Determination of the lipophilicity of some anti-hypoxia drugs: Comparison of TLC and HPLC methods

Summary The lipophilicity of 14 anti-hypoxia drugs has been determined by reversed phase thin-layer (RPTLC) and reversed phase high performance liquid chromatography (RPHPLC) in eluent systems containing different concentrations of acetonitrile and potassium dihydrogen phosphate. There was significant correlation between lipophilicity and the specific hydrophobic surface area of the drugs in RPTLC, indicating that the drugs behave as an homologous series of compounds. In RPTLC the concentration of buffer has a negligible effect on the retention of the drugs whereas in RPHPLC the buffer concentration influenced the retention. This discrepancy can be explained by the lower sensitivity of RPTLC. There was strong correlation between lipophilicity values determined by both methods, proving that both are suitable for the determination of molecular lipophilicity.     

Ultrasound-assisted extraction, HPLC analysis, and antioxidant activity of polyphenols from unripe apple

The polyphenols were extracted from the unripe apple assisted by a highly efficient and simple method of the ultrasound. Response surface methodology was used to investigate the effects of processing parameters, including ultrasound power, extraction time, temperature, and ethanol concentration on total polyphenols yield and polyphenols composition was analyzed by HPLC. Antioxidant activity of the polyphenols was evaluated as 2, 2-diphenyl-1-picrylhydracyl scavenging activity and inhibition activity of lipid peroxidation. The results showed that 10-100 times higher total polyphenols yield was obtained from the unripe apple than those from the reported apple pomace. The optimum extraction conditions were ultrasonic power of 519.39 W, extraction time of 30 min, extraction temperature 50°C, ethanol concentration of 50% gave the total polyphenols yield of 13.26 ± 0.56 mg GAE/g. HPLC analysis indicated that (-)-epicatechin, procyanidin B2, chlorogenic acid, and procyanidin B1 were the predominant polyphenols in unripe apple, which contributed to the higher antioxidant activity to 2, 2-diphenyl-1-picrylhydracyl of unripe apple polyphenols than other apple polyphenols. The extracted polyphenols had higher ability to inhibit lipid peroxidation than butylated hydroxy toluene, which demonstrated that the unripe apple polyphenols have the potential to be used as a substitute of some synthetic antioxidants.     

Effect of drying methods on the content of natural pigments and antioxidant capacity in extracts from medicinal plants: a spectroscopic study

Medicinal plants are a significant source of biological active and health protective compounds. Post-harvest treatments can affect, in different ways, the content and biological activity of such compounds. One of the most common post-harvesting methods is drying. In this study, we investigated the effect of drying method on the content of natural pigments (chlorophylls, carotenoids and anthocyanins) and on the antioxidant capacity of two traditionally used herbs, the Melissa officinalis (lemon balm) and the Urtica dioica (stinging nettle) both of them landraces collected from plants grown in Nitra region, West of Slovakia. The freeze-dried samples of both herbs exhibited the highest content of chlorophyll a (7.55 ± 0.13 mg/g dry mass for lemon balm and 9.41 ± 0.17 mg/g dry mass for stinging nettle), chlorophyll b (3.12 ± 0.28 mg/g dry mass for lemon balm and 3.34 ± 0.24 mg/g dry mass for stinging nettle) and carotenoids (2.11 ± 0.24 mg/g dry mass for lemon balm and 2.62 ± 0.06 mg/g dry mass for stinging nettle). The content of chlorophylls and carotenoids correlated with the DPPH antioxidant (radical scavenging) capacity. Higher antioxidant capacity of the lemon balm extracts compared to nettle samples was attributed to the higher content of polyphenol compounds anthocyanins. Despite the higher cost, the freeze drying (lyophilisation) was recommended as the most suitable drying method, mainly for reasons of preserving maximum pigment content and antioxidant capacity.     

甘松不同溶剂提取物的抗氧化活性研究

采用DPPH、ABTS、羟自由基、超氧阴离子和还原力五种体外抗氧化测定方法对甘松95%乙醇提取物以及石油醚萃取物,乙酸乙酯萃取物,正丁醇萃取物和水萃取物等4个不同极性部位的抗氧化活性进行评价,同时分析抗氧化活性与其总多酚和总黄酮含量的相关性。研究结果表明,除水和石油醚萃取物外,甘松其他3个萃取物均表现出一定的抗氧化活性,且与总多酚和总黄酮含量呈显著相关。其中,乙酸乙酯萃取物中总黄酮和总多酚含量最高,分别为(157.22±1.89)mg·g-1和(99.43±1.23)mg·g-1,其清除DPPH、ABTS、超氧阴离子和羟自由基的IC50分别为(0.20±0.02)mg·mL-1、(0.15±0.01)mg·mL-1、(0.29±0.02)mg·mL-1和(0.35±0.02)mg·mL-1。甘松的乙酸乙酯萃取物具有显著的抗氧化活性,可以成为天然抗氧化活性化合物的良好来源。     

Determination of free phenolic acids and antioxidant capacity of methanolic extracts obtained from leaves and flowers of camel thorn (Alhagi maurorum)

The present study comprises the determination of some phenolic acids from the leaves and flowers of Alhagi maurorum by HPLC-DAD, confirmed by LC-MS-APCI. The antioxidant properties and measurements of the total phenolic contents of the extracts were assessed by 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging and Folin-Ciocalteu methods, respectively. It was found that the leaf extract had higher antioxidant potential (83.5%) than the flower extract (72.3%). The antioxidant properties and total phenolic contents of the leaves were higher than those of the flowers.     

Phytochemical composition,antioxidant activity and HPLC profiles of Swertia species from Western Ghats

Swertia chirayita is one of the potential medicinal plants of the family Gentianaceae in traditional medicine. Due to its high demand and scarcity, trade of chirayita is affected by adulterants. Swertia species from Western Ghats were compared with S. chirayita for phytochemical characterisation and antioxidant activities by using different extracts. This study revealed that acetone is the best extraction solvent of phenolic and flavonoid compounds with antioxidant properties as compared with other extracts. S. chirayita showed better antioxidant activity than other species with highest content of phenolics and flavonoids. Among the species from Western Ghats, Swertia minor has better antioxidant properties with higher content of phenolics and flavonoids when compared with S. chirayita. Gallic acid was detected in all species under study by using HPLC analysis. The Swertia species under study showed similar phytochemical properties and antioxidant potential and hence their use as substitute to S. chirayita needs to be further investigated.     

Validated TLC and HPLC Stability-Indicating Methods for the Quantitative Determination of Dapsone

JPC – Journal of Planar Chromatography – Modern TLC - Two stability-indicating chromatographic methods were developed for the analysis of dapsone in presence of its oxidative...     

Determination of biologically active substances in roasted coffees using a diode-array HPLC system.

We studied the simultaneous quantitative analysis of biologically active substances, such as nicotinic acid, trigonelline, caffeine, qunolinic acid and tannic acid and pyrogallic acid, in several roasted coffees by an HPLC/diode-array system with a home-made sol-gel and ODS-2 columns. A simple method for simultaneous quantitative analysis of biologically active substances in the coffee brew became feasible by an HPLC/diode-array system with a sol-gel column at a single wavelength of 210 nm. The most efficient condition of the Rs value was above 1.05 when two sol-gel columns were connected. In addition, the elution behavior of nicotinic acid in brew extracted from commercially available coffee beans suggests the thermal decomposition process during roasting, and indicated the maximum value for full city roasted coffee.     

Phenolic profiles and antioxidant capacities of crude extracts and subsequent fractions from Potentilla fruticosa L. leaves

This work aimed to further investigate the phenolic profiles and antioxidant capacities of the crude extracts and the subsequent fractions of Potentilla fruticosa leaves. Result showed that P. fruticosa leaves contained high amounts for hyperoside, ellagic acid and (+)-catechin contents, and the highest amount being registered for hyperoside (17.67 mg g^?1). Nine sub-fractions were obtained after column chromatographic separation. EF-3, EF-4, EF-5 and BF-2 presented higher values for their total phenolic or flavonoid, (+)-catechin, ellagic acid and hyperoside content. Besides, EF-3, EF-4, BF-2 and BF-3 showed significant in vitro antioxidant capacities and protective effects on Escherichia coli under peroxide stress. The correlation between chromatograms and antioxidant activity showed that (+)-catechin, ellagic acid and hyperoside may play crucial roles in the antioxidant capacities of P. fruticosa and could be used as chemical markers for its quality assessment. Moreover, this is the first time P. fruticosa leaves have been systematically studied.     

Determination of the active metabolites of sibutramine in rat serum using column-switching HPLC

A simple and direct analysis using column-switching HPLC method was developed and validated for the quantification of active metabolites of sibutramine, N-mono-desmethyl metabolite (metabolite 1, M1) and N-di-desmethyl metabolite (metabolite 2, M2) in the serum of rats administered sibutramine HCl (5.0 mg/kg, p.o.). Rat serum was directly injected onto the precolumn without sample prepreparation step following dilution with mobile phase A, i. e., methanol-ACN-20 mM ammonium phosphate buffer (pH 6.0 with phosphoric acid) (8.3:4.5:87.2 by volume). After the endogenous serum components were eluted to waste, the system was switched and the analytes were eluted to the trap column. Active metabolites M1 and M2 were then back-flushed to the analytical column for separation with mobile phase B, i. e., methanol-ACN-20 mM ammonium phosphate buffer (pH 6.0 with phosphoric acid) (35.8:19.2:45 by volume) and detected at 223 nm. The calibration curves of active metabolites M1 and M2 were linear in the range of 0.1-1.0 microg/mL and 0.15-1.8 microg/mL. This method was fully validated and shown to be specific, accurate (10.4-10.7% error), and precise (1.97-8.79% CV). This simple and rapid analytical method using column-switching appears to be useful for the pharmacokinetic study of active metabolites (M1 and M2) of sibutramine.     

Anticholinesterase,antioxidant activity and phytochemical investigation into aqueous extracts from five species of Agrimonia genus

Aqueous extracts of aerial flowering parts of five Agrimonia species (Rosaceae): Agrimonia coreana Nakai, Agrimonia japonica (Miq.) Koidz, Agrimonia procera Wallr., Agrimonia eupatoria L. and Agrimonia leucantha Kunze were investigated on their antioxidant activity, measured using five different methods; the best was the extract from A. procera with IC₅₀ values from 6 to 29 μg/mL. All the extracts displayed inhibition of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) at the tested concentration of 100 μg/mL. We found the highest inhibition of cholinesterase in the extract of A. japonica with inhibition 70.4% for AChE and 79.8% for BuChE. These findings are statistically significant in comparison with those of other extracts (p < 0.001). The phytochemical analyses showed that the antioxidant activity of Agrimonia extracts can be affected especially by hexahydroxydiphenoyl (HHDP)-glucose and quercetin glycosides, and inhibition of cholinesterases by apigenin, luteolin and quercetin glycosides.     

Comparative study of chemical composition,antibacterial and antioxidant activity of essential oils isolated from the seeds of Amomum subulatum by using microwave extraction and hydro-distillation methods

Microwave assisted hydro-distillation (MAHD) and conventional hydro-distillation (HD) techniques were compared in the extraction of essential oils from Amomum subulatum seeds. The time required for MAHD method (70 ​min) is lesser than that for HD method (4 ​h). There is a slight increase in the yield of extracted oil in MAHD method (3.35%) compared to HD (3%). Gas chromatography–mass spectrometry GC-MS results show that MAHD extracted essential oil was wealthier in oxygenated compounds. 1, 8-Cineole was found to be a major compound in case of both the essential oil, followed by α-pinene. In MAHD the percentage of the major oxygenated monoterpene (1, 8- cineol) slightly increases from 88% to 89% as compared to hydrodistillation. Contrarily to this, the percentage of monoterpene hydrocarbon was decreased in MAHD than HD extracted oil. MAHD and HD extracted oils show good antibacterial activities against gram-negative and gram-positive bacteria. MAHD extracted oil shows better antibacterial activity than HD extracted against both gram positive and gram-negative bacteria. 2, 2-diphenyl-1-picrylhydrazyl (DPPH•) radical scavenging antioxidant activities show that MAHD extract has better inhibition percentage than HD extract, and the half-maximal inhibitory concentration IC50 value of MAHD was less than HD extracted oil.     

Determination of glimepiride in pharmaceutical formulations using HPLC and first-derivative spectrophotometric methods

Two validated analytical methods have been developed to determine glimepiride in pharmaceutical formulations using HPLC and 1st order derivative spectrophotometric techniques. Employing reverse phase HPLC method, the drug was analyzed by pumping a mixture of acetonitrile and 2% formic acid solution, pH 3.5 (80: 20 v/v) through a C₁₈ column (250 × 4.6 mm, 5 μm) and detecting the eluents at 228 nm. The linearity range was found to be 20–140 μg/mL with mean recovery of 100.52 ± 0.33%. The second method was based on the formation of a complex of the drug with 2,3,5-triphenyl-2H-tetrazolium chloride in basic media. 1st order derivative spectrum made it possible to detect the complex at 413.5 nm. The linearity range was found to be 40–160 μg/mL, with mean recovery of 100.33 ± 0.47%. Both the proposed methods can reliably be used for routine analysis of glimepiride in raw material as well as in pharmaceutical formulations. The article is published in the original.     

Profiling of phenolic compounds and antioxidant activity of dry extracts from the selected Sorbus species

The antioxidant efficiency of dry extracts from inflorescences and/or leaves of seven Sorbus species was studied using four in vitro tests of SET (single electron transfer) and HAT-type (hydrogen atom transfer) mechanisms. The 70% methanol extracts and its diethyl ether, ethyl acetate, n-butanol and water fractions were tested in parallel with the phenolic standards, e.g., caffeic acid, quercetin, BHA, BHT, and Trolox. The SET-type activity of the extracts depended primarily on the extraction solvent. The most valuable extracts were n-butanol and ethyl acetate ones, which activity was high in the DPPH (EC(50) = 3.2-5.2 μg/mL), TEAC (2.8-4.0 mmol Trolox/g), and FRAP (9.8-13.7 mmol Fe2+/g) tests, and strongly correlated with the total phenolic levels (39.6-58.2% of gallic acid equivalents). The HPLC-PDA analysis of the extracts led to the identification of chlorogenic acid, isoquercitrin, hyperoside, rutin, quercetin 3-O-sophoroside, and sexangularetin 3-O-β-D-glucopyranoside as the main components. Apart from flavonoids and hydroxycinnamic acids, proanthocyanidins have also a significant impact on the SET-type activity. The HAT-reactivity of the extracts in the linoleic acid peroxidation test (IC(50) = 36.9-228.3 μg/mL) depended more strongly on the plant tissue than on the extraction solvent, and its correlation with the phenolic content was weak. Both SET and HAT-type activity of the most potent Sorbus extracts was comparable with the activity of the standards, indicating their great potential as effective sources for health products.     

Antioxidant potential and phytochemical composition of extracts obtained from Phyllanthus phillyreifolius by different extraction methods

Abstract

Phyllanthus phillyreifolius (Euphorbiaceae), poorly studied plant species, was fractionated using conventional and high pressure extraction techniques such as supercritical fluid and pressurized liquid extractions. Lipophilic substances were extracted with n-hexane and supercritical CO₂ with or without co-solvent ethanol, meanwhile higher polarity fractions were recovered with acetone and 70% ethanol. Antioxidant potential was assessed by various chemical assays, which revealed that 70% ethanol was the most effective solvent for recovery of antioxidants. UPLC-MS phytochemical analysis of hydrophilic extracts confirmed geraniin as the main constituent of P. phillyreifolius. Other quantitatively important compounds were phyllanthusiin D and elaeocarpusin. Three isomers of tocopherol (α, β and γ) were quantified by HPLC in lipofhilic extracts. Generally, the results from this study revealed high antioxidant potential of P. phillyreifolius; consequently the plant may be considered as a promising source of antioxidants for functional foods, nutraceuticals and pharmaceutical formulations.     

A comparative analysis of raw and processed Radix Paeoniae Alba samples using HPLC,chemometrics, and antioxidant activity

Radix Paeoniae Alba (RPA) and its processed products are commonly used in traditional Chinese medicine, but the chaos phenomenon among processed products often occurs. In this study, we used multiple chemometric methods to analyze raw and six different processed products of RPA based on HPLC fingerprinting. Heat map analysis was used to assess the changes in chemical composition. Principal component analysis was used for classification, and the samples were divided into four classes: class 1 (raw, wine-processed, and vinegar-processed products), class 2 (bran-processed and soil-processed products), class 3 (stir-fried products), and class 4 (coke products). Further, the orthogonal partial least squares discriminant analysis model was used to obtain chemical markers among different classes. The antioxidant property of RPA is an important factor responsible for its pharmacological effects, and so the antioxidant activity of RPA was also investigated. We measured 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) and hydroxyl radical scavenging ability, and ferric reducing antioxidant power. The total antioxidant activity follows the order: coke > stir-fried > soil-processed > bran-processed > wine-processed > raw > vinegar-processed products. These results suggest that different processing methods affect the chemical composition and antioxidant power of RPA, and thus, different products of RPA should not be mixed.     

Pressurized liquid extracts from Spirulina platensis microalga. Determination of their antioxidant activity and preliminary analysis by micellar electrokinetic chromatography

In this work, different extracts from the microalga Spirulina platensis are obtained using pressurized liquid extraction (PLE) and four different solvents (hexane, light petroleum, ethanol and water). Different extraction temperatures (115 and 170 degrees C) were tested using extraction times ranging from 9 to 15 min. The antioxidant activity of the different extracts is determined by means of an in vitro assay using a free radical method. Moreover, a new and fast method is developed using micellar electrokinetic chromatography with diode array detection (MEKC-DAD) to provide a preliminary analysis on the composition of the extracts. This combined application (i.e., in vitro assays plus MEKC-DAD) allowed the fast characterization of the extracts based on their antioxidant activity and the UV-vis spectra of the different compounds found in the extracts. To our knowledge, this work shows for the first time the great possibilities of the combined use of PLE-in vitro assay-MEKC-DAD to investigate natural sources of antioxidants.     

Effect of electron-beam irradiation on the antioxidant activity of extracts from Citrus unshiu pomaces

After electron-beam irradiation of citrus pomaces (CP), the total phenolic content (TPC), radical scavenging activity (RSA), and reducing power (RP) were evaluated. When CP were irradiated at 37.9 kGy; the TPC, RSA and RP of water extract of CP increased from 6543.2 to 7405.4 μM, 37.6% to 52.9%, and 0.64 to 0.90, respectively, compared with the non-irradiated control. The results indicate that the electron-beam irradiation can be an efficient process for increasing the antioxidant activity of CP.