Preparation and characterization of biodegradable food packaging films using lemon peel pectin and chitosan incorporated with neem leaf extract and its application on apricot fruit.

The present study aims to develop and characterize biodegradable packaging films from lemon peel-derived pectin and chitosan incorporated with a bioactive extract from neem leaves. The films (PCNE) contained varying concentrations of neem leaf extract and were comprehensively assessed for their physical, optical, mechanical, and antimicrobial attributes. The thickness, moisture content, water solubility, and water vapor permeability of the biodegradable packaging films increased with the increasing concentration of neem leaf extract. Comparatively, the tensile strength of the films decreased by 42.05 % compared to the control film. The Scanning Electron Microscopy (SEM) confirmed that the resultant blended pectin-chitosan films showed a uniform structure without cracks. Furthermore, the analysis targeting Staphylococcus aureus and Aspergillus niger indicated that the films had potent antimicrobial activity. Based on these results, the optimum films were selected and subsequently applied on apricot fruits to increase their shelf life at ambient temperature. The findings, after examining factors such as colour, firmness, total soluble solids, shrinkage, weight loss, and appearance, concluded that the apricots coated by PCNE-5 had the most delayed signs of spoilage and increased their shelf life by 50 %. The results showed the potential applicability of lemon peel pectin-chitosan-neem leaf extract blend films in biodegradable food packaging.

Evolving therapeutic interventions for the management and treatment of Alzheimer's disease.

Alzheimer's Disease (AD) patients experience diverse symptoms, including memory loss, cognitive impairment, behavioral abnormalities, mood changes, and mental issues. The fundamental objective of this review is to discuss novel therapeutic approaches, with special emphasis on recently approved marketed formulations for the treatment of AD, especially Aducanumab, the first FDA approved moiety that surpasses the blood-brain barrier (BBB) and reduces amyloid plaques in the brain, thereby reducing associated cognitive decline. However, it is still in the phase IV trial and is to be completed by 2030. Other drugs such as lecanemab are also under clinical trial and has recently been approved by the FDA and is also discussed here. In this review, we also focus on active and passive immunotherapy for AD as well as several vaccines, such as amyloid-beta epitope-based vaccines, amyloid-beta DNA vaccines, and stem cell therapy for AD, which are in clinical trials. Furthermore, ongoing pre-clinical trials associated with AD and other novel strategies such as curcumin-loaded nanoparticles, Crispr/ cas9, precision medicine, as well as some emerging therapies like anti-sense therapy are also highlighted. Additionally, we discuss some off-labeled drugs like non-steroidal anti-inflammatory drugs (NSAID), anti-diabetic drugs, and lithium, which can manage symptoms of AD and different non-pharmacological approaches are also covered which can help to manage AD. In summary, we have tried to cover all the therapeutic interventions which are available for the treatment and management of AD under sections approved, clinical phase, pre-clinical phase or futuristic interventions, off-labelled drugs, and non-pharmacological interventions for AD, offering positive findings and well as challenges that remain.

A Spectrum of Solutions: Unveiling Non-Pharmacological Approaches to Manage Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a developmental disorder that causes difficulty while socializing and communicating and the performance of stereotyped behavior. ASD is thought to have a variety of causes when accompanied by genetic disorders and environmental variables together, resulting in abnormalities in the brain. A steep rise in ASD has been seen regardless of the numerous behavioral and pharmaceutical therapeutic techniques. Therefore, using complementary and alternative therapies to treat autism could be very significant. Thus, this review is completely focused on non-pharmacological therapeutic interventions which include different diets, supplements, antioxidants, hormones, vitamins and minerals to manage ASD. Additionally, we also focus on complementary and alternative medicine (CAM) therapies, herbal remedies, camel milk and cannabiodiol. Additionally, we concentrate on how palatable phytonutrients provide a fresh glimmer of hope in this situation. Moreover, in addition to phytochemicals/nutraceuticals, it also focuses on various microbiomes, i.e., gut, oral, and vaginal. Therefore, the current comprehensive review opens a new avenue for managing autistic patients through non-pharmacological intervention.

Structure-based investigation of MARK4 inhibitory potential of Naringenin for therapeutic management of cancer and neurodegenerative diseases.

MAP/microtubule affinity-regulating kinase 4 (MARK4) is a member of serine/threonine kinase family and considered an attractive drug target for many diseases. Screening of Indian Medicinal Plants, Phytochemistry, and Therapeutics (IMPPAT) using virtual high-throughput screening coupled with enzyme assay suggested that Naringenin (NAG) could be a potent inhibitor of MARK4. Structure-based molecular docking analysis showed that NAG binds to the critical residues found in the active site pocket of MARK4. Furthermore, molecular dynamics (MD) simulation studies for 100 ns have delineated the binding mechanism of NAG to MARK4. Results of MD simulation suggested that binding of NAG further stabilizes the structure of MARK4 by forming a stable complex. In addition, no significant conformational change in the MARK4 structure was observed. Fluorescence binding and isothermal titration calorimetric measurements revealed an excellent binding affinity of NAG to MARK4 with a binding constant (K) = 0.13 × 106 M-1 obtained from fluorescence binding studies. Further, enzyme inhibition studies showed that NAG has an admirable IC50 value of 4.11 µM for MARK4. Together, these findings suggest that NAG could be an effective MARK4 inhibitor that can potentially be used to treat cancer and neurodegenerative diseases.

Purification, modeling and structural insights of calmodulin-binding receptor like cytoplasmic kinase 2 from Oroxylum Indicum.

Calmodulin binding receptor like cytoplasmic kinase 2 (CRCK2) belongs to the family of receptor like kinases (RLKs) which is mainly implicated in pathways associated with the stress responses in plants. The protein from the stem of Oroxylum indicum was isolated and purified using anion-exchange followed by gel filtration chromatography. The purity of protein was checked using SDS-PAGE, which showed a single band of 50 kDa. The purified protein was identified as CRCK2 using MALDI-TOF. Using I-TASSER, a bioinformatics tools, the model of protein was constructed and its secondary structure was predicted using VADAR. The secondary structure content was also determined by far-UV CD, which indicated that the CRCK2 is mainly ß-sheet dominating protein (43% ß-sheet). The secondary structural content predication from computational method is in close agreement with the result obtained by CD spectropolarimeter. This study validates I-TASSER model for determination of structure of a protein. Moreover, stability of CRCK2 was monitored against heat- and guanidinium chloride (GdmCl)-induced denaturation by using circular dichroism (CD) and fluorescence spectroscopy. Denaturation curve analysis gave values of 2.88 ±â€¯0.12 kcal mol-1and 4.11 ±â€¯0.09 M for ∆°GD (Gibbs free energy change at 25 °C) and Cm (midpoint of denaturation), respectively. It has been observed that purified CRCK2 is quite stable protein against both heat-induced as well as GdmCl-induced denaturation. This is very first report of purification and biophysical characterization of CRCK2 protein from medicinal plant O. indicum.

Investigation of molecular mechanism of recognition between citral and MARK4: A newer therapeutic approach to attenuate cancer cell progression.

Microtubule affinity regulating kinase 4 (MARK4) is a member of AMP-activated protein kinase, found to be involved in apoptosis, inflammation and many other regulatory pathways. Since, its aberrant expression is directly associated with the cell cycle and thus cancer. Therefore, MARK4 is being considered as a potential drug target for cancer therapy. Here, we investigated the mechanism of inhibition of MARK4 activity by citral. Docking studies suggested that citral effectively binds to the active site cavity, and complex is stabilized by several interactions. We further performed molecular dynamics simulation of MARK4-citral complex under explicit water condition for 100ns and observed that binding of citral to MARK4 was quite stable. Fluorescence binding studies suggested that citral strongly binds to MARK4 and thereby inhibits its enzyme activity which was measured by the kinase inhibition assay. We further performed MTT assay and observed that citral inhibits proliferation of breast cancer cell line MCF-7. This work provides a newer insight into the use of citral as novel cancer therapeutics through the MARK4 inhibition. Results may be employed to design novel therapeutic molecule using citral as a scaffold for MARK4 inhibition to fight related diseases.

Elucidation of Dietary Polyphenolics as Potential Inhibitor of Microtubule Affinity Regulating Kinase 4: In silico and In vitro Studies.

Microtubule affinity regulating kinase 4 (MARK4) is a Ser/Thr kinase belonging to AMPK-like family, has recently become an important drug target against cancer and neurodegenerative disorders. In this study, we have evaluated different natural dietary polyphenolics including rutin, quercetin, ferulic acid, hesperidin, gallic acid and vanillin as MARK4 inhibitors. All compounds are primarily binds to the active site cavity of MARK4. In silico observations were further complemented by the fluorescence-binding studies and isothermal titration calorimetry (ITC) measurements. We found that rutin and vanillin bind to MARK4 with a reasonably high affinity. ATPase and tau-phosphorylation assay further suggesting that rutin and vanillin inhibit the enzyme activity of MARK4 to a great extent. Cell proliferation, ROS quantification and Annexin-V staining studies are clearly providing sufficient evidences for the apoptotic potential of rutin and vanillin. In conclusion, rutin and vanillin may be considered as potential inhibitors for MARK4 and further exploited to design novel therapeutic molecules against MARK4 associated diseases.

Therapeutic progress in amyotrophic lateral sclerosis-beginning to learning.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease associated with motor neuron degeneration, muscle weakness, paralysis and finally death. The proposed mechanisms of ALS include glutamate excitotoxicity, oxidative stress, inflammation, mitochondrial dysfunction, apoptosis and proteasomal dysfunction. Although numerous pathological mechanisms have been explained, ALS remains incurable disease because of failure of clinical trials and lack of any effective therapy. The rapid advancement in genetic discoveries in ALS emphasizes the point that ALS is a multi-subtype syndrome rather than a single disease. This can be argued as one of the single reason why many previous therapeutic drug trials have failed. Efforts to develop novel ALS treatments which target specific pathomechanisms are currently being pursued. Herein, we review the recent discovery and preclinical characterization of neuroprotective compounds and compare their effects on disease onset, duration and survival. Furthermore, the structure-activity relationships of these agents are analyzed with the overall goal of developing a screening strategy for future clinical applications.

Purification and characterization of Ras related protein, Rab5a from Tinospora cordifolia.

Ras related protein (Rab5a) is one of the most important member of the Rab family which regulates the early endosome fusion in endocytosis, and it also helps in the regulation of the budding process. Here, for the first time we report a simple and reproducible method for the purification of the Rab5a from a medicinal plant Tinospora cordifolia. We have used weak cation-exchange (CM-Sepharose-FF) followed by gel-filtration chromatography. A purified protein of 22-kDa was observed on SDS-PAGE which was identified as Rab5a using MALDI-TOF/MS. Our purification procedure is fast and simple with high yield. The purified protein was characterized using circular dichroism for the measurement of secondary structure followed by GdmCl- and urea-induced denaturation to calculate the values of Gibbs free energy change (ΔGD), ΔGD°, midpoint of the denaturation Cm, i.e. molar GdmCl [GdmCl] and molar urea [Urea] concentration at which ΔGD=0; and m, the slope (=∂ΔGD/∂[d]) values. Furthermore, thermodynamic properties of Rab5a were also measured by differential scanning calorimeter. Here, using isothermal calorimeteric measurements we further showed that Rab5a binds with the GTP. This is a first report on the purification and biophysical characterization of Rab5a protein from T. cordifolia.

Purification and characterization of oligonucleotide binding (OB)-fold protein from medicinal plant Tinospora cordifolia.

The oligonucleotide binding fold (OB-fold) is a small structural motif present in many proteins. It is originally named for its oligonucleotide or oligosaccharide binding properties. These proteins have been identified as essential for replication, recombination and repair of DNA. We have successfully purified a protein contains OB-fold from the stem of Tinospora cordifolia, a medicinal plants of north India. Stems were crushed and centrifuged, and fraction obtained at 60% ammonium sulphate was extensively dialyzed and applied to the weak anion exchange chromatography on Hi-Trap DEAE-FF in 50mM Tris-HCl buffer at pH 8.0. Eluted fractions were concentrated and applied to gel filtration column to get pure protein. We observed a single band of 20-kDa on SDS-PAGE. Finally, the protein was identified as OB-fold by MALDI-TOF. The purified OB-fold protein was characterized for its secondary structural elements using circular dichroism (CD) in the far-UV region. Generally the OB-fold has a characteristic feature as five-stranded beta-sheet coiled to form a closed beta- barrel. To estimate its chemical stability, guanidinium chloride-induced denaturation curve was followed by observing changes in the far-UV CD as a function of the denaturant concentration. Analysis of this denaturation curve gave values of 8.90±0.25kcalmol(-1) and 3.78±0.18M for ΔGD° (Gibbs free energy change at 25°C) and Cm (midpoint of denaturation), respectively. To determine heat stability parameters of OB-fold protein, differential scanning calorimetry was performed. Calorimetric values of ΔGD°, Tm (midpoint of denaturation), ΔHm (enthalpy change at Tm), and ΔCp (constant-pressure heat capacity change) are 9.05±0.27kcalmol(-1), 85.2±0,3°C, 105±4kcalmol(-1) and 1.6±0.08kcalmol(-1)K(-1). This is the first report on the isolation, purification and characterization of OB-fold protein from a medicinal plant T. cordifolia.

Purification and characterization of RGA2, a Rho2 GTPase-activating protein from Tinospora cordifolia.

Rho GTPases activating protein 2 (RGA2) is primarily involved in the modulation of numerous morphological events in eukaryotes. It protects plants by triggering the defense system which restricts the pathogen growth. This is the first report on the isolation, purification and characterization of RGA2 from the stems of Tinospora cordifolia, a medicinal plant. The RGA2 was purified using simple two-step process using DEAE-Hi-Trap FF and Superdex 200 chromatography columns, with a high yield. The purity of RGA2 was confirmed by SDS-PAGE and identified by MALDI-TOF/MS. The purified protein was further characterized for its secondary structural elements using the far-UV circular dichroism measurements. Our purification procedure is simple two-step process with high yield which can be further used to produce RGA2 for structural and functional studies.