Implementation of different separation techniques for resolving ternary mixture of Paracetamol, Pseudoephedrine Hydrochloride and Chlorpheniramine maleate with further quantification

Due to the wide applicability of separation techniques that rely on the property of differential migration in pharmaceutical formulations analysis, different analytical strategies have been proposed to resolve mixtures of multi-components pharmaceuticals. Three separation methods were developed and validated for the simultaneous determination of Paracetamol (PAR), Pseudoephedrine HCl (PSE) and Chlorpheniramine maleate (CHP). The first method is a thin-layer chromatographic (TLC) separation, followed by densitometric measurement. The separation was carried out on aluminium sheet of silica gel 60 F254 using ethanol:chloroform:ammonia (1:7:0.4, by volume) as the mobile phase. Determination of PAR, PSE and CHP was successfully applied over the concentration ranges of 3–25 µg/band, 0.5–10 µg/band and 0.1–6 µg/band, respectively. The second method is HPLC separation that was achieved on C18 column using the mobile phase acetonitrile:phosphate buffer pH 5 (10:90, v/v) at a flow rate 1 mL min−1. PAR, PSE and CHP were determined by HPLC in concentration ranges of 5–400 μg mL−1, 2–40 μg mL−1 and 0.5–16 μg mL−1, respectively. The third method is a capillary electrophoresis (CE) separation. The electrophoretic separation was achieved using 20 mM phosphate buffer (pH 6.5) at 20 kV. The linearity was reached over concentration ranges of 30–250 μg mL−1, 5–50 μg mL−1 and 0.8–20 μg mL−1 for PAR, PSE and CHP, respectively. The developed methods were validated with respect to linearity, precision, accuracy and system suitability. The proposed methods were successfully applied for bulk powder and dosage form analysis with RSD of precision <2%. Moreover, statistical comparison with the official methods confirms the methods' validity.

Simultaneous Densitometric and UPLC Methods for the Determination of Paracetamol and Metoclopramide HCL

Two simple, accurate and precise methods were presented for the simultaneous quantitative estimation of paracetamol and metoclopramide hydrochloride. The first is based on TLC separation of the two drugs, followed by densitometric measurement at 274 nm using a developing system composed of dichloromethane - methanol - conc. ammonia (8:2:0.05 by volume). The second method is based on UPLC separation of the cited drugs at 230 nm using C18 column and a mobile phase of 0.1% ortho-phosphoric acid (PH 3.5) – acetonitrile (70: 30, v/v). Regression analysis of Beer’s plots showed good correlations (r = 0.9996 - 0.9998) over concentration ranges of 1.5 – 15 and 0.1-0.6 μg/spot or 5-25 and 0.25 – 1.5 μg mL-l for paracetamol and metoclopramide HCL using the two suggested methods, respectively. The proposed methods were also successfully applied to analyze both drugs in their pharmaceutical formulation; Migracid® tablets with mean recoveries ranging from 99.03 to 101.1%. The results obtained were validated as per ICH guidelines and statistically analyzed and found to be in accordance with those given by a reported method.

Tetrahydrocannabinol Sensing in Complex Biofluid with Portable Raman Spectrometer Using Diatomaceous SERS Substrates

Using thin-layer chromatography in tandem with surface-enhanced Raman spectroscopy (TLC-SERS) and tetrahydrocannabinol (THC) sensing in complex biological fluids is successfully conducted with a portable Raman spectrometer. Both THC and THC metabolites are detected from the biofluid of marijuana-users as biomarkers for identifying cannabis exposure. In this article, ultra-sensitive SERS substrates based on diatomaceous earth integrated with gold nanoparticles (Au NPs) were employed to detect trace levels of cannabis biomarkers in saliva. Strong characteristic THC and THC metabolite SERS peaks at 1601 and 1681 cm−1 were obtained despite the moderate interference of biological molecules native to saliva. Urine samples were also analyzed, but they required TLC separation of THC from the urine sample to eliminate the strong influence of urea and other organic molecules. TLC separation of THC from the urine was performed by porous microfluidic channel devices using diatomaceous earth as the stationary phase. The experimental results showed clear separation between urea and THC, and strong THC SERS characteristic peaks. Principal component analysis (PCA) was used to analyze the SERS spectra collected from various THC samples. The spectra in the principal component space were well clustered for each sample type and share very similar scores in the main principal component (PC1), which can serve as the benchmark for THC sensing from complex SERS spectra. Therefore, we proved that portable Raman spectrometers can enable an on-site sensing capability using diatomaceous SERS substrates to detect THC in real biological solutions. This portable THC sensing technology will play pivotal roles in forensic analysis, medical diagnosis, and public health.

PRELIMINARY PHYTOCHEMICAL EVALUATION AND SPECTRAL CHARACTERIZATION OF ACTIVE CONSTITUENTS IN THE DRIED EXTRACTS OF THE WHOLE PLANT ARGYREIA IMBRICATA (ROX) SANT AND PATEL

Objective: The present study was designed with the aim of preliminary phytochemical evaluation and spectral characterization phytochemicals present in the extracts of whole plant Argyreia imbricata (Rox) Sant and Patel. Methods: The whole plant A. imbricata was collected, authenticated, and dried in the shade for powdering in mechanical grinder. The powdered plant material was extracted with Soxhlet apparatus using different solvents, and the dried extract obtained was subjected to preliminary phytochemical evaluation, and then column chromatography separation and the separated fractions were subjected to thin-layer chromatography (TLC) separation. Based on the yield in TLC, the selected compounds were subjected to carbon-13 nuclear magnetic resonance, proton NMR (1H NMR), and mass spectroscopic evaluation. Results: In the preliminary phytochemical evaluation, the presence of alkaloids, glycosides, phenolic compounds, flavonoids, carbohydrates, saponins, sterols, and terpenoids was found in different extracts. In column chromatography separation, totally 650 fractions were collected, and four compounds were isolated with the help of TLC. Among the four, two compounds were selected for spectral evaluation based on the yield obtained. The selected compounds were identified and named as, 3-ethyl-5-(hydroxymethyl) phenol and methyl 4-hydroxybenzoate with the help of spectral evaluation. Conclusion: The outcome of this study is beneficial, and further investigation in the future directed toward the detailed pharmacological screening of the extracts may give more valuable results.

LAJU HIDROLISIS HEROIN DALAM AIR DAN PLASMA

Heroine is hydrolyzed spontaneously in water and plasma. This will influence the determination, especially on the drug profiling. Spectrophotodensitometry has been used to analyze drug profiling of illicit heroine. This article reports the AL-TLC separation of heroine, 6-monoacetylmorphine (6-MAM), morphine, acetyl codeine, and the heroine hydrolysis in water and plasma. Heroin, 6-MAM, morphine, and acetyl codeine can be well-separated by mobile phase of toluene:siclohexane:diethyl amine (75:15:10, v/v). The limit of detection was 165.16 ng/spot and the limit of quantification was 550.55 ng/spot. Heroine was hydrolyzed in water and plasma under first order reaction. The rate of reaction was 0.55 min-1 in water with the half time reaction of 12.47 minutes. On the other hand the hydrolysis rate in plasma was slower with the half time of 16 minutes. Keywords : hydrolysis, heroin, water, TLC-spectrophotodensitometer

Isolation of novel quorum-sensing active bacteria from microbial mats in Shark Bay Australia

ABSTRACT Quorum sensing is a potent system of genetic control allowing phenotypes to be coordinated across localized communities. In this study, quorum sensing systems in Shark Bay microbial mats were delineated using a targeted approach analyzing whole mat extractions as well as the creation of an isolate library. A library of 165 isolates from different mat types were screened using the AHL biosensor E. coli MT102. Based on sequence identity 30 unique isolates belonging to Proteobacteria, Actinobacteria and Firmicutes were found to activate the AHL biosensor, suggesting AHLs or analogous compounds were potentially present. Several of the isolates have not been shown previously to produce signal molecules, particularly the members of the Actinobacteria and Firmicutes phyla including Virgibacillus, Halobacillius, Microbacterium and Brevibacterium. These active isolates were further screened using thin-layer chromatography (TLC) providing putative identities of AHL molecules present within the mat communities. Nine isolates were capable of producing several spots of varying sizes after TLC separation, suggesting the presence of multiple signalling molecules. This study is the first to delineate AHL-based signalling in the microbial mats of Shark Bay, and suggests quorum sensing may play a role in the ecosphysiological coordination of complex phenotypes across microbial mat communities.

QUANTIFICATION OF EXEMESTANE ACCUMULATION DURING MICROBIAL BIOCONVERSION BY TLC IMAGE ANALYSIS

Objective: The present study was aimed at developing a rapid, cost effective and accurate method for quantification of exemestane using thin layer chromatography (TLC) separation followed by image analysis and to test it for monitoring the accumulation of exemestane during microbial bioconversion.Methods: After microbial bioconversion and TLC separation of products formed, exemestane was quantified using ImageQuant TL v2003 image analysis software and the results were compared with high performance liquid chromatography (HPLC) analysis.Results: The percentage error between TLC and HPLC analyses was ranged from (-) 5.18 to (+) 5.51. Bacterial strains Arthrobacter simplex IAM 1660, Nocardia sp. MTCC 1534, Pseudomonas putida MTCC 1194 and Rhodococcus rhodochorus MTCC 291 respectively yielded 79.7 (72 h), 63.9 (72 h), 69.8 (96 h) and 83.2 (96 h) mole percent bioconversion of 6-methylene androstenedione to exemestane. Conclusion: Rhodococcus rhodochorus MTCC 291 was found to be the most suitable organism for the bioconversion and may be used to develop an eco-friendly route to replace chemical synthesis that eliminates the use of toxic chemicals and side products.