A lyophilized surfactant-based rutin formulation with improved physical characteristics and dissolution for oral delivery.

Rutin (RUT) is a phytochemical flavonoid with numerous therapeutic potentials including antihypertension, cardioprotective, neuroprotective, and anti-cancer activities. Its clinical use is inhibited due to its poor aqueous solubility and permeability over oral administration. The present study aimed to overcome these problems through micellization and entrapment of RUT in solid dispersion (SD) using Poloxamer (POL) 407 and 188 as surfactant-based matrices. The RUT/SD formulations were prepared in serial drug loading concentrations in weight percentage to the total solid. The physical properties of the formed RUT/SD solids were characterized by several methods including polarizing microscopy, differential thermal analysis (DTA), X-ray diffractometry (XRD), scanning electron microscopy (SEM) and dissolution study. The dissolution test was performed using a paddle dissolution apparatus and samples were analyzed using UV spectrophotometry. Polarized microscope confirmed that the optical behaviors of the RUT/SD implied a formation of miscible RUT with POL matrices. The morphology of RUT/SDs varied from porous matrices with craters to smoother surfaces as a function of RUT concentrations. XRD and DTA data exhibited that RUT existed as partially amorphous. These data indicated that the higher concentration of RUT in the RUT/SD formulations, the higher amorphous proportion of the RUT in the solid state. Henceforth, this led to an increase in the percentage of dissolved RUT from the developed RUT/SD formulations at 94 to 100% compared to pure RUT at only < 35% within an hour. The present study disclosed the successful improvements in the physical characteristics of the RUT/SD formulations and their potencies for the future development for oral formulation.

Development and validation of a HILIC-HPLC-ELSD method for simultaneous determination of glucosamine hydrochloride and chondroitin sulfate in dietary supplements

Abstract The objective of the present study is to develop and validate a simple, selective and accurate hydrophilic interaction liquid chromatography - a high performance liquid chromatography incorporating an evaporative light scattering detector (HILIC-HPLC-ELSD) method for simultaneously determining glucosamine hydrochloride and chondroitin sulfate in dietary supplements. The chromatographic separation was carried out on a ZIC-HILIC column (150 mm x 4.6 mm x 5µm) in isocratic system mode with a mobile phase of acetonitrile, 30 mM ammonium formate and water (77:20:3, v/v/v) at pH 4.5, a column temperature of 35°C, a flow rate of 1 mL.min-1, and an injection volume of 5 µL. An evaporative light scattering (ELS) detector was used. Effective separation was achieved by means of analyte resolution of more than 1.5 with an analysis run time of approximately 20 minutes. The linearity of glucosamine hydrochloride and chondroitin sulfate ranged from 0.4 to 2.5 mg.mL-1. The limits of the detection and quantification of glucosamine hydrochloride were 20 and 80 mg.mL-1 respectively, while for chondroitin sulfate they were 80 and 400 mg.mL-1. All validation parameters satisfied the acceptance criteria in accordance with International Conference on Harmonisation (ICH) guidelines. The method was successfully applied to the assay of commercial dietary supplement samples

GC-MS- and NMR-Based Metabolomics and Molecular Docking Reveal the Potential Alpha-Glucosidase Inhibitors from Psychotria malayana Jack Leaves.

Psychotria malayana Jack leaf, known in Indonesia as "daun salung", is traditionally used for the treatment of diabetes and other diseases. Despite its potential, the phytochemical study related to its anti-diabetic activity is still lacking. Thus, this study aimed to identify putative inhibitors of α-glucosidase, a prominent enzyme contributing to diabetes type 2 in P. malayana leaf extract using gas chromatography-mass spectrometry (GC-MS)- and nuclear magnetic resonance (NMR)-based metabolomics, and to investigate the molecular interaction between those inhibitors and the enzyme through in silico approach. Twenty samples were extracted with different solvent ratios of methanol-water (0, 25, 50, 75, and 100% v/v). All extracts were tested on the alpha-glucosidase inhibition (AGI) assay and analyzed using GC-MS and NMR. Multivariate data analysis through a partial least square (PLS) and orthogonal partial square (OPLS) models were developed in order to correlate the metabolite profile and the bioactivity leading to the annotation of the putative bioactive compounds in the plant extracts. A total of ten putative bioactive compounds were identified and some of them reported in this plant for the first time, namely 1,3,5-benzenetriol (1); palmitic acid (2); cholesta-7,9(11)-diene-3-ol (3); 1-monopalmitin (4); ß-tocopherol (5); α-tocopherol (6); 24-epicampesterol (7); stigmast-5-ene (8); 4-hydroxyphenylpyruvic acid (10); and glutamine (11). For the evaluation of the potential binding modes between the inhibitors and protein, the in silico study via molecular docking was performed where the crystal structure of Saccharomyces cerevisiae isomaltase (PDB code: 3A4A) was used. Ten amino acid residues, namely ASP352, HIE351, GLN182, ARG442, ASH215, SER311, ARG213, GLH277, GLN279, and PRO312 established hydrogen bond in the docked complex, as well as hydrophobic interaction of other amino acid residues with the putative compounds. The α-glucosidase inhibitors showed moderate to high binding affinities (-5.5 to -9.4 kcal/mol) towards the active site of the enzymatic protein, where compounds 3, 5, and 8 showed higher binding affinity compared to both quercetin and control ligand.

Analytical method for the determination of curcumin entrapped in polymeric micellar powder using HPLC.

OBJECTIVES: Curcumin belongs to the family of curcuminoids, natural polyphenolic compounds that possesses neuroprotective properties, anti inflammatory and anticancer. Its entrapment in the developed casein-based micellar powder (CMP) and poloxamer-based micellar powder (PMP) was to enhance the solubility and improve the bioavailability. Henceforth, the present study aimed to acquire an efficient analytical method for the curcumin analysis in polymeric micellar formulations. METHODS: A fast and specific HPLC method was developed for analyzing curcumin in two different micellar matrices using casein and poloxamer. The HPLC was equipped with a C18 column (250 × 4 mm, 5 µm) and diode array detector. A designated isocratic elution of curcumin was employed using mobile phase with a composition of water (1%, v/v acetic acid) and acetonitrile in a ratio of 50:50 v/v. The employed flow rate was 1.0 mL/min and the analyte was examined at 421 nm. RESULTS: An effective analysis in HPLC was successfully achieved by the predetermined HPLC condition. A good resolution of peaks at the employed flow rate was achieved. The linearity was excellent in two different range of concentrations, 2-20 and 10-50 µg/mL. The selectivity, accuracy and precision fulfilled the acceptable requirements. CONCLUSIONS: The developed method was practically effective to qualitatively identified curcumin. In addition, the assay also effectively quantified the amount of curcumin in the polymeric entrapping matrices which demonstrates that it has great potential to be used in natural compound analysis.

Antioxidants profile of Momordica charantia fruit extract analyzed using LC-MS-QTOF-based metabolomics.

Momordica charantia fruit is claimed to have healthy benefit. Despite this potential claim, the phytochemical study of this fruit is still lacking. Thus, this study aimed to evaluate the antioxidants profile of Momordica charantia (Cucurbitaceae) fruit. The antioxidant activity of the ethanolic extracts of various polarities was evaluated and the metabolites that are responsible for its activity were identified using metabolomics approach. Six different mixture of ethanol in water that are 0%, 20%, 40%, 60%, 80%, and 100% (v/v) was extracted using dveseeded fruit sample. Liquid chromatography-mass spectrometry-quadrupole time of flight and multivariate data analysis was used to identify the metabolites that were either antioxidants or pro-oxidants. The 80% ethanol extract exhibited the most antioxidant activity when tested in both 2, 2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) antioxidant assays. This extract showed the most intense LC-MS signals represented to ascorbic acid, margarolic acid, brevifolincarboxylic acid, quercetin 3-O-glycoside, kuguacin H, cucurbitacin E, 3-malonylmomordicin I, and goyaglycoside G correlating to the anti-oxidant activity. This study reports for the first time the existence of brevifolincarboxylic acid in this fruit, and the antioxidant activity of 3-malonylmomordicin I and goyaglycoside G. In addition, the loading plots revealed the unknown compounds possessing the antioxidant activity which are potential to be isolated in the future study.

Characterization of α-Glucosidase Inhibitors from Psychotria malayana Jack Leaves Extract Using LC-MS-Based Multivariate Data Analysis and In-Silico Molecular Docking.

Psychotria malayana Jack has traditionally been used to treat diabetes. Despite its potential, the scientific proof in relation to this plant is still lacking. Thus, the present study aimed to investigate the α-glucosidase inhibitors in P.malayana leaf extracts using a metabolomics approach and to elucidate the ligand-protein interactions through in silico techniques. The plant leaves were extracted with methanol and water at five various ratios (100, 75, 50, 25 and 0% v/v; water-methanol). Each extract was tested for α-glucosidase inhibition, followed by analysis using liquid chromatography tandem to mass spectrometry. The data were further subjected to multivariate data analysis by means of an orthogonal partial least square in order to correlate the chemical profile and the bioactivity. The loading plots revealed that the m/z signals correspond to the activity of α-glucosidase inhibitors, which led to the identification of three putative bioactive compounds, namely 5'-hydroxymethyl-1'-(1, 2, 3, 9-tetrahydro-pyrrolo (2, 1-b) quinazolin-1-yl)-heptan-1'-one (1), α-terpinyl-ß-glucoside (2), and machaeridiol-A (3). Molecular docking of the identified inhibitors was performed using Auto Dock Vina software against the crystal structure of Saccharomyces cerevisiae isomaltase (Protein Data Bank code: 3A4A). Four hydrogen bonds were detected in the docked complex, involving several residues, namely ASP352, ARG213, ARG442, GLU277, GLN279, HIE280, and GLU411. Compound 1, 2, and 3 showed binding affinity values of -8.3, -7.6, and -10.0 kcal/mol, respectively, which indicate the good binding ability of the compounds towards the enzyme when compared to that of quercetin, a known α-glucosidase inhibitor. The three identified compounds that showed potential binding affinity towards the enzymatic protein in molecular docking interactions could be the bioactive compounds associated with the traditional use of this plant.

Alpha-Glucosidase Inhibitory Effect of Psychotria malayana Jack Leaf: A Rapid Analysis Using Infrared Fingerprinting.

The plant Psychotria malayana Jack belongs to the Rubiaceae family and is known in Malaysia as "meroyan sakat/salung". A rapid analytical technique to facilitate the evaluation of the P. malayana leaves' quality has not been well-established yet. This work aimed therefore to develop a validated analytical technique in order to predict the alpha-glucosidase inhibitory action (AGI) of P. malayana leaves, applying a Fourier Transform Infrared Spectroscopy (FTIR) fingerprint and utilizing an orthogonal partial least square (OPLS). The dried leaf extracts were prepared by sonication of different ratios of methanol-water solvent (0, 25, 50, 75, and 100% v/v) prior to the assessment of alpha-glucosidase inhibition (AGI) and the following infrared spectroscopy. The correlation between the biological activity and the spectral data was evaluated using multivariate data analysis (MVDA). The 100% methanol extract possessed the highest inhibitory activity against the alpha-glucosidase (IC50 2.83 ± 0.32 µg/mL). Different bioactive functional groups, including hydroxyl (O-H), alkenyl (C=C), methylene (C-H), carbonyl (C=O), and secondary amine (N-H) groups, were detected by the multivariate analysis. These functional groups actively induced the alpha-glucosidase inhibition effect. This finding demonstrated the spectrum profile of the FTIR for the natural herb P. malayana Jack, further confirming its medicinal value. The developed validated model can be used to predict the AGI of P. malayana, which will be useful as a tool in the plant's quality control.

IN VITRO ANTIMALARIAL ACTIVITY AND TOXICITY STUDIES OF JOHAR (CASSIA SIAMEA) LEAVES FROM THREE DIFFERENT LOCATIONS.

BACKGROUND: Antimalarial activity of Cassia siamea leaves has been proven by the active compound that has been found, i.e. Cassiarin A. It is known that the quantity of the content of a compound that has the potential as a raw material for medicine can be influenced by various factors including differences in plant origin. This study aims at comparing the antimalarial activity and toxicity of C.siamea leaves from three regions with different meters location values above sea level (asl), i.e Pariaman (1,000 m asl), Palu (60 m asl), and Surabaya (2 m asl). MATERIALS AND METHODS: The materials used in this study were Johar leaves from Pariaman, Surabaya, and Palu extracted with n-hexane, and 90% ethanol containing 1% tartaric acid. The antimalarial activity test was done with Plasmodium falciparum 3D7. The toxicity test applied MTT ELISA method. RESULTS: C.siamea leaf that had highest antimalarial activity came from Pariaman with IC50 value of 0.006µg/ml, then from Palu was 0.037µg/ml, and the lowest antimalarial activity was from Surabaya that was 0.09µg/ml . In testing the toxicity to get CC50, the highest toxicity came from Surabaya with CC50 value of 135.81µg/ml, Pariaman with CC50 value of 220.82 µg/ml, and the least toxic came from Palu with CC50 value of 235.52µg/ml . CONCLUSION: C.siamea leaf obtained from Pariaman had a selectivity index value that satisfies the requirements of a promising antimalarial effect.