Jpc-journal of Planar Chromatography-modern Tlc最新文献

Traditional formulations owing to their holistic approach to health and wellness with minimum side effects are widely used all over the globe. Itrifal Muqawwi Dimagh (IMD) is a traditional polyherbal formulation used in the Unani system of medicine conventionally claimed to support and enhance cognitive function. The formulation is widely used by a large population due to its alleged therapeutic efficacy. Despite being widely used, the formulation lacks scientific validation and justification in establishing its therapeutic potential. The present study was designed to detect and identify bioactives responsible for acetylcholinesterase inhibitory activity by thin-layer chromatography (TLC)‒bioautography‒mass spectroscopy (MS) and its validation using in silico molecular approach. Quality control was done using ultraperformance liquid chromatography (UPLC)‒MS fingerprint analysis. The authentication of individual constituents of the formulation was done by using powder microscopy. Ethyl acetate and hydrolyzed ethyl acetate fraction of hydroalcoholic extract of IMD were prepared, and in vitro acetylcholinesterase (AChE) inhibitory activity by Ellman’s method was performed. The TLC‒MS bioautography revealed the presence of scopoletin, tannic acid, ellagic acid, and catechin as potential bioactive anticholinesterase metabolites. In silico analysis of the identified metabolites from IMD showed AChE activity of ten identified metabolites, moreover, catechin and naringenin showed the best of potential activity. UPLC‒MS analysis demonstrated separation of 33 phytocompounds in the best active fraction of formulation. Hence, based on our findings, it can be concluded that IMD has a great potential to overcome cholinergic deficiency and can be used for neuroprotection and other neurological disorders after successful in vivo pharmacokinetic and toxicity validation.

A novel, efficient, and sensitive high-performance thin-layer chromatography (HPTLC) method has been developed and validated for the concurrent measurement of amlodipine besylate (AML besylate) and azilsartan (AZL) in human plasma spiked with the mixture. Reflectance/absorbance densitometry was conducted using toluene‒ethyl acetate‒methanol‒acetone‒acetic acid (6:1.5:1:0.5:1, V/V) as the mobile phase, and separation was achieved on a precoated silica gel HPTLC plate. This chromatographic system yielded compact bands with excellent resolution at a retardation factor (R_F) of 0.22 ± 0.002 for AML besylate and 0.73 ± 0.001 for AZL. Quantification of AML besylate and AZL was performed at 244 nm within the ranges of 60‒600 ng per band and 90‒900 ng per band, respectively. Calibration plots exhibited strong linearity, with correlation coefficients of 0.9976 for AML besylate and 0.9974 for AZL. Following the International Council for Harmonisation (ICH) guidelines, the developed method was validated. The lowest detectable values for AML besylate and AZL were 13.79 ng per band and 18.62 ng per band, respectively. The recommended HPTLC methodology for the simultaneous determination of AML besylate and AZL is demonstrated to be sensitive, selective, accurate, and precise. This technique can effectively be applied to the simultaneous detection and quantification of AML besylate and AZL in synthetic mixtures and human plasma samples. The enhancing effect of ammonia on the absorption intensity and the bathochromic effect on the wavelength of absorbtion were investigated by molecular modeling and it is suggested that ammonia causes acrylamide to change into acrylamic acid with more conjugated double bonds that rationale the increase in the absorption intensity and the bathochromic shift in the wavelength of the absorption.

Lipophilicity is a physicochemical parameter well known as a decisive factor for predicting the successful development of a drug. Thus, a balance between potency and physicochemical properties during medicinal chemistry optimization is needed. In this study, the lipophilicity of isoindole-1,3(2H)-dione derivatives designed as phosphodiesterase 10A (PDE10A) inhibitors was determined by chromatographic [reversed-phase thin-layer chromatography (RP-TLC) and ultra-performance liquid chromatography/mass spectrometry (UPLC/MS)] and in silico methods. To assess the correlation between the obtained lipophilicity parameters, principal component analysis (PCA) was performed. logP values obtained by chromatographic (logP_RP-TLC and logP_UPLC/MS) and in silico methods were compared using the PCA method. The results of PCA revealed that logP_UPLC/MS and in silico clogP provided by the ChemDraw program were highly correlated. Compounds’ drug likeness was screened, and the pharmacokinetic properties were predicted. All the investigated compounds displayed drug-likeness properties, and they met the criteria of Lipinski’s rule of five, which predicted the oral bioavailability of drug candidates. Analysis of the influence of physicochemical properties on the biological activity showed that the compounds with increased potency on PDE10A had significantly higher topological polar surface area (TPSA) values. The blood‒brain barrier permeability and the hemolytic activity of model compound 18 were examined. The model compound 18 displayed no toxicity effect on erythrocytes in the hemolytic assay and good parallel artificial membrane permeability. The results showed that phthalimide compounds with benzimidazole moiety are a source of compound-targeted inhibition of PDE10A with balanced physicochemical and drug-likeness properties.

Desmodium oojeinensis (Roxb.) H. Ohashi (DO) plant has been traditionally known to possess astringent, anti-inflammatory, hepatoprotective, wound healing, antimicrobial, rejuvenating, etc., activities. Since standardization of medicinal plants is essential to assess their quality by determining the active chemical constituents, the present work was aimed to estimate lupeol, stigmasterol, and betulin in DO bark and roots by a validated thin-layer chromatography (TLC) method. The proposed instrumental TLC method was validated in terms of linearity and range, limit of detection (LOD) and limit of quantification (LOQ), accuracy, precision, robustness and specificity. The mobile phase containing hexane‒ethyl acetate (8.5:1.5, V/V) and TLC aluminum plate, precoated with silica gel 60 F₂₅₄ as stationary phase were used. Anisaldehyde‒H₂SO₄ reagent was used for post-chromatographic derivatization. Data for the calibration plots of lupeol, stigmasterol and betulin showed linearity with y = 20.097x + 5124.9 (r² = 0.9906), y = 12.466x + 1518.5 (r² = 0.99472), and y = 10.902x + 545.3 (r² = 0.9944) regression equations, respectively, in 100–500 ng band⁻¹ concentrations. Lupeol, stigmasterol, and betulin were eluted at R_F [± standard deviation (SD)] 0.42 ± 0.01, 0.27 ± 0.01, and 0.19 ± 0.01, respectively. The LOD and LOQ for lupeol, stigmasterol, and betulin were found to be 12.75 and 38.64 ng band⁻¹, 18.02 and 54.59 ng band⁻¹, and 13.35 and 40.46 ng band⁻¹, respectively. Stability study revealed that lupeol, stigmasterol, and betulin were stable in chloroform at 25 ± 2 °C for 24 h with %RSD value of 0.69, 1.07, and 1.64, respectively. The proposed work showed good precision, robustness and specificity of the method. Average percentage recoveries of lupeol, stigmasterol, and betulin from the chloroform extracts of DO bark and roots were 97.43 ± 0.47, 97.69 ± 0.59; 97.02 ± 1.04, 97.89 ± 0.65, 98.18 ± 0.73, and 97.68 ± 1.09, respectively. The contents (% w/w; n = 3) of lupeol, stigmasterol, and betulin in the chloroform extracts of bark and roots of DO were found to be 2.15 ± 0.03 and 0.45 ± 0.06, 0.43 ± 0.03 and 0.40 ± 0.05, and 2.65 ± 0.04 and 2.13 ± 0.05, respectively, on dry weight basis of the extracts.

A wide range of analytical techniques has been reported for the determination of amlodipine besylate (AML) and indapamide (IND) in their commercialized formulations. Nevertheless, sustainable/green analytical techniques for estimating AML and IND in combination are scarce in literature. Accordingly, the present research was carried out to establish a rapid, highly sensitive, and sustainable high-performance thin-layer chromatography (HPTLC) technique for the simultaneous estimation of AML and IND using the National Environmental Method Index (NEMI), Green Analytical Procedure Index (GAPI), and analytical greenness (AGREE) tools. Metal plates coated with 250-µm silica gel 60 GF₂₅₄ were utilized for chromatographic separation. Densitometric detection was carried out at 288 nm using mobile phase ethanol‒ethyl acetate‒triethylamine (5:5:0.3, V/V). The HPTLC with ultraviolet (UV) detection method was linear in the 300‒1500 ng per band and 100‒500 ng per band ranges for AML and IND, respectively. The assay of AML and IND in the marketed formulation was found to be 101.40 ± 1.55% and 99.33 ± 1.39%, respectively, using the suggested HPTLC‒UV method. The AGREE index for the greener HPTLC‒UV technique was found to be 0.73, suggesting an excellent greenness profile for the proposed HPTLC‒UV technique. The method was validated as per the International Council for Harmonisation (ICH) Q2(R1) guidelines.

Medicinal plant extracts include a multitude of chemical compounds. Therefore, there is a need for suitable analytical methods for their identification and standardization. The aim of the present study was to extract, isolate, and identify rutin from Capparis zeylanica (CZ) leaves and determine its content in the stems, leaves, roots, and flowers of CZ plant by a simple, sensitive, precise, accurate, and specific high-performance thin-layer chromatography (HPTLC) method for the first time. The HPTLC method used silica gel 60 F₂₅₄ for the stationary phase, ethyl acetate‒glacial acetic acid‒formic acid–water (10:1.1:1.1:2.6, V/V) as the mobile phase, and scanning under ultraviolet (UV) 264 nm light, by using deuterium lamp, without chemical reagents. The calibration curves for rutin were found to be linear in the range between 400 and 1400 ng per spot with R² = 0.9953 ± 0.0004. This method showed good peak symmetry for rutin (R_F: 0.418 ± 0.004) with limit of detection and limit of quantification of 14.10 and 42.73 ng/spot, respectively. The precision, accuracy, robustness, and specificity studies for CZ extracts and rutin are within the limits as per the International Council for Harmonisation guidelines. This study confirms the presence of rutin with content (% w/w) of 3.16 ± 0.01 in leaves, 5.92 ± 0.04 in stems, and 2.23 ± 0.02 in flowers, on dry weight basis of CZ extracts. However, it is not found at detectable levels in the roots of CZ plant. In conclusion, the proposed method is simple, reliable, and specific for the quantification of rutin. The results also show that CZ could be an important nutritional source of rutin.

Bisoprolol fumarate (BSF) is a cardioselective β-blocker, whereas telmisartan (TLM) is an angiotensin II receptor antagonist. Both drugs in combination are available as a formulation product effective in the management of hypertension. As a rapid and precise analytical method, there is still no high-performance thin-layer chromatographic (HPTLC) method reported for the simultaneous estimation of both the drugs in combination. Therefore, the aim of the present study was to develop a precise, accurate, and sensitive HPTLC method for simultaneous estimation of BSF and TLM in bulk and in formulation product. The separation was achieved by using a mobile phase mixture consisting of methanol‒ethyl acetate‒glacial acetic acid (MeOH‒EA‒GAA; 6:4:0.1, V/V). The retention factor (R_F) for BSF and TLM was found to be 0.435 ± 0.04 and 0.699 ± 0.04, respectively, and the peak areas were determined at 231 nm. The method was subsequently validated as per the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) Q2 (R1) guidelines. A linear relationship was obtained between the concentration ranges and peak areas with correlation coefficient (R²) ≥ 0.999. The developed method was found to be accurate, precise, and robust. The method was then applied for analysis of three different batches of marketed product, and the percent content for BSF and TLM in the formulation was found to be > 98.0% w/w.

Abstract

Majun Nisyan (MJN) is a polyherbal formulation used in Unani medicine for its therapeutic efficacy against Nisyan (forgetfulness/dementia). In the current work, an effort was made to develop and validate a new high-performance thin-layer chromatography (HPTLC) method for the quantitative estimation of the marker compounds (α + β) boswellic acids, β-asarone, isoeugenol, 6-gingerol and piperine from the formulation of MJN. The method employed HPTLC plates precoated with silica gel 60 F₂₅₄ as the stationary phase. The solvent system comprised of toluene‒ethyl acetate‒chloroform‒acetic acid (8:2:5:0.1, V/V). This method yields compact spots for (α + β) boswellic acids (R_F 0.102 ± 0.01), β-asarone (R_F 0.982 ± 0.04), isoeugenol (R_F 0.850 ± 0.03), 6-gingerol (R_F 0.698 ± 0.03) and piperine (R_F 0.355 ± 0.01). A CAMAG TLC Scanner 4 was used to carry out the densitometric scanning at 254 nm. The International Council for Harmonisation’s Q2(R1) guideline was used to validate the HPTLC method. Each gram of the MJN formulation was found to contain 0.0248 ± 0.0053 mg (α + β) boswellic acids, 0.0520 ± 0.0069 mg β-asarone, 0.0487 ± 0.0041 mg isoeugenol, 0.0468 ± 0.0023 mg 6-gingerol and 0.0609 ± 0.0084 mg piperine. The developed and validated HPTLC method was found to be simple, inexpensive, sensitive, precise and accurate. This method can be easily applied to routine control analysis of all commercially available Ayurvedic and herbal formulations containing (α + β) boswellic acids, β-asarone, isoeugenol, 6-gingerol and piperine.

The present work encompassed the assay quantification of diltiazem hydrochloride and its degradation impurity F by high-performance thin-layer chromatography (HPTLC). HPTLC was performed with chloroform‒methanol‒formic acid (7.5:1.5:0.2, V/V) as the mobile phase and aluminum-backed thin layer chromatography (TLC) plates precoated with a 200 μm layer of silica gel 60 F₂₅₄ as the stationary phase. The densitometric chromatograms were performed at 254 nm, and the method was validated as recommended in the International Council for Harmonisation (ICH) guidelines. Statistical data from the analysis showed that the method is precise and highly sensitive. The correlation coefficient values of diltiazem hydrochloride and its impurity F were 0.9998 and 0.9975, respectively. The limit of detection was 76.81 ng/zone for diltiazem hydrochloride and 29.21 ng/zone for impurity F, while the limit of quantification was 232.77 ng/zone for diltiazem hydrochloride and 87.05 ng/zone for impurity F. The preciseness of the method was demonstrated and calculated as the % relative standard deviation for diltiazem hydrochloride and its impurity F. The mean recovery of the accuracy for diltiazem hydrochloride was between 98.23% and 100.29%, whereas for impurity F it was 100.02‒100.29%. The simplicity of the method proves its applicability for the quantification of impurity F during the synthesis of diltiazem hydrochloride.

Currently, the antimicrobial activity of essential oils (EOs) is an outstanding research field due to antibiotic resistance of microorganisms. Thin-layer chromatography‒direct bioautography (TLC‒DB) is an effective, fast method to find components with antimicrobial activity in a mixture of plant compounds, e.g., in EOs. The volatility and hydrophobic characters of EOs require special experimental conditions, and disc diffusion assay is not appropriate to explore the antimicrobial activity of them. The aim of this study was to use “R” mutants, which are more sensitive to synthetic antimicrobial drugs, in DB to increase the sensitivity of this method. Our hypothesis was that these mutants show sensitivity to some EOs (thyme, clove, and peppermint) as well. The chemical composition of our tested EOs was measured with gas chromatography‒mass spectrometry (GC‒MS). The main compounds (39.8% thymol, 78.8% eugenol, and 50.4% menthol) of EOs showed notable antibacterial activity in TLC‒DB. Based on our results, we suggest to use Salmonella minnesota Re595 rough strain as test bacterium in bioautography, because it showed the highest sensitivity to the tested antibiotics (gentamicin and cephalexin) and EOs. Furthermore, this rough mutant could make TLC‒DB more faster, because only 4 h incubation time was enough to detect the inhibition zones of the active compounds used in this study.