Utilizing Andrographis paniculata leaves and roots by effective usage of the bioactive andrographolide and its nanodelivery: investigation of antikindling and antioxidant activities through in silico and in vivo studies.

To valorise the bioactive constituents abundant in leaves and other parts of medicinal plants with the objective to minimize the plant-based wastes, this study was undertaken. The main bioactive constituent of Andrographis paniculata, an Asian medicinal plant, is andrographolide (AG, a diterpenoid), which has shown promising results in the treatment of neurodegenerative illnesses. Continuous electrical activity in the brain is a hallmark of the abnormal neurological conditions such as epilepsy (EY). This can lead to neurological sequelae. In this study, we used GSE28674 as a microarray expression profiling dataset to identify DEGs associated with andrographolide and those with fold changes >1 and p-value <0.05 GEO2R. We obtained eight DEG datasets (two up and six down). There was marked enrichment under various Kyoto Encyclopaedia of Genes and Genomes (KEGG) and Gene Ontology (GO) terms for these DEGs (DUSP10, FN1, AR, PRKCE, CA12, RBP4, GABRG2, and GABRA2). Synaptic vesicles and plasma membranes were the predominant sites of DEG expression. AG acts as an antiepileptic agent by upregulating GABA levels. The low bioavailability of AG is a significant limitation of its application. To control these limitations, andrographolide nanoparticles (AGNPs) were prepared and their neuroprotective effect against pentylenetetrazol (PTZ)-induced kindling epilepsy was investigated using network pharmacology (NP) and docking studies to evaluate the antiepileptic multi-target mechanisms of AG. Andrographolide is associated with eight targets in the treatment of epilepsy. Nicotine addiction, GABAergic synapse, and morphine addiction were mainly related to epilepsy, according to KEGG pathway enrichment analysis (p < 0.05). A docking study showed that andrographolide interacted with the key targets. AG regulates epilepsy and exerts its therapeutic effects by stimulating GABA production. Rats received 80 mg/kg body weight of AG and AGNP, phenytoin and PTZ (30 mg/kg i.p. injection on alternate days), brain MDA, SOD, GSH, GABAand histological changes of hippocampus and cortex were observed. PTZ injected rats showed significantly (***p < 0.001) increased kindling behavior, increased MDA, decreased GSH, SOD, GABA activities, compared with normal rats, while treatment AGNPs significantly reduced kindling score and reversed oxidative damage. Finally, we conclude that the leaves and roots of A. Paniculata can be effectively utilized for its major bioactive constituent, andrographolide as a potent anti-epileptic agent. Furthermore, the findings of novel nanotherapeutic approach claim that nano-andrographolide can be successfully in the management of kindling seizures and neurodegenerative disorders.

Bioproduction of eriodictyol by Escherichia coli engineered co-culture.

Eriodictyol (ED) is a flavonoid in the flavanones subclass. It is abundantly present in a wide range of medicinal plants, citrus fruits, and vegetables. In addition, ED owns numerous importantly medicinal bioactivities such as inhibition of proliferation, metastasis and induction of apoptosis in glioma cells or inhibition of glioblastoma migration, and invasion. This study described the heterologous production of ED by E. coli based co-culture engineering system from the simple carbon substrate D-glucose. Two E. coli strains were engineered and functioned as constitutive components of biological system. Specifically, the first strain (upstream module) contained genes for synthesis of p-coumaric acid (pCA) from D-glucose. And, the second strain (downstream module) consisted of genes for the synthesis of ED from pCA. The highest yield in ED production was achieved 51.5 ± 0.4 mg/L using stepwise optimal culture conditions, while monoculture was achieved 21.3 ± 0.2 mg/L only. In conclusion, co-culture was the most efficient alternative approach for the synthesis of ED and other natural products.

Protective effects of extracts of lichen Dirinaria consimilis (Stirton) D.D. Awasthi in bifenthrin- and diazinon-induced oxidative stress in rat erythrocytes in vitro.

The intoxication of insecticides such as bifenthrin and diazinon has been reported to generate free radicals, and thereby alter the antioxidant defense system in erythrocytes. The present study is aimed to investigate the protective effects of acetone (DA) and methanolic (DM) extracts of lichen Dirinaria consimilis against bifenthrin and diazinon toxicity in rats' erythrocytes in vitro. Rats' erythrocytes were exposed to bifenthrin and diazinon, individually and also in combination with DA or DM at 1 ppm for 3 h at 37 ˚C. By using spectrophotometric methods, all the samples were estimated for changes in hemoglobin (Hb) concentration, malondialdehyde (MDA) levels, and enzyme [Superoxide dismutase (SOD), catalase (CAT), and glutathione S-transferases (GST)] activities. The outcomes showed that both the insecticides were highly toxic to rats' erythrocytes. Among all groups, both the insecticides and DA exposed groups showed very low levels of MDA content, and GST activity in rats' erythrocytes, when compared to the control. Besides, DA groups pretreated with both insecticides showed significant improvement of total Hb concentration, SOD, and CAT activities, with respect to the control. Hence, the present results indicate that the extracts of D. consimilis act as an antioxidant agent that reduces oxidative stress burden in insecticides toxicity.

Antimycobacterial activity of acetone extract and isolated metabolites from folklore medicinal lichen Usnea laevis Nyl. against drug-sensitive and multidrug-resistant tuberculosis strains.

ETHNOPHARMACOLOGICAL RELEVANCE: Tuberculosis (Tb) is one of the most infectious diseases caused by Mycobacterium tuberculosis (M.t) with almost 2 million deaths yearly. Although many Tb control programs have been organised, there is an elevated number of Tb cases due to the appearance of extremely drug-resistant and multidrug-resistant (MDR) Tb strains. In the cultures of Venezuelan Andes, fruticose lichen Usnea laevis Nyl. (Usneaceae) with folklore name 'Barba de Piedra, Tusinya' is used as a natural remedy for Tb. AIM OF THE STUDY: This study was performed to provide a scientific rationale for the folklore usage of U. laevis in treating Tb by validating its antimycobacterial activity against two drug-sensitive and four MDR-Tb strains. MATERIALS AND METHODS: The mycobacterial inhibitory activities of acetone extract (Ul), fractions (F1-10), and isolated metabolites (1-4) of U. laevis were evaluated against M.t H37Ra using 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide reduction menadione assay (XRMA). Furthermore, Ul and 1-4 were subjected to antimycobacterial activity against M.t H37Ra, Mycobacterium smegmatis, and four MDR-Tb (MDR-A8, MDR-V791, MDR-R and MDR-40) strains using resazurin microtitre plate assay (REMA) and cytotoxicity against THP-1 macrophages using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and their selectivity index values were also calculated. RESULTS: Initially, Ul has shown prominent inhibitory activity (IC50 value: 5.44 ± 0.36 µg/ml) and four of its fractions (F1, F2, F5 and F7) also exhibited the best inhibitory activity (IC50 values ranged from 7.46 ± 0.19 to 71.38 ± 2.57 µg/ml) against M.t H37Ra using XRMA. Purification of these bioactive fractions identified four metabolites, namely usnic acid (1), atranorin (2), salazinic acid (3), and lobaric acid (4). From the MIC values of REMA, it was identified that Ul, 1 and 4 were more effective in inhibiting the growth of all four MDR-Tb strains, compared to first-line drug rifampicin. Interestingly, Ul has shown better antimycobacterial activity than 1-4 and rifampicin against MDR-Tb strains may be due to the synergistic effect of its metabolites. Also, the IC50 values of Ul and 1-4 on THP-1 macrophages were found to be far higher than MIC values against tested Tb strains, indicating that THP-1 macrophages were not harmfully affected at concentrations that were effective against Tb strains. Further, the calculated selectivity index values revealed the more active and non-toxicity of Ul, 1 and 4 against MDR-Tb strains than rifampicin. CONCLUSIONS: The current study lends the first evidence for the presence of antimycobacterial metabolites in U. laevis. The results exposed the Andean folklore use of U. laevis for treating Tb, and the key biomarker metabolites were found to be 1 and 4. Hence, it can be concluded that U. laevis can be used as a potential source for the novel drug development for MDR-Tb.

Cleroda-4(18),13-dien-15,16-olide as novel xanthine oxidase inhibitors: An integrated in silico and in vitro study.

In the present study, in silico predictions and molecular docking were performed on five clerodane diterpenes (1-5) from Polyalthia longifolia seeds to evaluate their potential as xanthine oxidase (XO) inhibitors. The initial screening was conducted by target prediction using TargetNet web server application and only compounds 3 and 4 showed a potential interaction with XO. Compounds 3 and 4 were subsequently subjected to in silico analyses on XO protein structure (PDB: 1N5X) using Schrödinger Release 2020-3 followed by structural modeling & molecular simulation studies to confirm the initial prediction result and identify the binding mode of these compounds to the XO. Molecular docking results revealed that compounds 3 (-37.3 kcal/mol) and 4 (-32.0 kcal/mol) binds more stably to XO than the reference drug allopurinol (-27.0 kcal/mol). Interestingly, two residues Glu 802 and Thr 1010 were observed as the two main H-bond binding sites for both tested compounds and the allopurinol. The center scaffold of allopurinol was positioned by some π-π stacking with Phe 914 and Phe 1009, while that of compounds 3 and 4 were supported by many hydrophobic interactions mainly with Leu 648, Phe 649, Phe 1013, and Leu 1014. Additionally, the docking simulation predicted that the inhibitory effect of compounds 3 and 4 was mediated by creating H-bond with particularly Glu 802, which is a key amino acid for XO enzyme inhibition. Altogether, in vitro studies showed that compounds 3 and 4 had better inhibitory capacity against XO enzyme with IC50 values significantly (p < 0.001) lower than that of allopurinol. In short, the present study identified cleroda-4(18),13-dien-15,16-olide as novel potential XO inhibitors, which can be potentially used for the treatment of gout.

In vitro antimycobacterial studies of flavonols from Bauhinia vahlii Wight and Arn.

Mycobacterial infections and fast-growing strains are increasing globally with 8 million new cases and 1.8 million fatalities per annum worldwide. The acid-fast bacterium, Mycobacterium tuberculosis (M.t), can spread diseases like tuberculosis (Tb) and weaken the immune system. In Ayurveda, the Bauhinia genus is most valued for the treatment of tuberculosis lymphadenitis. The objective of the present study is to identify anti-tubercular compounds from the under-investigated medicinal plant B. vahlii Wight and Arn. using bioassay guided isolation. The antimycobacterial activity was evaluated against non-virulent strains: Mycobacterium tuberculosis H37Ra (ATCC 25177) and Mycobacterium bovis BCG (ATCC 35743). Also, antibacterial and cytotoxicity activities were tested to identify the specificity of the isolated metabolites. Bioassay-guided isolation yielded three known flavonols, namely quercetin (1), ombuin (2), and kaempferol (3), from the methanolic extract of bark of B. vahlii. The results of antimycobacterial activity tests revealed that 2 showed much better mycobactericidal activity than 1 and 3 under ex vivo condition with minimum inhibitory concentration (MIC) values ranged from 0.05 ± 0.01 to 0.26 ± 0.01 nM, and half-maximal inhibitory concentration (IC50) values ranged from 2.85 ± 0.14 to 7.21 ± 1.09 nM against dormant and active forms, respectively. Also, compound 2 showed higher resistance with MIC values > 100 µg/mL against both Gram-positive and Gram-negative bacteria and the least cytotoxicity up to 100 µg/mL concentration against the tested series of cancer cell lines. The results revealed the Ayurvedic use of extracts of the Bauhinia genus for treating tuberculosis, and the key bioactive compounds were found to be flavonols (1-3). The present work provides the first evidence for the presence of antimycobacterial compounds in B. vahlii. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02672-4.

Sekikaic acid modulates pancreatic ß-cells in streptozotocin-induced type 2 diabetic rats by inhibiting digestive enzymes.

The antioxidant and antidiabetic effects of sekikaic acid (SA) were investigated using in vitro and in vivo study models. SA possessed good antioxidant activity as assessed through hydroxyl radicals (IC50 value = 41.5 µg/mL) and ferric ions assay (IC50 value = 42.0 µg/mL). SA exhibited stronger α-glucosidase and α-amylase inhibition than that of aldose-reductase and protein tyrosine phosphatase 1B. The hypoglycemic activity of SA caused significant reduction of plasma glucose levels in normal and glucose loaded rats. The anti-hyperglycemic activity of SA (2 mg/Kg body weight) was indicated by the reduction of blood glucose by 44.17 ± 3.78% in the third week in streptozotocin-induced diabetic rats. The hypolipidaemic action of SA was evident by the significant decrease in the levels of low-density lipoprotein, total cholesterol, and total glycerides. Histologically, the pancreas of the treated groups showed significant regeneration of the pancreatic ß-cells compared to diabetic control, possibly due to the inhibition of digestive enzymes.

Dual COX and 5-LOX inhibition by clerodane diterpenes from seeds of Polyalthia longifolia (Sonn.) Thwaites.

Natural metabolites with their specific bioactivities are being considered as a potential source of materials for pharmacological studies. In this study, we successfully isolated and identified five known clerodane diterpenes, namely 16-oxo-cleroda-3,13(14)E-dien-15-oic acid (1), 16-hydroxy-cleroda-3,13-dien-15-oic acid (2), 16-hydroxy-cleroda-4(18),13-dien-16,15-olide (3), 3α,16α-dihydroxy-cleroda-4(18),13(14)Z-dien-15,16-olide (4), and 16α-hydroxy-cleroda-3,13(14)Z-dien-15,16-olide (5) from the methanolic extract of seeds of Polyalthia longifolia. Initially, all the isolated metabolites were investigated for COX-1, COX-2, and 5-LOX inhibitory activities using the standard inhibitory kits. Of which, compounds 3, 4, and 5 exhibited to be potent COX-1, COX-2, and 5-LOX inhibitors with the IC50 values similar or lower to those of the reference drugs. To understand the underlying mechanism, these compounds were subjected to molecular docking on COX-1, COX-2, and 5-LOX proteins. Interestingly, the in silico study results were in high accordance with in vitro studies where compounds 3, 4, and 5 hits assumed interactions and binding pattern comparable to that of reference drugs (indomethacin and diclofenac), as a co-crystallized ligand explaining their remarkable dual (COX/LOX) inhibitor actions. Taken together, our findings demonstrated that compounds 3, 4, and 5 functioned as dual inhibitors of COX/5-LOX and can contribute to the development of novel, more effective anti-inflammatory drugs with minimal side-effects.