Authors:Omer A. Basudan, Perwez Alam, Nasir A. Siddiqui, Mohamed F. Alajmi, Adnan J. Alrehaily, Saleh I. Alqasoumi, Maged S. Abdel-Kader, Prawez Alam, and Abd El Raheim M. Donia
A simple and sensitive high-performance thin-layer chromatographic (HPTLC) method was developed for the evaluation of biomarker β-amyrin in the leaves of fve different species of genus Ficus (Ficus carica, Ficus nitida, Ficus ingens, Ficus palmata, and Ficus vasta) grown in the Kingdom of Saudi Arabia. Chromatography was performed on glass-backed silica gel 60 F254 HPTLC plates with solvents toluene–methanol (9:1, v/v) as the mobile phase. After development, the HPTLC plate was derivatized with p-anisalde-hydereagent to give well-resolved and compact spot of β-amyrin. Scanning and quantifcation were done at 550 nm. The system was found to give compact spot for β-amyrin at RF = 0.58. The linear regression analysis data for the calibration plots showed good linear relationship with r2 = 0.998 with respect to area in the concentration range of 100–900 ng. The regression equation for β-amyrin standard was found to be Y = 5.835X + 87. The precisions (n = 6) for β-amyrin were found to be 1.64–1.77% and 1.68–1.84%, respectively, for intra-day and inter-day batches, and the recovery values were found to be 97.6–98.3%. β-Amyrin was found to be present in three species, i.e., F. carica (0.29%, w/w), F. nitida (0. 5 4% w/w), and F. p almata (0.31%, w/w), while it was absent in F. vasta and F. ingens. The statistical analysis proves that the developed method for the quantifcation of β-amyrin is reproducible; hence, it can beemployed for the determination of β-amyrin in plasma and other biological fuids as well as in fnished products avai lable in the market.
Authors:Mohamed AlAjmi, Perwez Alam, and Faiyaz Shakeel
A simple and sensitive high-performance thin-layer chromatographic (HPTLC)-densitometric method was developed and validated for quantification of β-amyrin in the crude extracts of two species of Maytenus (Maytenus obscura and Maytenus parviflora) grown in Saudi Arabia. HPTLC-densitometry was performed on glass-backed silica gel 60 F254 TLC plates with the binary mobile phase hexane-ethyl acetate (3:1, v/v). The developed plate was derivatized with p-anisaldehyde, then scanned and quantified densitometrically at 550 nm. The system was found to give a compact spot for β-amyrin at RF value of 0.38 ± 0.01. The method was found to be satisfactory in terms of sensitivity, accuracy, precision, and recovery. The content of β-amyrin was estimated as 0.42% ± 0.01% and 0.88% ± 0.01% w/w in M. obscura and M. parviflora, respectively. The developed HPTLC technique can be very useful for the quantification of β-amyrin present in various medicinal plants.
Authors:Mohammed F. Alajmi, Afzal Hussain, and Perwez Alam
In this study, we have developed a validated high-performance thin-layer chromatography (HPTLC) method for the concurrent estimation of the biomarkers β-amyrin and β-sitosterol in dichloromethane and ethanol extracts of the aerial parts of Tinospora cordifolia (TCDC and TCET) and Calotropis gigantia (CGDC and CGET). Chromatographic estimations were carried out on HPTLC (glass-backed silica gel 60 F254) plates with solvents hexane and ethyl acetate in the ratio of 7.5:2.5, v/v (as the mobile phase). Post development, the plate was derivatized with p-anisaldehyde reagent to furnish compact spots of β-amyrin and β-sitosterol and scanned at λmax = 530 nm. Well-resolved, compact as well as intense peaks of β-sitosterol (RF = 0.26 ± 0.001) and β-amyrin (RF = 0.39 ± 0.001) were found. The linear regression equation and the correlation coefficient square (r2) for β-amyrin (Y = 6.118x + 460.76 and 0.9959) and β-sitosterol (Y = 7.109x + 1069.1 and 0.9967) in the concentration range of 100–1400 ng spot−1 indicated good linear relationship. The low values of the percent relative standard deviation (% RSD) for intra-day and inter-day precisions for β-amyrin (1.003–1.148 and 0.993–1.105) and β-sitosterol (0.578– 0.969 and 0.513–0.813) suggested that the method is precise. The % recovery and % RSD values were found to be 98.42–99.29% and 1.103–2.103, respectively, for β-amyrin and 98.33–99.39% and 1.375–2.346, respectively, for β-sitosterol, which confirms the good accuracy of the proposed method. The quantity of β-amyrin in TCDC, CGDC, TCET, and CGET was found to be 70.14, 10.76, 4.85, and 0.87 μg mg−1, respectively, of the dried weight of the extracts, while the β-sitosterol content was found to be 19.4, 18.5, 1.18, and 0.27 μg mg−1, respectively. The highest quantity of β-amyrin and β-sitosterol in the dichloromethane fractions of T. cordifolia and C. gigantia validated the wide range of their proved activities including antioxidant, anticancer, and hepatoprotective features. The above developed method can be further employed in the analysis of these markers in marketed preparations.
Authors:A. Ferchichi, S. Harrabi, M. Feki, and H. Fellah
been previously reported. The GC-MS analysis of terpene alcohol fraction of the studied oil samples revealed the presence of two pentacyclictriterpene alcohols, β-amyrin and lupeol, and one non-cyclic diterpene alcohol, phytol. For both cultivars, the
Authors:Katarzyna Szewczyk, Łukasz Komsta, and Agnieszka Skalska-Kamińska
High performance thin-layer chromatography (HPTLC) combined with densitometry has been used for analysis of the triterpenoid content of acetone and ethyl acetate extracts of the leaves of
(Sims.) Opiz. After optimization of the extraction process, HPTLC separation was successfully standardized, using silica gel plates washed with methanol and dichloromethane, dichloromethane-ethyl acetate 18.5:1.5 as mobile phase, and 8% H
in ethanol (
) as spray reagent. Triterpenoids in the leaves of
were quantified by measurement of absorbance. Three spots were observed on chromatograms — the sum of α and β-amyrin, oleanolic acid, and an unidentified compound. All of the triterpenoid components were analyzed for the first time in this plant. The triterpenoid content was calculated statistically.
Authors:S. Trdan, N. Valič, L. Andjus, I. Vovk, M. Martelanc, B. Simonovska, J. Jerman, R. Vidrih, M. Vidrih, and D. Žnidarčič
In a field experiment on the natural resistance of cabbage (
) against onion thrips
, 20 cabbage genotypes were included: 9 early, 5 mid-early, 6 mid-late (regarding the longevity of the growing period), 3 red, 17 white (regarding colour), 14 hybrids and 6 varieties (regarding genetic origin). For comparisons between genotypes, we determined significant differences in the mean index of damage on the exterior leaves of cabbage heads (1.12–2.83), the net weight of heads (281.40,151169.6 g), and yield loss (5.8–47.4%). The concentration of several compounds from cabbage leaves (epicuticular wax, α-amyrin, β-amyrin, lupeol, sucrose, glucose, fructose, vitamin C, palmitic acid, stearic acid, and arachidic acid) are measured. For all groups of cabbage genotypes the only confirmed negative correlation was between the extent of damage caused by the sucking of onion thrips and epicuticular wax content on the cabbage leaves (in groups where we artificially placed genotypes with similar characteristics). After this preliminary experiment, the research must be extended in order to determine the effect of the remaining analysed compounds in cabbage leaves for resistance (sensitivity) of cabbage to onion thrips attack.
Authors:Katerina Naumoska, Breda Simonovska, Alen Albreht, and Irena Vovk
Separation of three triterpenic acids (ursolic, oleanolic and betulinic acid) was achieved on different thin-layer chromatography (TLC) (silica gel 60) and high-performance thin-layer chromatography (HPTLC) sorbents (silica gel 60, C2 RP and C18 RP) using several developing solvents, based on the non-polar diluent n-hexane, and ester (methyl acetate, ethyl acetate, ethyl propionate) as selector. Anisaldehyde and molybdophosphoric acid detection reagents were used. Finally, a simple method on a C18 RP HPTLC plate was developed using n-hexane-ethyl acetate (5:1 v/v) as a developing solvent in a horizontal developing chamber. The method was used for the screening of ursolic, oleanolic and betulinic acids in different vegetable extracts. Other plant triterpenoids (lupeol, α-amyrin, β-amyrin, cycloartenol, lupenone, friedelin, lupeol acetate, cycloartenol acetate) and phytosterols (β-sitosterol, stigmasterol) did not interfere. TLC-MS was used as a tool for the additional confirmation of the presence of ursolic, oleanolic, and betulinic acids in some of the studied vegetable extracts. Ursolic and oleanolic acids were found in radicchio Leonardo and white-colored radicchio di Castelfranco extracts for the first time, while betulinic acid was not detected in the eggplant extract by MS, although it was suggested at first by TLC analysis. Pre-chromatographic bromination on the HPTLC silica gel 60 plates and subsequent development in toluene-chloroform-diethyl ether-formic acid (20:16:4:0.1, v/v) provided a superior resolution of these compounds.
Authors:A. Böszörményi, Sz. Szarka, É. Héthelyi, I. Gyurján, M. László, B. Simándi, É. Szőke, and É. Lemberkovics
Our objectives were to establish a GC method capable of quantitative analysis of terpenoids without derivatisation and to examine the amount of β-sitosterol extracted from Morus alba L. leaf and stem bark by use of traditional organic solvent extraction and supercritical-fluid extraction (SFE). To measure β-sitosterol content without derivatization, GC-FID was used with 5-α-cholestan-3-one as internal standard. To identify terpenoid constituents, GC-MS was used; β-sitosterol, phytol, lanost-7-en-3-on, α-amyrin, β-amyrin, and lupeol were identified. We established that for Morus leaf the best SFE method for β-sitosterol was pilot scale SFE; the β-sitosterol content of this extract was higher than that of the hexane solvent extract. Among analytical SFE conditions, 200 bar for 90 min and 300 bar for 60 min resulted in extraction of the most β-sitosterol. For mulberry stem bark, solvent extraction with hexane and SFE at 400 bar and 40°C for 60 min proved the best methods.