Fractionation Of Plant Extracts Research Articles

Screening of the Medicinal Plant-Based Metabolites Responsible for Silver Nanoparticle Phytofabrication

In this study, the detailed phytochemical composition of two medicinal plants was investigated and their importance in silver nanoparticles (AgNPs) synthesis was highlighted. Aqueous extracts from the leaves of both plants were used for various analytical techniques, including UV-vis spectroscopy, Gas chromatography-mass spectrometry (GC-MS), Fourier transform infrared spectroscopy (FTIR) and Thin layer chromatography (TLC). Quantification of total phenolics and flavonoids in different fractions of plant extracts revealed significant concentrations. TLC profiling showed a higher abundance of free phenolic and flavonoid compounds compared to bound forms in both plants, suggesting that they contribute significantly to total phenolic and flavonoid content. UV-vis spectra from 200 to 600 nm revealed the presence of aromatic rings and chromophores in leaf extracts. Furthermore, the GC-MS analysis identified several bioactive compounds, some of which were found to be common between these two species. The results demonstrate that various bioactive compounds such as phenols, flavonoids, alkaloids, carotenoids and terpenoids were present in these plant species. These compounds efficiently serve as both reducing and stabilizing agents during phytofabrication of AgNPs, consequently eliminating the need for the use of hazardous chemicals.

Shining insights: Deciphering the biogenic synthesis of Ajuga bracteosa-mediated gold nanoparticles with advanced microscopy techniques.

In this study, gold nanoparticles (AuNPs) were bioreduced from Ajuga bracteosa, a medicinal herb known for its therapeutic properties against various diseases. Different fractions of the plant extract were used, including the methanolic fraction (ABMF), the n-hexane fraction (ABHF), the chloroform fraction (ABCF), and the aqueous extract for AuNPs synthesis. The characterization of AuNPs was performed using UV-Vis spectrophotometry, FT-IR, XRD, EDX, and TEM. UV-Vis spectroscopy confirmed the formation of AuNPs, with peaks observed at 555 nm. FT-IR analysis indicated strong capping of phytochemicals on the surface of AuNPs, which was supported by higher total phenolic contents (TPC) and total flavonoid contents (TFC) in AuNPs. XRD results showed high crystallinity and a smaller size distribution of AuNPs. TEM analysis revealed the spherical shape of AuNPs, with an average size of 29 ± 10 nm. The biologically synthesized AuNPs exhibited superior antibacterial, antioxidant, and cytotoxic activities compared to the plant extract fractions. The presence of active biomolecules in A. bracteosa, such as neoclerodan flavonol glycosides, diterpenoids, phytoecdysone, and iridoid glycosides, contributed to the enhanced biological activities of AuNPs. Overall, this research highlights the potential of A. bracteosa-derived AuNPs for various biomedical applications due to their remarkable therapeutic properties and effective capping by phytochemicals. RESEARCH HIGHLIGHTS: This research underscores the growing significance of herbal medicine in contemporary healthcare by exploring the therapeutic potential of Ajuga bracteosa and gold nanoparticles (AuNPs). The study highlights the notable efficacy of A. bracteosa leaf extracts and AuNPs in treating bacterial infections, demonstrating their bactericidal effects on a range of strains. The anti-inflammatory properties of plant extracts and nanoparticles are evidenced through paw edema method suggesting their applicability in managing inflammatory conditions. These findings position A. bracteosa and AuNPs as potential candidates for alternative and effective approaches to modern medication.

Chloride, glutathiones, and insect-derived elicitors introduced into the xylem trigger electrical signaling.

Ricca assays allow the direct introduction of compounds extracted from plants or the organisms that attack them into the leaf vasculature. Using chromatographic fractionation of Arabidopsis (Arabidopsis thaliana) leaf extracts, we found glutamate was the most active low mass elicitor of membrane depolarization. However, other known elicitors of membrane depolarization are generated in the wound response. These include unstable aglycones generated by glucosinolate (GSL) breakdown. None of the aglycone-derived GSL-breakdown products, including nitriles and isothiocyanates, that we tested using Ricca assays triggered electrical activity. Instead, we found that glutathione and the GSL-derived compound sulforaphane glutathione triggered membrane depolarizations. These findings identify a potential link between GSL breakdown and glutathione in the generation of membrane depolarizing signals. Noting that the chromatographic fractionation of plant extracts can dilute or exchange ions, we found that Cl- caused glutamate receptor-like3.3-dependent membrane depolarizations. In summary, we show that, in addition to glutamate, glutathione derivatives as well as chloride ions will need to be considered as potential elicitors of wound-response membrane potential change. Finally, by introducing aphid (Brevicoryne brassicae) extracts or the flagellin-derived peptide flg22 into the leaf vasculature we extend the use of Ricca assays for the exploration of insect/plant and bacteria/plant interactions.

Inhibition of α-glucosidase activity by polyphenol compounds from C. occidentalis: Phytochemical screening and antidiabetic studies

Medicinal plants are recognized as a source of active molecules that can treat several diseases. Cassia occidentalis (C. occidentalis) is a medicinal plant traditionally used in rural areas of Burkina Faso for the treatment of diabetes. The objective of this work is to evaluate the antioxidant and antidiabetic properties of the fractions of this plant. The antidiabetic activity was investigated by following the inhibitory effect of the different fractions of plant extract on α-glucosidase. The antioxidant activities were performed using 2, 2-diphenyl-1-picryl-hydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) methods. TLC analysis revealed the presence of quercetin in the ethyl acetate fractions of stems and leaves of C. occidentalis. Ethyl acetate fractions of stems and leaves of C. occidentalis demonstrated significant antidiabetic properties with IC50 values ca. 0.274 ± 0.003 and 0.538 ± 0.011 mg/mL respectively compared to the reference acarbose with IC50 ca. 0.215 ± 0.004 mg/mL. The same fractions exhibited respectively the highest FRAP values close to 167.055 ± 0.008 and 128.490 ± 5.227 μg ET/mg and DPPH values of 114.062 ± 2.698 and 82.962 ± 3.189 μg ET/mg. Phytochemical screening revealed the presence of flavonoids, tannins and quinones. These results show that C. occidentalis possess molecules with interesting antidiabetic properties and demonstrate its use in the treatment of diabetes.

In-vitro and in-silico evaluation of Brugmansia sauveolens' ability to treat asthma

Asthma is a common chronic inflammatory disease of the airways. The airway epithelium produces bioactive cellular mediators that trigger the chronic inflammatory response. These mediators then attract and activate inflammatory cells in the airways of the lungs and release more biochemical mediators. With several benefits, such as a significant reduction in the cost and time of drug discovery as well as the use of laboratory animals, in-silico studies have grown to be an important and simple method for examining various chemical compounds against various targets. One of the common herbs found in the local Himalayan area is Brugmansia sauveolens, known as Datura, which is a source of tropane alkaloids and asthma can be effectively managed with alkaloids. Using a Soxhlet extractor, the collected plant components were first extracted with petroleum ether (to remove fats) followed by methanol. Online technologies like PASS Online, Molinspiration, SEA, SWISS, and SLAP data were used for a variety of in-silico predictions. After that, we tested the ability of Brugmansia extracts to reduce inflammation using an in-vitro approach. Rat RBC membrane lyses were stabilized using the plant extract fractions. The outcomes were also compared with aspirin (a common anti-inflammatory drug). Bioinformatics techniques were used to detect structural activity. After investigating all the data It was shown that B. sauveolens had anti-asthmatic activity.

Comparative determination of total antioxidant effects of ethanol extract of Phyllanthus amarus leaves

Abstract. Yakubu OE, Abu MS, Akighir J, Onuche JI, Arabi A. 2021. Comparative determination of total antioxidant effects of ethanol extract of Phyllanthus amarus leaves. Asian J Nat Prod Biochem 19: 81-85. Some medicinal and therapeutic potency have been reported of Phyllanthus in traditional settings. Hence, the current study evaluated the total antioxidant potential, phenolics, and flavonoids of fractions of ethanol extract of Phyllanthus amarus leave obtained using column chromatography. Extraction was done using absolute ethanol; elution was performed using different organic solvents of varying polarities, from n-Hexane, Chloroform, Ethylacetate, Ethanol, Methanol, and finally, water. The results obtained from the study revealed that fraction 10b (83 mg/mL) has the highest concentration of total Antioxidant Capacity (TAC), the next being 1a, 5a, and 11b (48mg/mL). The reduction in the TAC of the fractions was in the order; of 3b>7a>6a>5b>4a> with F7b (13mg/mL) having the lowest TAC. The result for Total phenolic Content (TPC) shows that fraction 10b (430 mg/mL) has the highest concentration of TPC, just like in antioxidant activity, followed by F4b (428 mg/mL). Fraction 2a (9 mg/mL) has the lowest concentration of TPC. Total Flavonoid Content (TFC) showed that fraction 3b (201 mg/mL) has the most concentration of total flavonoids, followed by 4b (120mg/mL). Meanwhile, fractions 1a, 2b, and 9a have the same TFC. While fraction 2a showed the lowest flavonoid concentration (25mg/mL). The plant extract fractions were found to contain more both phenolics and flavonoids, which may have contributed to the antioxidant activity observed.

Creating optimized peptide libraries for AMP discovery via PepSAVI-MS.

Antimicrobial peptides (AMPs) are host defense peptides with a range of functions/activities/modes of action that are ubiquitously expressed across all forms of life. Continued discovery of novel AMPs presents exciting opportunities to address evolving resistance to existing treatments in multiple fields, including agricultural pathogens/pests as well as antimicrobial and chemotherapeutics for human health. However, typical discovery methods including bioassay-guided fractionation and genome mining generally lack the capacity for robust AMP discovery in non-model/unsequenced organisms. PepSAVI-MS (Statistically guided bioactive peptides prioritized via mass spectrometry) was developed as an AMP discovery approach that addresses some of the limitations associated with these standard methods. PepSAVI-MS is a multi-pronged pipeline that includes peptide library creation, bioactivity screening, LC-MS analysis, and statistical modeling for putative AMP identification. The original implementation of PepSAVI-MS outlined strategies for the fractionation of plant extracts with strong cation exchange chromatography (SCX). Herein, we elaborate on recent improvements to peptide library creation through the use of orthogonal fractionation methods, specifically crude SCX chromatography and reversed-phase liquid chromatography (RPLC). This optimization of the "peptide library creation" step has demonstrated improvements for processing and AMP identifications via PepSAVI-MS.

A Photoactivable Natural Product with Broad Antiviral Activity against Enveloped Viruses, Including Highly Pathogenic Coronaviruses.

ABSTRACTThe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreak has highlighted the need for broad-spectrum antivirals against coronaviruses (CoVs). Here, pheophorbide a (Pba) was identified as a highly active antiviral molecule against human CoV-229E after bioguided fractionation of plant extracts. The antiviral activity of Pba was subsequently shown for SARS-CoV-2 and Middle East respiratory syndrome coronavirus (MERS-CoV), and its mechanism of action was further assessed, showing that Pba is an inhibitor of coronavirus entry by directly targeting the viral particle. Interestingly, the antiviral activity of Pba depends on light exposure, and Pba was shown to inhibit virus-cell fusion by stiffening the viral membrane, as demonstrated by cryoelectron microscopy. Moreover, Pba was shown to be broadly active against several other enveloped viruses and reduced SARS-CoV-2 and MERS-CoV replication in primary human bronchial epithelial cells. Pba is the first described natural antiviral against SARS-CoV-2 with direct photosensitive virucidal activity that holds potential for COVID-19 therapy or disinfection of SARS-CoV-2-contaminated surfaces.

Anti-Arthritic and Anti-Inflammatory Potential of Spondias mangifera Extract Fractions: An In Silico, In Vitro and In Vivo Approach.

The fruits of Spondias mangifera (S. mangifera) have traditionally been used for the management of rheumatism in the northeast region of India. The present study explores the probable anti-arthritis and anti-inflammatory potential of S. mangifera fruit extract’s ethanolic fraction (EtoH-F). To support this study, we first approached the parameters in silico by means of the active constituents of the plant (beta amyrin, beta sitosterol, oleonolic acid and co-crystallised ligands, i.e., SPD-304) via molecular docking on COX-1, COX-2 and TNF-α. Thereafter, the absorption, distribution, metabolism, excretion and toxicity properties were also determined, and finally experimental activity was performed in vitro and in vivo. The in vitro activities of the plant extract fractions were evaluated by means of parameters like 1,1-Diphenyl-2- picrylhydrazyl (DPPH), free radical-reducing potential, albumin denaturation, and protease inhibitory activity. The in vivo activity was evaluated using parameters like COX, TNF-α and IL-6 inhibition assay and arthritis score in Freund Adjuvant (CFA) models at a dose of 400 mg/kg b.w. per day of different fractions (hexane, chloroform, alcoholic). The molecular docking assay was performed on COX-1, COX-2 and TNF-α. The results of in vitro studies showed concentration-dependent reduction in albumin denaturation, protease inhibitors and scavenging activity at 500 µg/mL. Administration of the S. mangifera alcoholic fraction at the abovementioned dose resulted in a significant reduction (p < 0.01) in arthritis score, paw diameters, TNF-α, IL-6 as compared to diseased animals. The docking results showed that residues show a critical binding affinity with TNF-α and act as the TNF-α antagonist. The alcoholic fraction of S. mangifera extract possesses beneficial effects on rheumatoid arthritis as well as anti-inflammatory potential, and can further can be used as a possible agent for novel target-based therapies for the management of arthritis.

Seasonal antimicrobial activity of volatile substances emitted by the representatives of &lt;i&gt;Begonia&lt;/i&gt; L. genus (Begoniaceae)

The paper continues multi-year integrated studies of the numerous taxons of tropical Begonia genus, which allowed us to evaluate the adaptive capacity of the representatives of this genus relying on the revealed morphological, rhythmological and biochemical features. The investigation of antimicrobial activity of intact plants and different fractions of plant extracts allowed us to distinguish several promising representatives of this genus exhibiting activity against a broad range of test microorganisms. The involvement of these plants within phytomodules in child care centers caused a substantial decrease in total microbial contamination of the air. The paper contains results of the experimental investigation of seasonal antimicrobial activity of 13 species, 8 hybrids. The phytoncide activity was measured by exposing the streak cultures of microbial test species bacteria Staphylococcus epidermidis, Esсherichia coli and yeast-like fungi Candida albicans to the volatile emissions of plants. It has been established that 95% of begonias are distinguished by the pronounced activity against S. epidermidis bacteria, 48% against E. coli bacteria and 43% against fungi Candida albicans. The seasonal specificity of the antimicrobial action of begonias was detected: the growth of the colonies of S. epidermidis bacteria is inhibited to the highest extent during spring and summer, at the phase of intense growth, while the activity against E. coli is the highest during autumn and winter at the phase of moderate growth. The taxons with clearly pronounced, long-term and universal antimicrobial action to the studied test objects were revealed and recommended for practical planting in various types of indoor environments. The data obtained in the study may serve as the basis for further investigation of the chemical composition of volatile exometabolites by the example of the representatives of Gireoudia and Coelocentrum sections that have exhibited high antimicrobial activity against S. epidermidis and E. coli.

Antidiabetic profiling, cytotoxicity and acute toxicity evaluation of aerial parts of Phragmites karka (Retz.)

Ethnopharmacological relevancePhragmites karka (Retz.) of family Poaceae is a pristine tropical plant that is well known to the local healers for ailments of diabetes, fever, diarrhea and CNS depression but lacks the scientific evidence behind its traditional usage. Hence, we explicated this plant to find the scientific basis of its traditional utilization. Aim of the studyThe current study aims to find out the antidiabetic potential, toxicity after oral administration and in vitro cytotoxic activity of aerial parts of the plant on HeLa cells. MethodsThe plant was extracted with methanol by maceration and the crude extract was then subjected to solvent partitioning with modified Kupchan method for preparing several fractions. Phytochemical screening and total phenolic content of the plant was first determined through established procedures. Acute toxicity of the plant was studied by orally administering a single high dose (5000 mg/kg) of drug. Cytotoxicity of the methanolic plant extract was determined by measuring the percentage of cell viability on human cervical cancer cell lines, HeLa. In vitro antidiabetic activity was determined through iodine starch and DNSA (3,5-dinitrosalicylic acid) method of α-amylase inhibition. Finally, in vivo oral glucose tolerance test and alloxan induced antidiabetic activity test was performed at 150 and 300 mg/kg body weight doses of plant extract to confirm the in vivo antidiabetic activity. ResultsNo mortality was demonstrated by Phragmites karka in the acute toxicity test. However, signs of cellular toxicity was observed and histopathological studies on major organs exhibited necrosis in liver and kidney. In vitro cytotoxicity assay revealed the death of HeLa cells by DCM (dichloromethane) and n-hexane fractions of plant extract at 100 and 10 μg/mL concentrations. The IC50 value of the fractions were later evaluated by MTT assay (316.1 and 96.7 μg/mL for n-hexane and DCM fractions, respectively). In the iodine starch and DNSA method of α-amylase enzyme inhibitory activity test, substantial inhibition of enzyme was observed with the IC50 values of 2.05 and 2.08 mg/mL, respectively. In the in vivo antidiabetic activity test, considerable reduction in blood glucose level of diabetic mice was detected in both oral glucose tolerance test and alloxan induced antidiabetic activity test. In addition, the microscopic examination of pancreas showed noticeable recovery of pancreatic β cells and the blood lipid profile analysis represented a significant (p < 0.05) reduction of total cholesterol, LDL (low density lipoprotein) and triglyceride level in plant extract treated mice. ConclusionResults of this study reveals that the Phragmites karka extract is toxic at cellular level after oral administration and cytotoxic when tested on HeLa cells. The plant also evidenced hypoglycemic property, possibly through the inhibition of α-amylase enzyme and recovered the pancreatic beta cells along with the improvement of lipid profile of diabetic mice. However, robust studies on this plant is required to isolate the bioactive compounds, elucidate structures and evaluate their mechanism of actions in support of our findings. ClassificationToxicology and Safety, Quality Traditional Medicine.

Marshmallow root extract and honey: cough relief through mucoprotection

The experimental study investigated the bioadhesive properties of cough syrups containing partial fractions of plant extracts with or without added honey. The study authors established that addition of honey prolonged adhesion time on the mucosa in the model, thereby improving the physicochemical relief of irritation. As an additional soothing action, the tested syrups also reduced mucosal permeability to noxious substances and here too, the addition of honey contributed to the overall effect.

Marshmallow root extract and honey: cough relief through mucoprotection

The experimental study investigated the bioadhesive properties of cough syrups containing partial fractions of plant extracts with or without added honey. The study authors established that addition of honey prolonged adhesion time on the mucosa in the model, thereby improving the physicochemical relief of irritation. As an additional soothing action, the tested syrups also reduced mucosal permeability to noxious substances and here too, the addition of honey contributed to the overall effect.

PHYTOCHEMICAL ANALYSIS, ANTIMICROBIAL AND ANTIOXIDANT ACTIVITIES OF THYMUS VULGARIS L.

Medicinal activities of Thymus vulgaris L. were studied in the underlying research which was conducted in University of Central Punjab, Lahore. The powdered plant sample was collected and subjected to extraction and filtration by dissolving it in seven different solvents and thus the fractions obtained, were used for further analysis. The qualitative and quantitative analysis was carried out which revealed that this plant contained many biologically active compounds like saponin, alkaloids etc. These compounds have enormous bioactive roles. Antimicrobial activity was also determined in which four bacterial strains were used and the susceptibility of all the fractions of plant extract was determined by their inhibition zones which were calculated in mm. The results showed that water and chloroform extracts of plant has highest activity. The antioxidant activity was also determined by DPPH scavenging assay for which different concentrations of all plant extracts were made and their absorbance was noted to measure the percentage inhibition of each concentration, which revealed that this plant has significant antioxidant activity. The results have been represented using statistical approach that revealed that T. vulgaris plant has significant antibacterial and anti-oxidant activities and phytochemical analysis also proved that it has many useful compounds which have voluminous biological activity and these could be used in the formation of novel drugs.

Potential Nutraceutical Properties of Leaves from Several Commonly Cultivated Plants.

Chronic dietary ingestion of suitable phytochemicals may assist with limiting or negating neurodegenerative decline. Current therapeutics used to treat Alzheimer disease elicit broad adverse drug reactions, and alternative sources of cholinesterase inhibitors (ChEIs) are required. Herein, we screened methanolic extracts from seven commonly cultivated plants for their nutraceutical potential; ability to inhibit acetylcholinesterase (AChE) and butyryl-cholinesterase (BuChE), and provision of antioxidant activity through their 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) free radical scavenging capabilities. Potential neurotoxicity of plant extracts was examined via application to SHSY-5Y neuroblastoma cells and quantitation of cell viability. Methanolic extracts of Citrus limon (Lemon), Bombax ceiba (Red silk-cotton), Lawsonia inermis (Henna), Eucalyptus globulus (Eucalyptus), Ocimum basilicum (Basil), Citrus reticulata (Mandarin orange), and Mentha spicata (Spearmint) all displayed concentration-dependent inhibition of AChE and BuChE. The majority of extracts inhibited AChE and BuChE to near equipotency, with Henna and Eucalyptus extracts the two most potent ChEIs. All plant extracts were able to scavenge free radicals in a concentration-dependent manner, with Eucalyptus the most potent antioxidant. Toxicity of plant extracts to neuronal cells was concentration dependent, with Eucalyptus also the most toxic extract. Fractionation of plant extracts and analysis by mass spectrometry identified a number of plant polyphenols that might have contributed to the cholinesterase inhibition: 3-caffeoylquinic acid, methyl 4-caffeoylquinate, kaempferol-acetyl-glycoside, quercetin 3-rutinoside, quercetin-acetyl-glycoside, kaempferol 3-O-glucoside, and quercetin 3-O-glucoside. In silico molecular modeling of these polyphenols demonstrated their improved AChE and BuChE binding affinities compared to the current FDA-approved dual ChEI, galantamine. Collectively, all the plant extracts contained nutraceutical agents as antioxidants and ChEIs and, therefore, their chronic consumption may prove beneficial to combat the pathological deficits that accrue in Alzheimer disease.

Plasmodium falciparum purine nucleoside phosphorylase as a model in the search for new inhibitors by high throughput screening

Studies have shown that inhibition of Plasmodium falciparum Purine Nucleoside Phosphorylase (PfPNP) blocks the purine salvage pathway in vitro and in vivo. In this study, PfPNP was evaluated as a model in the search for new inhibitors using surface plasmon resonance (SPR). Its expression, purification, oligomeric state, kinetic constants, calorimetric parameters and kinetic mechanisms were obtained. PfPNP was immobilized on a CM5 sensor chip and sensorgrams were produced through binding the enzyme to the substrate MESG and interactions between molecules contained in 10 fractions of natural extracts. The oligomeric state showed that recombinant PfPNP is a hexamer. The true steady-state kinetic parameters for the substrate inosine were: KM 17 μM, kcat 1.2 s−1, VMax 2.2 U/mg and kcat/KM 7 × 10−4; for MESG they were: KM 131 μM, kcat 2.4 s−1, VMax 4.4 U/mg and kcat/KM 1.8 × 10−4. The thermodynamic parameters for the substrate Phosphate were: ΔG − 5.8 cal mol−1, ΔH − 6.5 cal mol−1 and ΔS − 2.25 cal mol−1/degree. The ITC results demonstrated that the binding of phosphate to free PfPNP led to a significant change in heat and association constants and thermodynamic parameters. A sequential ordered mechanism was proposed as the kinetic mechanism. Three plant extracts contained molecules capable of interacting with PfPNP, showing different levels of affinity. The identification of plant extract fractions containing molecules that interact with recombinant PfPNP using SRP validates this target as a model in the search for new inhibitors. In this study, we showed for the first time the true steady-state kinetic parameters for reactions catalyzed by PfPNP and a model using PfPNP as a target for High-throughput Screening for new inhibitors through SPR. This knowledge will allow for the development of more efficient research methods in the search for new drugs against malaria.

Hypercarb™: Unique Retentivity

Porous graphitic carbon HPLC packing material developed by John Knox’s research group and commercialised as Hypercarb™ has been available on the market for over thirty years. However, the unique properties of this phase still prove to be fundamentally novel and provide solutions to difficult separation applications. Of these properties, its lack of an elutropic series and hence ability to provide good retention not just in reversed-phase HPLC but even with non-aqueous solvents was put to good effect in this work. Herein it was demonstrated, the resolution of permethrin isomers, the isolation of coal tar extract constituents, the separation of a homologous series of garlic oil constituents, the clean-up of plant extract fractions and the provision of orthogonal selectivity for the separation of flurbiprofen and its related substances.

Plant expression of NifD protein variants resistant to mitochondrial degradation

To engineer Mo-dependent nitrogenase function in plants, expression of the structural proteins NifD and NifK will be an absolute requirement. Although mitochondria have been established as a suitable eukaryotic environment for biosynthesis of oxygen-sensitive enzymes such as NifH, expression of NifD in this organelle has proven difficult due to cryptic NifD degradation. Here, we describe a solution to this problem. Using molecular and proteomic methods, we found NifD degradation to be a consequence of mitochondrial endoprotease activity at a specific motif within NifD. Focusing on this functionally sensitive region, we designed NifD variants comprising between one and three amino acid substitutions and distinguished several that were resistant to degradation when expressed in both plant and yeast mitochondria. Nitrogenase activity assays of these resistant variants in Escherichia coli identified a subset that retained function, including a single amino acid variant (Y100Q). We found that other naturally occurring NifD proteins containing alternate amino acids at the Y100 position were also less susceptible to degradation. The Y100Q variant also enabled expression of a NifD(Y100Q)-linker-NifK translational polyprotein in plant mitochondria, confirmed by identification of the polyprotein in the soluble fraction of plant extracts. The NifD(Y100Q)-linker-NifK retained function in bacterial nitrogenase assays, demonstrating that this polyprotein permits expression of NifD and NifK in a defined stoichiometry supportive of activity. Our results exemplify how protein design can overcome impediments encountered when expressing synthetic proteins in novel environments. Specifically, these findings outline our progress toward the assembly of the catalytic unit of nitrogenase within mitochondria.

Phytochemical-assisted biosynthesis of silver nanoparticles from Ajuga bracteosa for biomedical applications

Silver nanoparticles (AgNPs) synthesized from plant extracts are widely used for the cure of many diseases from fever to cancers. Keeping in view the medicinal value of AgNPs, here we report a cost-effective phytochemical method for the biosynthesis of silver nanoparticles from Ajuga bracteosa. A. bracteosa is an important medicinal plant used to cure fever, appetite-loss, and cancer. Silver-nanoparticles were prepared from the aqueous extract of the plant. The methanolic extract of A. bracteosa (ABMF) was separated and n-hexane (ABHF) and chloroform (ABCF) fractions were obtained from the methanolic crude extract. The AgNPs were characterized by UV-Visible spectrophotometer, FTIR, XRD, and TEM. The total phenolic contents (TPC) and total flavonoid contents (TFC) in different fractions were determined and compared with AgNPs. The medicinal value of ABMF, ABHF, ABCF, and AgNPs was evaluated by antibacterial, antioxidant, anti-inflammatory, and cytotoxicity bioassays. The UV-visible spectrum showed a peak at 484 nm while FTIR results suggested strong capping of phytochemicals on AgNPs which was confirmed by a high amount of TPC and TFC. XRD analysis depicted a high degree of crystallinity and smaller size of AgNPs. TEM results showed spherical shaped AgNPs of size range 50 ± 12 nm. The biosynthesized AgNPs showed better antibacterial activity than plant extract fractions. Similarly, AgNPs have shown better antioxidant, cytotoxicity against cancer cell lines in-vitro, and anti-inflammatory activity in-vivo than a plant extract. The great medicinal value of A. bracteosa might be due to the presence of pharmacologically active phytochemicals such as diterpenoids, neo-clerodane flavonol glycosides, ergosterol, iridoid glycosides, phytoecdysones, and other polyphenols. These phytochemicals surround the silver nanoparticles during green synthesis and therefore, this capping of phytochemicals over silver nanoparticles results in enhanced biomedical applications of plant extracts.

Macrophylloflavone: A New Biflavonoid from Garcinia macrophylla Mart. (Clusiaceae) for Antibacterial, Antioxidant, and Anti-Type 2 Diabetes Mellitus Activities.

Investigations of antibacterial, antioxidant, and anti-type 2 diabetes mellitus activities have been carried out on Garcinia macrophylla Mart. plant extract fractions. An isolate from a fraction of ethyl acetate extract was characterized with spectroscopic data. A new biflavonoid compound was found to have a skeleton of 5,7,4′,5″,7″,3‴,4‴-heptahydroxyflavanone[3-6″] flavones which was named macrophylloflavone (1). The compound was evaluated for its antibacterial activity against Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923 with cephazolin as a positive control, antioxidant assay against 2,2 diphenyl-1-picrylhydrazyl (DPPH) with ascorbic acid as the positive control, and anti-type 2 diabetes mellitus treatment with metformin as a positive control. The biflavonoid compound exhibited a good inhibition for bacteria and free radical DPPH. Furthermore, biflavonoid compound treatment on the diabetic rats suggested its ability to decrease the blood glucose level. This study provided evidence that the plant has antibacterial, antioxidant, and antidiabetic properties.