glucose levels, serum fasting insulin levels, Homeostatic Model Assessment for Insulin Resistance (HOMA-IR), serum triglycerides, cholesterol, low-density lipoprotein-cholesterol, serum and urinary albumin, and creatinine and urea levels were measured. Apart from this, glutathione, malondialdehyde, superoxide dismutase, tumor necrosis factor-α, and interleukine-6 in the kidney homogenates of all groups of rats were measured, and the histopathological evaluation of the kidney was also performed. Results: It was observed that treatment with BE increased body weight significantly (p ≤ 0.05) to be similar to that of control groups. Fasting glucose, insulin, HOMA-IR levels, and lipid profile in the plasma of the pre-established T2DM rats groups decreased to p ≤ 0.05 in the BE-treated rats as the BE concentration increased. Treatment with BE also improved the renal levels of oxidative stress and inflammatory markers, urinary albumin, and serum creatinine and urea levels. Unlike all other groups, only the kidney tissues of the T2DM + BE (500 mg/kg) rats group showed normal kidney tissue structure, which appears to be similar to those found in the kidney tissues of the control rats groups. Conclusion: we found that streptozotocin administration disturbed markers of kidney dysfunction. However, Beta vulgaris L. root extract reversed these changes through antioxidant, anti-inflammatory, and antiapoptotic mechanisms.

Evaluation of the antimalarial and CD4+ T-cell modulatory effects of leaf methanol extract of Phyllanthus muellerianus (Kuntze) Exell (Phyllanthaceae) in Plasmodium berghei-infected mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Phyllanthus muellerianus (Kunze) Exell, a member of the Phyllanthaceae family, is a medicinal plant widely distributed in Africa. Decoctions from the leaves are used in Nigeria to treat fevers, convulsions, some neurological disorders and malaria. AIM OF THE STUDY: This study is to evaluate the anti-malarial properties of methanol extract of Phyllanthus muellerianus (MEPM) leaves and its ethyl acetate fraction using a murine malaria model infected with Plasmodium berghei. Additionally, we seek to investigate the potential modulatory effects of this extract and fraction on CD4+ T-cell populations in the context of malaria infection. MATERIALS AND METHODS: The anti-malarial effects of the leaf methanol extract of Phyllanthus muellerianus (MEPM) were screened using three established in vivo models of anti-plasmodial screening namely the curative, suppressive and prophylactic models. The methanol extract (MEPM) was afterwards fractionated into hexane (HFPM), ethyl acetate (EAFPM), and methanol (MFPM) fractions. In the pilot anti-malarial screening of the fractions, EAFPM exhibited the best antiparasitic activity. Subsequently, EAFPM was screened for anti-malarial activity using the three models above. The effects of the MEPM and EAFPM on haematological indices (Hb and PCV) of the inoculated animals were further screened and the mean survival time (MST) of the animals was monitored. CD4+ T cells of various groups were counted before and after treatment using a flow cytometer. The EAFPM was further subjected to HPLC analysis for identification of its major compounds. RESULTS: The EAFPM (100 and 200 mg/kg) elicited 88% and 93% cure respectively in the curative model, while artesunate (5 mg/kg,- the positive control) gave 87% protection. The MEPM and EAFPM also gave significant suppression of parasitemia in the suppressive model. The treated groups survived beyond 28 days as against 11 days by the control group (infected but not treated). The treated groups also prevented anaemia seen in the negative control. The EAFPM group significantly modulated the CD4+ T cell. Compounds identified were Gallocatechin, Quercetin -3-O-gallate, Ellagic acid, and Methylellagic acid rhamnoside). CONCLUSION: The study established that the leaf of Phyllanthus muellerianus possesses antimalarial activity, thus lending support to its use in the folkloric treatment of malaria.

Phytochemical characterization and phospholipase A2 inhibitory effect of Vitex negundo L. root extracts.

ETHNOPHARMACOLOGICAL RELEVANCE: Snake bites are a critical health issue in many parts of the world particularly in Asian countries lacking efficient health facilities in rural areas. Cobra is the most common snake type in Asia and is responsible for a large number of mortalities particularly in rural areas. Plants are usually considered the most effective and easy-to-approach treatment for snake bites in rural areas of various countries. Vitex negundo L. is an important medicinal plant traditionally used to treat snake bite envenomation in many countries of Asia. AIM OF THE STUDY: From literature survey of plants traditionally used in the treatment of snake bites in Asian countries including India, Pakistan and Sri Lanka, roots of V. negundo were selected for the present study. Anti-snake venom potential of its roots was assessed through various in vitro assays targeting the phospholipase A2 enzyme. MATERIALS AND METHODS: V. negundo roots were sequentially extracted in different organic solvents to get fractions and in methanol to get total extract. The extracts were evaluated for phospholipase A2 (PLA2) inhibitory potential through inhibition of venom-induced hemolysis, ADP-induced platelet aggregation, PLA2-induced fatty acid hydrolysis and anticoagulant effect of cobra venom. Antioxidant power was determined using DPPH and superoxide radical scavenging assays. GC-MS and HPLC analysis was performed for the total methanol extract. RESULTS: Strong PLA2 inhibitory effect was observed for all the extracts. The ethyl acetate, acetone and methanol fractions significantly inhibited toxic effects of cobra venom under in vitro conditions. Radical scavenging potential of these fractions was also significantly high as compared to non-polar fractions in both DPPH and superoxide scavenging assays. Phytochemical analysis indicated high phenolic and flavonoid contents in these fractions. GC-MS and HPLC analysis of total methanol extract confirmed the presence of bis(2-ethylhexyl) phthalate, phenol, o-Guaiacol, palmitic acid-methyl ester, methyl stearate, quercetin and kaempferol in the plant. CONCLUSION: The study concluded that the roots of V. negundo, particularly their polar extracts, have strong PLA2 inhibitory effect against cobra venom confirming their traditional use to manage snake bites. The roots of this plant can be further studied for isolation of plant-based antisera.

Exploring the efficacy of ethnomedicinal plants of Himalayan region against the malaria parasite.

ETHNOPHARMACOLOGICAL RELEVANCE: Plasmodium falciparum multi-drug resistant (MDR) strains are a great challenge to global health care. This predicament implies the urgent need to discover novel antimalarial drugs candidate from alternative natural sources. The Himalaya constitute a rich repository of medicinal plants which have been used traditionally in the folklore medicine since ages and having no scientific evidence for their activity. Crambe kotschyana Boiss. and Eremurus himalaicus Baker are used for their antipyretic and hepatoprotective properties in Kinnaur district of Himachal Pradesh, India. AIM OF THE STUDY: This study would investigate the antiplasmodial efficacy of C. kotschyana and E. himalaicus extracts, their fractions and active components using in vitro, in vivo and in silico approaches to provide a scientific insight into their activity. METHODS: The methanol extracts of C. kotschyana (CKME) and E. himalaicus (EHME) were prepared by maceration followed by fractionation using ethyl acetate. The isolation of flavonoid glycosides isorhamnetin-3, 7-di-O-glucoside from C. kotschyana and luteolin-6-C-glucoside (isoorientin) from E. himalaicus was carried out by antiplasmodial activity-guided isolation. In vitro antimalarial activity was assessed by WHO method while in vitro cytotoxicity was ascertained employing the MTT assay. Molecular docking and molecular dynamics simulation were performed using the Glide module of Schrödinger Software and Gromacs-2022 software package respectively. In vivo curative activity was assessed by Ryley and Peters method. RESULTS: The methanol extracts of both the plants illustrated the best antiplasmodial activity followed by the ethyl acetate fractions. Iso-orientin (IC50 6.49 µg/ml) and Isorhamnetin-3,7-di-O-glucoside (IC50 9.22 µg/ml) illustrated considerable in vitro activity even against P. falciparum resistant strain. Extracts/fractions as well as the isolated compounds were found to be non-toxic with CC50 > 640 µg/ml. Molecular docking studies were performed with these 2 O-glucosides against four malaria targets to understand the binding pose of these molecules and the results suggested that these molecules have selectivity for lactate dehydrogenase enzyme. CKME and EHME exhibited curative activity in vivo along with increase in Mean Survival Time of mice. CONCLUSION: The research delineated the scientific evidence that both the therapeutic herbs possessed antimalarial activity and notably, bioactive compounds responsible to exhibit the antimalarial activity have been isolated, identified and characterized. Further studies are underway to assess the antiplasmodial efficacy of isolated compounds alone and in combination with standard antimalarials.

Application of validated UV spectrophotometric and colorimetric method to quantify minoxidil in the development of trilayer dissolving microneedle: Proof of concept in ex vivo and in vivo studies in rats.

Alopecia areata (AA) is an autoimmune-induced hair loss condition, by utilizing MNX, a hair growth-promoting compound. However, minoxidil (MNX) administration's efficacy is hindered by low bioavailability and adverse effects. To enhance its delivery, Trilayer Dissolving Microneedles (TDMN) are introduced, enabling controlled drug release. The study's primary was to establish a validated UV-Vis Spectrophotometer method for Minoxidil analysis in rat skin affected by alopecia areata. This method adheres to International Conference Harmonization (ICH) and FDA guidelines, encompassing accuracy, precision, linearity, quantification limit (QL), and detection limit (DL). The validation method was conducted through two approaches, namely UV region validation using PBS and the colorimetric method in the visible region (Vis). The validated approach is then employed for assessing in vitro release, ex vivo permeation, and in vivo pharmacokinetics. Results indicate superior MNX extraction recovery using methanol compared to acetonitrile. Method C (5mL methanol) is optimal, offering high recovery with minimal solvent usage. Precision assessments demonstrate %RSD values within MNX guidelines (≤15%), affirming accuracy and reproducibility. UV-Vis spectroscopy quantifies MNX integration into TDMN, using PVA-PVP, with concentrations aligning with ICH standards (95% to 105%). In conclusion, TDMN holds promise for enhancing MNX delivery, mitigating bioavailability and side effect challenges. The validated UV-Vis Spectrophotometer method effectively analyzes MNX in skin tissues, providing insights into AA treatment and establishing a robust analytical foundation for future studies.

Methanolic extract of S. securidaca flowers, leaves, and seeds' antihyperlipidemic effects on high fat diet-induced hyperlipidemia in Wistar rats.

Significant risk factors for atherosclerosis include hyperlipidemia and oxidative stress, which together rank as three of the most significant risk factors for cardiovascular diseases. Securigera securidaca lowers cholesterol levels in diabetic rats' blood. This investigation's objective was to determine how methanolic extracts affected the flowers, leaves, and seeds of plants in rats that were fed a high-fat diet (HFD). Five groups of animals were created (n = 5). A total of 35 days, divided into two intervals, were used for the study. Rats received HFD during the first 15-day interval, while during the second 20-day interval, they also received extracts or the Atorvastatin reference drug. The extract of seeds has a high phenol content as well as DPPH radical antioxidant activity. Extracts were given at a dose of 200 mg/kg; p.o. Methanolic treatment of S. securidaca flowers, leaves, and seeds in HFD-induced hyperlipidemic rats resulted in significant reductions in total cholesterol, triglycerides, LDLC, and VLDL-C levels. HDL-C levels increased significantly because of the leaves. While in hyperlipidemic rats, seeds significantly reduced the activities of the enzymes ALT and ALP. The findings showed that, to a certain extent, seeds, flowers, and leaves may have benefits in reducing hyperlipidemia brought on by HFD in terms of lipid profiles and liver function enzymes. The findings of this study indicate a promising application prospect, but more research is needed to determine the exact mechanism of these novel compounds as antihyperlipidemic agents and to clarify their potential combination effect with synthetic drugs such as Atorvastatin. Combinations can reduce the dose of chemical medications required, which lowers the risk of side effects.

In vitro antimalarial activity of Syzygium cumini fruit fraction.

Background: Malaria is still one of the most severe public health problems worldwide. The development of treatment, prevention, and control of malaria is one of the substantial problems in the world. Aims: To investigate the in vitro antimalarial activity of Syzygium cumini methanol fruit fraction. Methods: Syzygium cumini L fruit powder was macerated with methanol (PA) and the extract obtained was fractionated using the liquid-liquid partition method with n-hexane, ethyl acetate, butanol, chloroform, methanol, and water solvents. In vitro antimalarial assay was conducted using the culture of Plasmodium falciparum 3D7 strain culture that had reached >5% growth and was examined for IC50 values using a 24-well microplate in duplicate. Each treatment and control well contained 1,080 µl of complete media. Well, number 1 was added with 120 µl fraction, and then the solution was diluted until it reached 0.01, 0.1, 1, 10, and 100 µg/ml the final concentration in the microtiter well. The control only contained complete media and infected erythrocytes without the addition of anti-malarial drugs. The microplate was incubated for 48 hours. After 48 hours, a thin blood smear was made fixed with methanol and stained with 20% Giemsa for 20 minutes to determine the IC50 value by plotting sample concentrations and percentage of parasitemia in Excel. Results: The IC50 values of ethyl acetate fraction, n.hexane fraction, butanol fraction, and water fraction were 1.189, 76.996, 1,769, and 15.058 µg/ml, respectively. Whereases the IC50 values of C1 fraction (mix fraction from chloroform: methanol 100:0 and 90:10) and C4 fraction (mix fraction from chloroform: methanol 20:80, 10:90, and 0:100) were 100.126 and 1.015 µg/ml, respectively. The results showed that the IC50 value of ethyl acetate, butanol, and C4 fraction were lower than 10µg/ml and were considered as good activity (strong antimalarial activity). Conclusion: The ethyl acetate, butanol and C4 subfraction from S. cumini fruit have the potential to be developed as an antimalarial agent.

Quantifying metabolic activity of Ascaris suum L3 using resazurin reduction.

BACKGROUND: Helminth infections are an important public health problem in humans and have an even greater impact on domestic animal and livestock welfare. Current readouts for anthelmintic drug screening assays are stage development, migration, or motility that can be subjective, laborious, and low in throughput. The aim of this study was to apply and optimize a fluorometric technique using resazurin for evaluating changes in the metabolic activity of Ascaris suum third-stage larvae (L3), a parasite of high economic relevance in swine. METHODS: Ascaris suum L3 were mechanically hatched from 6- to 8-week embryonated and sucrose-gradient-enriched eggs. Resazurin dye and A. suum L3 were titrated in 96-well microtiter plates, and resazurin reduction activity was assessed by fluorometry after 24 h of incubation. Fluorescence microscopy was used to localize the resazurin reduction site within the larvae. Finally, we exposed A. suum L3 to various stress conditions including heat, methanol, and anthelmintics, and investigated their impact on larval metabolism through resazurin reduction activity. RESULTS: We show that the non-fluorescent dye resazurin is reduced inside vital A. suum L3 to fluorescent resorufin and released into the culture media. Optimal assay parameters are 100-1000 L3 per well, a resazurin concentration of 7.5 µg/ml, and incubation at 37 °C/5% CO2 for 24 h. An intact L2 sheath around the L3 of A. suum completely prevents the uptake of resazurin, while in unsheathed L3, the most intense fluorescence signal is observed along the larval midgut. L3 exposed to methanol or heat show a gradually decreased resazurin reduction activity. In addition, 24 h exposure to ivermectin at 0.625 µM, mebendazole at 5 µM, and thiabendazole from 10 to 100 µM significantly decreased larval metabolic activity by 55%, 73%, and 70% to 89%, respectively. CONCLUSIONS: Together, our results show that both metabolic stressors and anthelmintic drugs significantly and reproducibly reduce the resazurin reduction activity of A. suum L3, making the proposed assay a sensitive and easy-to-use method to evaluate metabolic activity of A. suum L3 in vitro.

In vitro and in vivo anti-inflammatory activity of Cupressus torulosa D.DON needles extract and its chemical characterization.

ETHNOPHARMACOLOGICAL RELEVANCE: Cupressus torulosa (family Cupressaceae), widely distributed in the north western Himalayan region of India, is a coniferous aromatic tree with various traditional uses of its aerial parts. Its needles have been used for anti-inflammatory, anticonvulsant, antimicrobial, and wound-healing properties. AIM OF THE STUDY: The study aimed at investigating the previously unknown anti-inflammatory activity of the hydromethanolic extract of the needles employing in vitro and in vivo assays and scientifically validate traditional claim of their use in treatment of inflammation. Chemical characterization of the extract with the aid of UPLCQTOFMS was also of interest. MATERIALS AND METHODS: C. torulosa needles were first defatted with hexane and sequentially extracted with chloroform and 25% aqueous methanol (AM). Since the presence of phenolics (TPCs, 208.21 ± 0.95 mg GAE/g needles) and flavonoids (TFCs, 84.61 ± 1.21 mg QE/g needles) was observed in the AM extract only, it was chosen for biological and chemical examinations. Acute toxicity of the AM extract on female mice was evaluated following the OECD guideline 423. In vitro anti-inflammatory activity of the AM extract was examined using egg albumin denaturation assay while carrageenan-induced paw edema and formalin-induced paw edema models at doses of 100, 200 and 400 mg/kg po were used to determine the in vivo activity of the AM extract on Wistar rats of either sex. The components of the AM extract were analyzed by UPLC-QTOF-MS method using non-targeted metabolomics approach. RESULTS: AM extract was found to be non-toxic at 2000 mg/kg b.w. with no signs of abnormal locomotion, seizures and writhing. The extract demonstrated promising in vitro anti-inflammatory activity (IC50 160.01 µg/mL) compared to standard diclofenac sodium (IC50 73.94 µg/mL) in egg albumin denaturation assay. In carrageenan-induced paw edema and formalin-induced paw edema tests the extract showed significant anti- inflammatory activity (57.28% and 51.04% inhibition of paw edema, respectively) at the dose of 400 mg/kg p.o. after 4 h in comparison to the standard diclofenac sodium which displayed 61.39% and 52.90% inhibition, respectively, at the dose of 10 mg/kg p.o. after 4 h in these models. A total of 63 chemical constituents, majority of them being phenolics, were found in the AM extract of the needles. Two compounds namely monotropein (iridoid glycoside), (±)12-HETE (eicosanoid) and fraxin (coumarin glycoside) were reported to have anti-inflammatory effect. CONCLUSIONS: For the first time our study demonstrated that hydro-methanolic extract of C. torulosa needles exhibit anti-inflammatory activity thereby supporting their traditional use in the treatment of inflammatory disorders. UPLCQTOFMS assisted chemical profile of the extract was also unveiled.

Anti-inflammatory, anti-nociceptive and anti-pyretic activities of Cenchrus ciliaris L.

ETHNOPHARMACOLOGICAL RELEVANCE: Cenchrus ciliaris L. belongs to the family Poaceae and is found all over the world. It is native to the Cholistan desert of Pakistan where it is locally known as 'Dhaman'. Owing to high nutritional value, C. ciliaris is used as fodder while seeds are used for bread making which are consumed by locals. It also possesses medicinal value and is extensively employed to treat pain, inflammation, urinary tract infection, and tumors. AIM OF STUDY: Studies on the pharmacological activities of C. ciliaris are scarce in spite of its several traditional uses. To the best of our knowledge, no comprehensive study has been conducted on anti-inflammatory, analgesic and anti-pyretic activity of C. ciliaris until now. Here we employed an integrative phytochemical and in - vivo framework to evaluate the potential biological activities of C. ciliaris against inflammation, nociception and pyrexia experimentally induced in rodents. MATERIAL AND METHODS: C. ciliaris was collected from the desert of Cholistan, Bahawalpur, Pakistan. Phytochemical profiling of C. ciliaris was done by employing GC-MS analysis. Anti-inflammatory activity of plant extract was initially determined by various in - vitro assays including albumin denaturation assay and RBC membrane stabilization assays. Finally, rodents were utilized to evaluate in - vivo anti-inflammatory, antipyretic and anti-nociceptive activities. RESULTS: Our data revealed the presence of 67 phytochemicals in methanolic extract of C. ciliaris. The methanolic extract of C. ciliaris provided RBC membrane stabilization by 65.89 ± 0.32% and protection against albumin denaturation by 71.91 ± 3.42% at 1 mg/ml concentration. In in - vivo acute inflammatory models, C. ciliaris exhibited 70.33 ± 1.03, 62.09 ± 8.98, 70.24 ± 0.95% anti-inflammatory activity at concentration of 300 mg/ml against carrageenan, histamine and serotonin induced inflammation. In CFA induced arthritis, inhibition of inflammation was found to be 48.85 ± 5.11% at 300 mg/ml dose after 28 days of treatment. In anti-nociceptive assays C. ciliaris exhibited significant analgesic activity in both peripheral and centrally mediated pain. The C. ciliaris also reduced the temperature by 75.26 ± 1.41% in yeast induced pyrexia. CONCLUSION: C. ciliaris exhibited anti-inflammatory effect against acute and chronic inflammation. It also showed significant anti-nociceptive and anti-pyretic activity which endorses its traditional use in the management of pain and inflammatory disorders.

In vitro and in vivo antileishmanial activity of the hanging sedge flavonoids based on bio- guided fractionation assay.

As a major public health issue, cutaneous leishmaniasis (CL) has a number of complications, including drug resistance and poor response to conventional treatments. Over the last decade, research on natural sources for finding new antileishmanial agents has been a critical part of tropical disease research. Natural products also should be regarded as one of the most valuable applications for CL infection drug development. In this study, we assessed the in vitro and in vivo antileishmanial potential of Carex pendula Huds. (hanging sedge) methanolic extract and its fractions against Leishmania major produced cutaneous infection. Although the methanolic extract and its fractions exhibited suitable activity, the ethyl acetate fraction showed the best activity (with the half maximal inhibitory concentration IC50  = 1.627 ± 0.211 mg/mL). The toxicity and selectivity indices (SI) of all samples were determined in murine peritoneal macrophage cells (J774A.1) using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test. The flavonoid components of the ethyl acetate fraction were identified using liquid chromatography electrospray ionization mass spectrometry (LC-ESI MS/MS). Nine chemical compounds were identified in this fraction, including three flavonols, four flavanonols, and two flavan derivatives. L. major-infected mice were used as an in vivo model because the methanolic extract was effective against L. major promastigotes in the mammalian cell line J774A.1 with SI = 2.514 (tail lesion size model). In silico analysis of identified compounds also revealed a favorable interaction between compounds 2-5 and L. major protein targets (3UIB, 4JZX, 4JZB, 5L4N, and 5L42). According to the findings of this study, the ethyl acetate fraction (as flavonoid fraction) exhibited considerable in vitro antileishmanial activity.

Analgesic, anti-inflammatory activity and metabolite profiling of the methanolic extract of Callicarpa arborea Roxb. leaves.

ETHNOPHARMACOLOGICAL RELEVANCE: Callicarpa arborea Roxb. is widely used as traditional medicine especially by the tribal people of Bangladesh in the management of wide range of ailments. In addition to Bangladesh, the leaves of this plant is utilized as a remedy to various painful and inflammatory conditions including rheumatism, toothache and stomachache in other countries of Indian subcontinent. AIM OF THE STUDY: Depending on the ethnomedicinal uses, we undertook this study to investigate the in-vivo analgesic and anti-inflammatory activities of the methanolic extract of C. arborea Roxb. leaves in Swiss albino mice as well as its chemical composition. MATERIALS AND METHODS: We evaluated the analgesic activity of Callicarpa arborea Roxb. leaves by the acetic acid induced writhing test, the hot plate test, and the formalin test. We undertook the egg albumin induced paw edema test to determine the anti-inflammatory activity of the plant. Furthermore, we conducted the phytochemical profiling by gas chromatography-mass spectrometry (GC-MS). RESULTS: In acute toxicity test, no mortality was observed at the highest dose of 2000 mg/kg b.w. Significant (p < 0.005) inhibition of acetic acid induced writhing was observed at both doses of the extract. A dose dependent increase in the response time was seen in the hot-plate test. In the formalin test, the extract significantly inhibited pain response in both early and late phase. We observed marked anti-inflammatory activity manifested by a significant (p < 0.005) reduction in egg albumin induced paw edema. We identified a total of twenty one compounds in the extract of by GC-MS analysis. CONCLUSION: Taken all into consideration we conclude that the leaves of C. arborea Roxb. possesses potent analgesic and anti-inflammatory activity, thus justifying its's ethnomedicinal use against painful and inflammatory pathological conditions.

Gastroprotective effect methanol extract of Caesalpinia coriaria pods against indomethacin- and ethanol-induced gastric lesions in Wistar rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Caesalpinia coriaria (Jacq.) Willd is widely used as a traditional medinal plant in Mexico for protective and healing purposes and the treatment of gastrointestinal diseases. AIM OF THE STUDY: To investigate the gastroprotective effect of extract of Caesalpinia coriaria pods against ethanol-induced and indomethacin-induced gastric lesion models, its anti-inflammatory and antioxidative activities, and its main compounds through LC-MS analysis. MATERIALS AND METHODS: Male Wistar rats were orally administered a methanol extract obtained from the pods of C. coriaria at doses of 10, 30, 100, and 300 mg/kg prior to inducing gastric lesions with ethanol or indomethacin. Gastric mucosal lesions were evaluated by macroscopic and histopathological alterations. Determination of prostaglandin E2 (PGE2), alpha tumor necrosis factor (TNF-α), leukotriene B4 (LTB4), nitrites/nitrates, superoxide dismutase (SOD), and H2S gastric levels were investigated. Its main compounds of the active extract through LC-MS analysis. RESULTS: Phenolic compounds were identified as major components of methanol extract. LC-MS analysis identified 15 constituents, and the significant compounds were gallic acid, 3-O-galloylquinic acid, digalloylglucose, tetragalloylglucose, valoneic acid dilactone, pentagalloylglucose, digalloylshikimic acid, and ellagic acid. Pretreatment with the extract at doses of 100 and 300 mg/kg significantly reduced gastric ulcer lesions in both models. Compared with the reference drugs (omeprazole or ranitidine, respectively), no significant difference was found (p < 0.05). The extract's gastroprotective effect was accompanied by significant decreases in leukocyte recruitment, and gastric levels of TNF-α and LTB4 by two to fourfold (p < 0.05). Also, gastric levels of PGE2 gastric levels were maintained and the antioxidant enzyme activities of SOD and nitrate/nitrite in the gastric tissue were improved (p < 0.05). The LC-MS analysis indicated the presence of hydrolyzable tannins (mainly gallic acid derivatives). CONCLUSION: The results suggest that the gastroprotective effect of the methanol extract of C. coriaria pods occurs through anti-inflammatory, antioxidant, and NO modulation properties, and gallic acid derivatives may be the main possible compounds responsible for its actions.

In vitro and in vivo anthelmintic properties of Caesalpinia coriaria fruits against Haemonchus contortus.

The purpose of the present study was to assay the in vitro and in vivo anthelmintic activity (AA) of Caesalpinia coriaria (Cc) mature fruits against the nematode Haemonchus contortus (Hc). The Hc infective larvae were used to assess the in vitro AA through larval mortality assay. The exposure of larvae to the different treatments was performed in 96-well microtitration plates. The treatments were as follows: hydroalcoholic extract (HA-E, at 25-100 mg/mL), aqueous fraction (Aq-F, at 12.5-50 mg/mL), organic fraction (EtOAc-F at 12.5-50 mg/mL), compounds (1, methyl gallate and 2, gallic acid at 1.25-10 mg/mL), positive control (ivermectin at 5 mg/mL) and two negative controls (distilled water and 4% methanol). After exposure, dead and live larvae were quantified and results were compared to their controls. The in vivo assay was carried out by a faecal egg count reduction test (FECRT); artificially infected goat kids (F1: Boer x Nubia) were treated with Cc ground dried fruits to assess the AA. The treatments were established as follows: G1-untreated goats (negative control), G2-goats dewormed with ivermectin (positive control), G3-goats fed with Cc mature fruits (10% of their diet). Results in both in vitro and in vivo assays were analysed using an ANOVA through random design, applying a general linear model and mixed models. The in vitro results showed an evident larvicidal effect of the HA-E, EtOAc-F from Cc, indicating that the compound responsible for the AA was gallic acid. The results of the in vivo study corroborated the anthelmintic properties of Cc, reaching 78.6% reduction in the elimination of Hc eggs per gram of faeces. This plant represents a potential natural anthelmintic for the control of haemonchosis in goats under grazing conditions. Future studies should standardise the Cc extract or dried fruits for use in the management of nematodiasis in goat herds.

Methanolic neem (Azadirachta indica) stem bark extract induces cell cycle arrest, apoptosis and inhibits the migration of cervical cancer cells in vitro.

BACKGROUND: Cervical cancer remains one of the significant causes of mortality in women due to the limitations of current treatment strategies and their associated side effects. Investigation of alternative medicine, including phytomedicine, has shown effective anti-cancer potential with fewer side effects. Azadirachta indica (commonly known as neem) is known for its medicinal properties. The present study investigated the anti-cancer potential of methanolic neem stem bark extract (MNBE) against cervical cancer using HeLa, SiHa, and ME-180 cell lines. METHODS: Cytotoxic effect of MNBE on cultured cell lines was evaluated by MTT and clonogenic assay. The growth-inhibiting effect of MNBE was further confirmed by performing cell cycle analysis and apoptosis assay using flow cytometry. The anti-migratory effect of MNBE was evaluated by using wound healing and Boyden chamber assay. Real-time PCR was used to determine the mRNA expression, and western blot and flow cytometry was used to determine the protein levels of growth and migration-related genes. RESULTS: MNBE significantly suppressed the growth and survival of cervical cancer cells in a dose-dependent manner by inducing cell cycle arrest and apoptosis. In addition, the growth inhibitory effect of MNBE was specific to cervical cancer cells than normal cells. Cell cycle arrest was correlated to transcriptional downregulation of cyclin dependent kinase 1 (CDK1), cyclin A, and cyclin B. Additionally, MNBE treatment resulted in the upregulation of active caspase-3 protein and downregulation of prosurvival genes, Bcl2, and survivin at mRNA level and NFkB-p65 at the protein level. Furthermore, MNBE inhibited the migration of cervical cancer cells accompanied by modulation of migration-related genes, including zona occludens-1 (ZO-1), matrix metalloproteinase 2 (MMP2), focal adhesion kinase (FAK), N-cadherin, snail, and E-cadherin. CONCLUSION: In summary, the present study provides the first evidence of MNBE in restricting cervical cancer cell growth and migration, which warrants further investigation for developing novel anti-cancer drugs.

The Effects of Methanol Extracts of Hyssopus officinalis on Model of Induced Glioblastoma Multiforme (GBM) in Rats.

Given the complexity of pathophysiological processes of brain tumors, ineffective therapies, and high mortality rate, new therapeutic options with less toxicity are necessary. Hyssopus officinalis (hyssop) is an aromatic plant with important biological activities. The aim of this study is to assess the anti-cancer effect of hyssop extract on damages of glioblastoma multiforme. In this study, total flavonoids, phenolic content, and quantification of phenolic compound of hyssop extracts were analyzed. In vitro antioxidant properties of hyssop extract were also examined. In addition, cell viability, apoptosis, and cell cycle were evaluated in C6 glioma cell culture. In vivo, the rats were divided randomly into four main groups: intact, control, vehicle, and treatment groups. 1 × 106 C6 rat glioma cells were implanted into the right caudate nucleus of the rat's brain. The treatment group received the methanol extract of hyssop (100 mg/kg) for 7 days. Evolution of locomotor activity, tumor volume, survival rate, activities of antioxidant enzymes (superoxide dismutase (SOD) and catalase (CAT)), vascular endothelial growth factor (VEGF) expression, TUNEL-positive cells, p53 and p21 mRNA expression, and histological alterations were performed. The results showed that the methanol extract of hyssop increased the apoptosis and reduced the cell division of C6 glioma cells in cell culture. Moreover, methanol extract decreased the tumor volume and prolonged survival. Also, the activity of SOD and CAT enzymes was reduced in tumor tissue and enhanced in surrounding tissue. TUNEL-positive cells were increased in methanol extract of hyssop group. The expression of p53 and p21 mRNA was upregulated in the treatment group. Moreover, the histological analysis indicated a considerable decrease in invasion of tumor cells and inflammation in the hyssop-treated rats. According to the achieved results, it can be stated that hyssop has sufficient potential to inhibit damage of brain tumors, at least in part, by affecting the oxidative stress and cell proliferation pathways.

In Vivo Therapeutic Effect of Some Medicinal Plants' Methanolic Extracts on the Growth and Development of Secondary Hydatid Cyst Infection.

PURPOSE: The current study aimed to explore the in vivo therapeutic effects of the methanolic extracts of Citrullus colocynthis, Ruta graveolens, and Peganum harmala against hydatid cyst secondary infection. METHODS: Aerial parts of P. harmala and R. graveolens, including leaves and stems, and seeds of C. colocynthis were collected and extracted using absolute methanol. Rats that are infected with secondary infection of hydatid cysts were treated orally and intraperitoneally according to the determined lethal doses for 30 days. Histological, hematological, and biochemical investigations were done 8 months after the infection. RESULTS: Compared to Albendazole drug, C. colocynthis, and P. harmala, the methanol extract of R. graveolens showed higher and significant (P < 0.05) therapeutic effects on the secondary hydatid cysts growth. Those effects were represented by the reduction in the cysts' number, size, and weight; as well as the significant changes (P < 0.05) in values of hematological and biochemical parameters, the elevation of IFN-γ levels, and the decline of IL-10 and IL-4 cytokines, compared to the negative control group in both routes of treatment (oral and IP). Moreover, the histological sections showed that R. graveolens has a clear damaging effect on the hydatid cysts GL in the infected rats represented by the detachment of GL from LL and AL. CONCLUSION: This study can open an avenue to find new therapeutics for secondary hydatid cyst infections using the studied plant extracts, especially the extract of R. graveolens.

Phytochemical investigation and pharmacological evaluation of methanolic flower extract of Saussurea heteromalla.

Saussurea heteromalla is the one of specie of Saussurea plant belonging to family Asteraceae. The Saussurea heteromalla found extensively in Pakistan. The literature review highlights numerous biological aspects of Saussurea heteromalla. This research therefore aims to assess its potential anti-inflammatory, antioxidant, anti-cancer. Carrageenan induced rat paw edema model was used for the evaluation of anti-inflammatory activity. DPPH method was used to evaluate anti-oxidant activity. The MTT (3- [4, 5-dimethylthiazol-2-yl]-2, 5 diphenyl tetrazolium bromide) test was used to assess the viability of the cells for the assessment of the cytotoxic effect of the extract. Methanolic Saussurea heteromalla flowers extract analysis carried by GC-MS, result 19 different peaks. The methanolic extract of Saussurea heteromalla at 400mg/kg dose have equal anti-inflammatory action when compare with standard that is diclofenac sodium. Anti-oxidant activity of methanolic extract is also very good. IC50 value of methanol extract was 25µg/ml. 18.72% cell survive out of 100 when methanolic flower extract of Saussurea heteromalla was given at the dose of 400mg, which shows the cytotoxic effect. This activity shows that plant Saussurea heteromalla methanolic flowers extract have anti-inflammatory, anti-oxidant, cytotoxic effect. The isolation and characterization-based investigations proclaiming the biologically leading active molecule are worthy for further study in this regard.

In vitro antiplasmodial activity and cytotoxicity of extracts and chromatographic fractions of twigs from Pappea capensis EckI & Zeyh. (Sapindaceae).

ETHNOPHARMACOLOGICAL RELEVANCE: The Vha-Venda people of South Africa use Pappea capensis EckI & Zeyh. (Sapindaceae) twigs to treat malaria and its related symptoms. AIM OF STUDY: The main aim of this study was to evaluate the antiplasmodial and cytotoxic activity of P. capensis extracts and chromatographic fractions. Spectroscopy analysis was conducted using 1H NMR and GC-MS to tentatively identify the major classes of compounds and phytoconstituents that can be attributed to the observed antiplasmodial bioactivity. MATERIALS AND METHODS: Pappea capensis twigs were dried and then ground to fine powder. A solvent mixture of dichloromethane: methanol: water (1:0.5:0.5, v/v) was used to extract. The polar extract was separated from the non-polar. The organic extract was dried to yield a DCM (I = 60 g) extract. The methanol in the aqueous extract was evaporated using a rotary vapour and the remaining water freeze dried to yield a water extract (II = 287 g). Extract I was further partitioned using a solvent mixture of DCM: MeOH (1:1, v/v), separated and concentrated under vacuum to yield dichloromethane (III = 40 g) and methanol (IV = 15 g) extracts. A water-based decoction (V = 10 g) was also prepared to establish the clinical relevance of the preparation administered by Vha-Venda people in South Africa. Extracts II, III and IV were further subjected to silica column chromatography, eluting with a series of different solvents with increasing polarity to yield a total of 25 fractions (A - Y). In vitro antiplasmodial tests on Plasmodium falciparum (NF54) and cytotoxicity screens on mammalian L-6 rat skeletal myoblast cells were performed on all extracts and fractions. Selectivity indices (SI) were also computed for all tested extracts and fractions which were further subjected to 1H NMR spectroscopy and GC-MS analysis for the identification of the major classes of compounds present in the extracts. RESULTS: From the assayed extracts, only extract I (IC50 = 2.93 µg/ml; SI = 14), III (IC50 = 2.59 µg/ml; SI = 21) and IV (IC50 = 3.56 µg/ml; SI = 13) demonstrated the best antiplasmodial activity and selectivity. Of all assayed fractions, only N (0.6 µg/ml; SI = 91), D (0.85 µg/ml; SI = 37) and E (0.91 µg/ml; SI = 30) depicted the best antiplasmodial activity and selectivity. The 1H NMR analysis of the extracts and fractions identified the prominent class of constituents to be aliphatic based which was tentatively identified as terpenoids. When further GC-MS analysis was conducted, the presence of lupin-3-one, lupeol acetate, α-amyrin, and ß-amyrin phytoconstituents were tentatively confirmed. These constituents are triterpenoids with established antiplasmodial activity which can be tentatively attributed to the bioactivity observed in P. capensis twigs. CONCLUSION: The study validates the ethnomedicinal use of P. capensis for malaria treatment. It demonstrated the potential of discovering novel antiplasmodial constituents that could serve as drug hits through dereplication approaches where known compounds with established antimalarial activity can be bypassed to focus on the unknown.