Acorus calamus L. rhizome extract and its bioactive fraction exhibits antibacterial effect by modulating membrane permeability and fatty acid composition.

ETHNOPHARMACOLOGICAL RELEVANCE: India's ancient texts, the Charak Samhita and Sushruta Samhita, make reference to the traditional medicinal usage of Acorus calamus L. In India and China, it has long been used to cure stomach aches, cuts, diarrhea, and skin conditions. This ability of the rhizome is attributed to its antimicrobial properties. Research studies to date have shown its antimicrobial properties. However, scientific evidence on its mode of action is still lacking. AIM OF THE STUDY: Acorus calamus L. rhizome extract and its bioactive fraction exhibits antibacterial effect by modulating membrane permeability and fatty acid composition. MATERIAL AND METHOD: The secondary metabolites in the rhizome of A. calamus L. were extracted in hexane using Soxhlet apparatus. The ability of the extract to inhibit multidrug resistant bacterial isolates, namely Bacillus cereus, Escherichia coli, Acinetobacter baumannii, and Pseudomonas aeruginosa were evaluated using checkerboard assay. Further, the extract was purified using thin layer chromatography, gravity column chromatography, and combiflash chromatography. Structure elucidation of the active compound was done using GC-MS, FT-IR, and UV-Vis spectral scan. The mode of action of the bioactive fraction was determined. Bacterial membrane damage was analyzed using SEM, membrane permeability was determined using SYBR green I and PI dye, leakage of cytoplasmic contents were analyzed using Bradford assay and Fehling's reagent. The ability to inhibit efflux pump of A. baumannii was determined using EtBr accumulation assay and ß-lactamase inhibition was analyzed using nitrocefin as substrate. Also, the biofilm inhibition of B. cereus was determined using crystal violet dye. Moreover, the effect of the bioactive fraction on the fatty acid profile of the bacterial membrane was determined by GC-FAME analysis using 37 component FAME mix as standard. RESULTS: Acorus calamus L. rhizome hexane extract (AC-R-H) demonstrated broad-spectrum antibacterial activity against all the isolates tested. AC-R-H extract also significantly reduced the MIC of ampicillin against all tested bacteria, indicating its bacterial resistance modulating properties. The assay guided purification determined Asarone as the major compound present in the bioactive fraction (S-III-BAF). S-III-BAF was found to reduce the MIC of ampicillin against Escherichia coli (100-25 mg/mL), Pseudomonas aeruginosa (15-3.25 mg/mL), Acinetobacter baumannii (12.5-1.56 mg/ml), and Bacillus cereus (10-1.25 mg/mL). Further, it recorded synergistic activity with ampicillin against B. cereus (FICI = 0.365), P. aeruginosa (FICI = 0.456), and A. baumannii (FICI = 0.245). The mode of action of S-III-BAF can be attributed to its ability to disturb the membrane integrity, enhance membrane permeability, reduce biofilm formation, and possibly alter the fatty acid composition of the bacterial cell membranes. CONCLUSION: The bioactive fraction of AC-R-H extract containing Asarone as the active compound showed antibacterial activity and synergistic interactions with ampicillin against the tested bacterial isolates. Such activity can be attributed to the modulation of fatty acids present in bacterial membranes, which enhances membrane permeability and causes membrane damage.

Computational insights into the role of structurally diverse plant secondary metabolites as inhibitors against Imidazole Glycerol Phosphate Dehydratase of Mycobacterium tuberculosis.

Mycobacterium tuberculosis (Mtb) is one of the major causes of death worldwide and there is a pressing need for the development of novel drug leads. The Imidazole Glycerol Phosphate Dehydratase (IGPD) of Mtb is one of the key enzymes in the histidine biosynthesis pathway and has been recognized as the potentially underexploited drug target for anti-tuberculosis treatment. In the present study, 6063 structurally diverse plant secondary metabolites (PSM) were screened for their efficiency in inhibiting the catalytic activity of IGPD through molecular docking. The top 150 PSMs with the lowest binding energy represent the chemical classes, including Tannins (34%), Flavonoid Glycosides (14%), Terpene Glycosides (10%), Steroid Lactones (9.3%), Flavonoids (6.6%), Steroidal Glycosides (4.6%), etc. Bismahanine, Ashwagandhanolide, and Daurisoline form stable IGPD-inhibitor complexes with binding free energies of -291.3 ± 16.5, -279.0 ± 25.0, and -279.8 ± 17.6 KJ/mol, respectively, as determined by molecular dynamics simulations. These PSM demonstrated strong H-bond interactions with the amino acid residues Ile279, Arg281, and Lys276 in the catalytic region of IGPD, as revealed by structural snapshots. On the basis of our findings, these three PSM could be considered as possible leads against IGPD and should be explored in vitro and in vivo.Communicated by Ramaswamy H. Sarma.

Imidazole Glycerol Phosphate Dehydratase (IGPD) is an unexplored drug target in tuberculosis therapy.Inhibitory potential of 6063 plant secondary metabolites (PSM) against IGPD enzyme was studied.Ensemble docking and structural-activity relationship studies ascertained the group of diverse molecules.MD simulations predicted Bismahanine and Ashwagandhanolide as possible inhibitors of IGPD.

In vitro and computational studies of the ß-lactamase inhibition and ß-lactam potentiating properties of plant secondary metabolites.

ß-lactam resistance in bacteria is primarily mediated through the production of ß-lactamases. Among the several strategies explored to mitigate the issue of ß-lactam resistance, the use of plant secondary metabolites in combination with existing ß-lactams seem promising. The present study aims to identify possible ß-lactam potentiating plant secondary metabolites following in vitro and in silico approaches. Among 180 extracts from selected 30 medicinal plants, acetone extract of Ficus religiosa (FRAE) bark recorded the least IC50 value of 3.9 mg/ml. Under in vitro conditions, FRAE potentiated the activity of ampicillin, which was evidenced by the significant reduction in IC50 values of ampicillin against multidrug resistant bacteria. Metabolic profiling following HR-LCMS analysis revealed the presence of diverse metabolites viz. flavonoids, alkaloids, terpenoids, etc. in FRAE. Further, ensemble docking of the FRAE metabolites against four Class A ß-lactamase (SHV1, TEM1, KPC2 and CTX-M-27) showed quercetin, taxifolin, myricetin, luteolin, and miquelianin as potential inhibitors with the least average binding energy. In molecular dynamic simulation studies, myricetin formed the most stable complex with SHV1 and KPC-2 while miquelianin with TEM1 and CTX-M-27. Further, all five metabolites interacted with amino acid residue Glu166 in Ω loop of ß-lactamase, interfering with the deacylation step, thereby disrupting the enzyme activity. The pharmacokinetics and ADMET profile indicate their drug-likeness and non-toxic nature, making them ideal ß-lactam potentiators. This study highlights the ability of metabolites present in FRAE to act as ß-lactamase inhibitors.Communicated by Ramaswamy H. Sarma.

Development of Wash-Durable Antimicrobial Cotton Fabrics by In Situ Green Synthesis of Silver Nanoparticles and Investigation of Their Antimicrobial Efficacy against Drug-Resistant Bacteria.

An environment friendly and wash-durable silver nanoparticle treatment of cotton fabrics was carried out by in situ reduction of silver nitrate using Azadirachta indica leaf extract. The wash durability of the silver nanoparticles treatment on the cotton fabric was improved by pretreating the fabrics by mercerization and by adopting hydrothermal conditions of 120 °C temperature and 15 psi pressure for the in situ synthesis. The silver nanoparticle treated fabrics were characterized using scanning electron microscopy, colorimetric analysis and inductively coupled plasma mass spectroscopy. The coating of silver nanoparticles was seen to be dense and uniform in the scanning electron micrographs of the treated fabrics. An evaluation of the antibacterial efficacy of the silver nanoparticle treated fabric against antibiotic-resistant Gram-positive and Gram-negative strains was carried out. The antibacterial efficacy was found to be the highest against Bacillus licheniformis, showing 93.3% inhibition, whereas it was moderate against Klebsiella pneumoniae (20%) and Escherichia coli (10%). The transmittance data of a UV spectrophotometer (290-400nm) was used for measuring the UV protection factor of the silver nanoparticle treated fabrics. All the silver nanoparticle treated fabrics showed good antimicrobial and UV protection activity. The treatment was also seen to be durable against repeated laundering. This paper contributes the first report on a novel green synthesis approach integrating mercerization of cotton fabrics and in situ synthesis of nanoparticles under hydrothermal conditions using Azadirachta indica leaf extract for improved wash durability of the multifunctional fabric.

In silico studies evidenced the role of structurally diverse plant secondary metabolites in reducing SARS-CoV-2 pathogenesis.

Plants are endowed with a large pool of structurally diverse small molecules known as secondary metabolites. The present study aims to virtually screen these plant secondary metabolites (PSM) for their possible anti-SARS-CoV-2 properties targeting four proteins/ enzymes which govern viral pathogenesis. Results of molecular docking with 4,704 ligands against four target proteins, and data analysis revealed a unique pattern of structurally similar PSM interacting with the target proteins. Among the top-ranked PSM which recorded lower binding energy (BE), > 50% were triterpenoids which interacted strongly with viral spike protein-receptor binding domain, > 32% molecules which showed better interaction with the active site of human transmembrane serine protease were belongs to flavonoids and their glycosides, > 16% of flavonol glycosides and > 16% anthocyanidins recorded lower BE against active site of viral main protease and > 13% flavonol glycoside strongly interacted with active site of viral RNA-dependent RNA polymerase. The primary concern about these PSM is their bioavailability. However, several PSM recorded higher bioavailability score and found fulfilling most of the drug-likeness characters as per Lipinski's rule (Coagulin K, Kamalachalcone C, Ginkgetin, Isoginkgetin, 3,3'-Biplumbagin, Chrysophanein, Aromoline, etc.). Natural occurrence, bio-transformation, bioavailability of selected PSM and their interaction with the target site of selected proteins were discussed in detail. Present study provides a platform for researchers to explore the possible use of selected PSM to prevent/ cure the COVID-19 by subjecting them for thorough in vitro and in vivo evaluation for the capabilities to interfering with the process of viral host cell recognition, entry and replication.

Opportunities and challenges in managing antibiotic resistance in bacteria using plant secondary metabolites.

Development of antibiotic resistance (ABR) in bacteria and its multidimensional spread is an emerging global threat that needs immediate attention. Extensive antibiotics (AB) usage results in development of ABR in bacteria by target modification, production of AB degrading enzymes, porin modifications, efflux pumps overexpression, etc. To counter this, apart from strict regulation of AB use and behavioural changes, research and development (R&D) of newer antimicrobials are in place. One such emerging approach to combat ABR is the use of structurally and functionally diverse plant secondary metabolites (PSMs) in combination with the conventional AB. Either the PSMs are themselves antimicrobial or they potentiate the activity of the AB through a range of mechanisms. However, their use is lagging due to poor knowledge of mode of action, structure-activity relationships, pharmacokinetics, etc. This review paper discussed the opportunities and challenges in managing ABR using PSMs. Mechanisms of ABR development in bacteria and current strategies to counter them were studied and the areas where PSMs can play an important role were highlighted. The use of PSMs, both as an anti-resistance and anti-virulence agent in combination therapy to counter multi-drug resistance along with their mechanisms of action, has been discussed in detail. The difficulties in the commercialisation of PSMs and strategies to overcome them along with future priority areas of research have also been given. Following the given R&D path will definitely help in better understanding and utilising the full potential of PSMs in solving the problem of antimicrobial resistance (AMR).