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Summary

Clerodendrum viscosum leaves are used in indigenous systems of medicines of mainland and maritime Southeast Asian countries for the treatment of fever, pain, dysentery, colic, and removal of Ascarids. The Clerodendrum species under study exhibit various phytochemical and morphological similarities. Therefore, it is very challenging to distinguish raw powdered materials used for therapeutic purposes. A validated high-performance thin-layer chromatography (HPTLC) method with 4 key markers, viz., 24ß-ethylcho-lesta-5,22E,25-triene-3ß-O-D-glucoside, clerodinin-A, 24ß-ethyl-cholesta-5,22E,25-triene-3ß-ol, and lupeol coupled with a chemometric analysis was used to distinguish 3 closely related Clerodendrum species, viz., C. inerme, C. multiforum, and C. viscosum. PRISMA approach was applied for effective HPTLC fingerprint development. The HPTLC-densitometry method was validated following the current International Conference on Harmonisation (ICH) guidelines. Taxonomic differentiation was established by fingerprint-based similarity analysis, a chemotaxonomic study using hierarchical clustering analysis (HCA), and principal component analysis (PCA) was done. HPTLC chromatogram similarity was calculated as correlation coefficient and congruence coefficient values, demonstrating poor similarities (0.26–0.86). However, PCA has resulted in 2 principal component (PC) loadings. PC1 separated C. multiforum, explaining 85.48% of variance mainly due to distribution of 2 triterpenoids. The present HPTLC method is coupled to marker-based quality determination of raw plants as well as discrimination of Clerodendrum species. Chemometric analysis based on 4 metabolites clearly establishes a practical identification of Clerodendrum species intended for therapeutic use.

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A two-dimensional thin-layer chromatographic fingerprint has been developed on a polyamide plate for the quality control of Helleborus thibetanus Franch. The investigated sample was separated by chloroform-ethyl acetate-methanol (3.0:8.0:4.4, v/v) in the first dimension and isooctane-n-propyl alcohol-water (10:2.5:1.0, v/v) adding 0.28 mol L−1 sodium dodecyl sulfate, a reversed micelle, in the second dimension. The plate was dried in the air at room temperature and examined in ultraviolet (UV) light at λ = 365 nm after development. The two developments were carried out over a distance of 70 mm. The two-dimensional thin-layer chromatographic method was validated in terms of repeatability, stability, and robustness. For fingerprint analysis, nine spots were identified as common spots. Statistical method (canonical correlation analysis) was first used to calculate the degree of similarity between the two-dimensional chromatograms. The proposed method was novel and accurate for the identification and quality evaluation of H. thibetanus Franch.

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Summary

A new objective chromatographic response function, R K, based on the kernel density estimate, is introduced for estimation of the fingerprinting performance of a particular TLC system (uniformity of retention) for which a large set of experimental R F values of possible components of the mixture is available. The R K criterion is insensitive to large numbers (hundreds or thousands) of R F values, when the previously proposed criteria cease. It can be applied to one and two-dimensional TLC and is easily computed. As an example of its application, the performance of twelve general screening systems was evaluated in the context of herbal extract fingerprinting (88 phytochemical standards) by both one and two-dimensional TLC.

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Shenqi Fuzheng Injection (SFI) is a traditional Chinese medicine injection, widely used to enhance immune function of clinical cancer patients undergoing chemotherapy. In this study, a high-performance liquid chromatography-diode array detection-evaporative light scattering detection (HPLC-DAD-ELSD) method was established for quality control of SFI, which could simultaneously semiquantitatively reflect the constituents displayed in the chromatographic profile of SFI. The relative retention time and relative peak areas of the 21 common peaks related to the reference peak were calculated. The validity and advantage of this method were validated by systematically comparing chromatograms of 10 batches of SFI samples with the analytical methods of principal component analysis and angle cosine method recommended by the State Food and Drug Administration of China. Moreover, a total of 21 constituents of SFI were identified or tentatively characterized in the fingerprint via ultrafast liquid chromatography-diode array detection-quadrupole time-of-flight (UFLC-DAD-Q-TOF) tandem mass spectrometry technique on the basis of the retention time, ultraviolet spectra, fragmentation patterns, and reported literatures. All the results proved that the technique was useful in comprehensive quality evaluation of SFI and further study.

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Summary

A high-performance liquid chromatographic method with diode-array detection (HPLC-DAD) has been developed and validated to establish the fingerprint of Turnera diffusa. Hydroalcoholic extracts were obtained from 19 raw herbal samples collected in different regions of México. Separation was performed on a 150 mm × 3.9 mm (4-μm particle) C18 column, using a gradient of methanol and 0.1% aqueous trifluoroacetic acid as the mobile phase. Chromatograms were recorded at 254 nm. To identify each peak, both retention time and peak spectrum were used. Intraday and interday relative standard deviations were <3% for retention time and <12% for relative areas. Extracts were stable in solution for up to 60 days. Results from a robustness study showed that the amount of ethanol in the mobile phase had a substantial effect on retention time. The relative areas of 12 peaks common to the chromatograms obtained from 19 authenticated T. diffusa samples were chosen to construct a principal-components analysis (PCA) model. The soft independent modeling of class analogy (SIMCA) method based on the PCA model was used to evaluate the quality of eleven commercial products.

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Three molecular markering techniques: inter-simple sequence repeat (ISSR); start codon targeted (SCoT), conserved DNA-derived polymorphism (CDDP) markers were compared for fingerprinting of 40 varieties of bread wheat. The number of scoreable and polymorphic bands produced using the ISSR, SCoT and CDDP primers for varieties was more than that of genotypes. Average polymorphism information content (PIC) for ISSR, SCoT and CDDP markers was 0.39, 0.41 and 0.34, respectively, and this revealed that three different marker types were equal for the assessment of diversity amongst genotypes. Cluster analysis for three different molecular types revealed that genotypes taken for the analysis can be divided in three and four distinct clusters. There were no significant differences among these markers in terms of the evaluation of genotypes. These results suggest that efficiency of SCoT, CDDP and ISSR markers was relatively the same in fingerprinting of genotypes but SCoT and CDDP analysis are more effective in fingerprinting of wheat genotypes. To our knowledge, this was the first detailed report of a comparison of performance among two targeted DNA region molecular markers (SCoT and CDDP) in comparision with ISSR technique on a set of samples of wheat cultivars. Overall, our results indicate that SCoT, ISSR and CDDP fingerprinting could be used to detect polymorphism for genotypes of wheat.

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Salvadora persica plant having a number of antimicrobial substances and the roots of the S. persica shrub have been demonstrated to possess antimicrobial activity. Sticks from the roots of S. persica have been used for centuries as a traditional method of cleaning teeth. Benzyl isothiocyanate is the main antimicrobial agent found in the root of the plant. Here, we developed high-performance thin-layer chromatography (HPTLC) and reversed-phase high-performance liquid chromatography (RP-HPLC) methods for the qualitative analysis of benzyl isothiocyanate in the S. persica root powder and the dental care herbal formulation labeled to contain Miswak extract. A HPTLC method was carried out using pre-coated silica gel aluminum TLC plates 60 F254, and the mobile phase consisting of n-hexane (10 mL) furnished compact spots at R F 0.39 ± 0.01. A HPLC method was carried out using a column Phenomenex C18(150 mm × 4.6 mm, 5 μm) and methanol‒10 mm ammonium acetate (70:30, v/v) as the mobile phase with a good retention time, 8.179 ± 0.05. The developed method was validated by HPTLC in accordance with the International Council of Harmonization (ICH) Q2 (R1) guidelines for precision, accuracy, and robustness. The developed plate was scanned and quantified densitometrically at 250 nm. Linear regression analysis revealed a good linear relationship between the peak area and the amount of benzyl isothiocyanate in the range of 3–15 μg per band (r 2 = 0.997). Fingerprinting was done by HPTLC and HPLC method by using a dental care herbal formulation labeled to contain Miswak extract. The proposed method will be useful to evaluate the therapeutic efficacy of Miswak extract and dental care herbal formulations containing Miswak extract.

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The evolution of individual anthocyanins during vinification of Merlot and Pinot Noir grapes was studied using two different winemaking procedures for each grape variety. Additionally, the effect of the applied vinification on the anthocyanin composition of the obtained wine at the end of maceration and wine aged 6 months was investigated and compared with the anthocyanin patterns of the original grape. The dynamics of the extraction process was monitored daily during maceration by analysing the anthocyanins in the must using HPLC. The results showed that the anthocyanin composition of young wines was different from that of the grapes. The proportions of malvidin-3- glucoside and malvidin-acetate were higher in wines than in the grape skins, but this was not the case for malvidin coumarate. Application of different vinification procedures to the same raw material resulted in wines with similar anthocyanin patterns. However, the anthocyanin profiles changed with the ageing of the wines.

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Detailed studies of the gas release profiles of minerals and rocks reveal differences in the degassing rates and degassing temperatures and also in the compositions of the fluids evolved from the samples.

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