Method for the analysis of intracellular adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) in Mycobacterium smegmatis that involves rapid extraction procedure based on sonication of cells in perchloric acid, as well as separation of nucleotides by ion-pair reversed-phase high-performance liquid chromatography and ultraviolet (UV) detection at 254 nm, is developed. The analytes were separated with mobile phase consisted of acetonitrile and 50 mM monobasic potassium phosphate (pH 4.6) with 25 mM tetrabutylammonium hydrogensulfate in a ratio of 0.5:99.5% within 30 min. The calibration curves were linear in the range of 20–1000 pmol of ATP and 10–1000 pmol of ADP and AMP with correlation coefficient (r
) of ≥0.9998. The proposed method is applicable for mycobacterium cultures taken over a wide range of optical density and physiological states. Concentrations of ATP, ADP, and AMP in mycobacterial extracts varied from 2.61 ± 0.27 to 9.60 ± 0.19 nmol/mg dry weight, from 1.75 ± 0.12 to 5.86 ± 0.09 nmol/mg dry weight, and from 0.55 ± 0.08 to 4.40 ± 0.07 nmol/mg dry weight, respectively, depending on the physiological state.
Authors:Na Zhou, Tao Li, Lianfeng Ai, Chunhai Guo, Juan Zhang, Shan Fu and Qiao Wang
Lacosamide, a new type of antiepileptic drug, was subjected to forced degradation under the conditions of hydrolysis (acidic and alkaline), oxidation, dry heat, and photolysis to characterize its possible degradation products. The drug showed significant degradation under acidic, alkaline and oxidative conditions. The degradation products were separated on an Agilent Zorbax SB-C18 column with gradient elution using a mobile phase consisting of acetonitrile and ammonium acetate (0.002 mol/L) with formic acid as additive. A combination of liquid chromatography hybrid triple quadrupole-linear ion trap mass spectrometry (LC–QqLIT-MS) and liquid chromatography hybrid ion trap/time-of-flight mass spectrometry (LC-IT/TOF-MS) was used to identify degradation products. A total of 7 products including 4 novel degradation products were characterized. The mechanisms of degradation products of lacosamide were discussed. Application of the method to study degradation products of lacosamide provided fragment information, allowing further investigation of the degradation pathways and intrinsic stability of the drug.
A new reliable simple solvent extraction method for the endocrine disruptor bisphenol A (BPA) in canned food was developed employing an aqueous basic extraction solution of 0.25 M K2CO3/0.10 M NaOH after spiking with BPA-d16 as internal standard. The BPA was next extracted into diethyl ether after solution acidification to pH = 4 and filtration. Homogenous acetylation at dry basic conditions (acetic anhydride as derivatization agent and solvent with sodium acetate as catalyst) after diethyl ether evaporation was carried out for 30 min at 110 °C. Detection of the acetylated BPA was carried out by gas chromatography–electrospray ionization/mass spectrometry (GC–EI/MS) in the selected ion monitoring (SIM) mode with pulsed split-less mode. The method was applicable in terms of eliminating the use of solvents like acetonitrile for the extraction step, where relatively long evaporation times may have been needed to evaporate acetonitrile. Also, removing lipids and precipitating most of the proteins at acidic conditions (pH = 4) prior to diethyl ether extraction can replace the often used heptane or hexane or solid sorbents. The method was tested linear with limit of linearity (LOL = 750 μg/L) and with coefficient of determination (R
= 0.998), repeatable with relative standard deviation (RSDr < 7%) with instrument detection limit (IDL) of 0.01 μg/L and limit of quantitation (LOQ) of 0.034 μg/L. The method detection limit (MDL) ranged from 0.3 μg/kg to 3.2 μg/kg based on 1 g sample (wet weight). Recovery ranged from 85% to 94% with the relative standard deviations of 2%–13%. BPA concentrations in tested canned foods from outlet stores ranged from <MDL to 57.4 ± (2.6) μg/kg which were below the specific limit for BPA migration in food proposed by the European Union (EU) and within the food safety and quality criteria. The extraction and derivatization steps for BPA were unique and have not been reported in literature.
Authors:H. W. Zhao, Y. L. Geng, H. Zhu, P. Yang and J. Q. Yu
Seven compounds, including two flavanones, dihydrokaempferol (1) and naringenin (2), and five terpenoids, boscartol A (3), 3,7-dioxo-tirucalla-8,24-dien-21-oic acid (4), 3α-acetoxyl-7-oxo-tirucalla-8,24-dien-21-oic acid (5), 11-keto-β-boswellic acid (6), and acetyl-11-keto-boswellic acid (7), have been purified by high-speed counter-current chromatography (HSCCC) from olibanum. For the separation, from 250 mg of the crude extract, 3.1 mg of 1 (95.2% purity), 2.7 mg of 2 (96.1% purity), 9.1 mg of 3 (96.7% purity), 4.5 mg of 4 (95.3% purity), 5.4 mg of 5 (96.3% purity), 48.1 mg of 6 (96.8% purity), and 45.5 mg of 7 (98.1% purity) were obtained by HSCCC with petroleum ether–ethyl acetate–methanol–water (1:0.8:1.1:0.6, v/v). The structures of these seven compounds were elucidated by a combination of electrospray ionization mass spectrometry (ESI–MS) and extensive nuclear magnetic resonance (NMR) spectroscopic.
Authors:Shiming Song, Huili Huang, Zhaojie Chen, Jie Wei, Cheng Deng, Huihua Tan and Xuesheng Li
A residue analytical method was developed for the determination of trichlorfon, chlorpyrifos, dimethoate, β cypermethrin, deltamethrin, and chlorothalonilin in six leafy vegetables by gas chromatography–electron capture detector (GC–ECD) and gas chromatography–flame photometric detector (GC–FPD). The method had a good linearity (R2 ≥ 0.9924) and precision (RSD ≤ 14.0%). The limits of quantification (LOQ) of six pesticides were all 0.01 mg/kg. Average recoveries of six pesticides ranged from 81% to 119%. The developed method was successfully applied to study the initial deposits, degrade characteristics, and terminal residues for six pesticides applied to six leafy vegetables under the same dose of formulation. The half-life of six pesticides was in the range of 0.8–8.8 days. The highest initial deposits, maximal residues, and terminal residues were found on leaf mustard and sweet potato leaves as the same pesticides were applied in different crops. Therefore, leaf mustard can be selected as representative commodity in the same subgroup to realize the residual extrapolation. This conclusion should be considered as a complement on crop classification of China.
Authors:Lianguo Chen, Haiya Wu, Xiaoting Tu, Yi Zhao, Yanyan Jiang, Congcong Wen and Yue Luo
Atractylodis exerted a variety of pharmacological effects such as anti-tumor, anti-inflammatory, anti-bacterial, and anti-aging effects etc. The major ingredients of Atractylodis are atractylenolide I and II that exhibited activities in anti-inflammatory and anticancer. In this work, a sensitive and selective ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method for determination of atractylenolide I and II in rat plasma was developed. The UPLC–MS/MS method was validated for selectivity, linearity, accuracy, precision, recovery, and stability with a total run time of 4.0 min. After addition of atractylenolide III as an internal standard (IS), protein precipitation by acetonitrile was used to prepare samples. Chromatographic separation was achieved on a UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm) with 0.1% formic acid and acetonitrile as the mobile phase with gradient elution. An electrospray ionization source was applied and operated in positive ion mode; multiple reactions monitoring (MRM) mode was used for quantification using target fragment ions m/z 231.1 → 185.1 for atractylenolide I, m/z 233.1 → 91.0 for II, and m/z 249.0 → 231.1 for IS. Calibration plots were linear throughout the range 1–1000 ng/mL for atractylenolide I and II in rat plasma. Mean recoveries of atractylenolide I and II in rat plasma ranged from 86.2% to 96.3%. Relative standard deviation (RSD) of intra-day and inter-day precision was both less than 12%. The accuracy of the method was between 91.0% and 109.0%. The method was successfully applied to pharmacokinetic study of atractylenolide I and II after intravenous administration in rats.
Authors:JianHua Zhang, Ling Wang, Xi Zhu, Xue Bai and Hua Yin
You-Gui-Yin (YGY), a famous traditional Chinese medicine, has been widely used in clinics for the treatment of kidney-yang deficiency, yang deficiency caused by excessive yin, and osteoporosis. A rapid and sensitive ultraperformance liquid chromatography–electrospray ionization–mass spectrometry (UPLC–ESI–MS) method for simultaneous determination of six Aconitum alkaloids including aconitine (AC), hypaconitine (HA), mesaconitine (MA), benzoylaconine (BAC), benzoylhypaconine (BHA), and benzoylmesaconine (BMA) in rat plasma after oral administration of YGY was developed in this study. Chromatographic separation was performed on an ACQUITY UPLC™ BEH C18 column (2.1 × 100 mm, 1.7 μm) using gradient elution with the mobile phase consisting of 2 mmol/L ammonium formate in 0.05% formic acid aqueous solution and 0.05% formic acid methanol solution, at a flow rate of 0.20 mL/min. MS detection was performed in the positive ion mode. The calibration curves were linear in the concentration range of 0.04160–41.60 ng/mL, 0.1070–107.0 ng/mL, 0.07358–73.50 ng/mL, 0.03228–32.28 ng/mL, 0.01809–18.09 ng/mL, and 0.1320–132.0 ng/mL for AC, HA, MA, BAC, BHA, and BMA, respectively. The intra- and inter-day precisions (relative standard deviation [RSD]) were less than 11.6% and 12.6%, respectively. The accuracies Relative Error (RE) ranged from −10.2% to 5.6%, while the recoveries ranged from 70.4% to 99.3%. The method for simultaneous quantitation of Aconitum alkaloids of You-Gui-Yin in rat plasma is accurate and repeatable, and this method was successfully applied to investigate the pharmacokinetics of the six Aconitum alkaloids in rat plasma after oral administration of YGY. For the pharmacokinetic study, the pharmacokinetics of the six Aconitum alkaloids were best described by a two-compartment open model.
Authors:Libin Wang, Xi Li, Le Mi, Xin Shen, Tian Feng, Xueying Liu and Qingwei Wang
2′,4′,6′,4-Tetra-O-acetylphloretin (TAPHL) is a prodrug of phloretin (PHL) in which the OH groups are protected by acetylation. A validated liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for the determination of PHL in rat biological matrices was developed and applied to investigate and compare the pharmacokinetics, tissue distribution, and excretion of PHL and TAPHL in rats following a single oral administration. The method was validated for accuracy, precision, linearity, range, selectivity, lower limit of quantification (LLOQ), recovery, and matrix effect. All validation parameters met the acceptance criteria according to regulatory guidelines. The mean pharmacokinetic parameters of tmax, Cmax, AUC(0 − t), CL/F, and t1/2 were observed after oral administration in rats. The data showed that PHL was absorbed and eliminated rapidly from plasma after oral administration. The pharmacokinetic properties are improved, such as the tmax has been prolonged and the area under the curve (AUC) has been enhanced after oral administration of TAPHL to rats. Tissue distribution results indicated that PHL could be rapidly and widely distributed into tissues but could not effectively cross the blood–brain barrier in rats. After oral administration of TAPHL to rats, its tissue distribution to rats was similar as that after oral administration of equimolar PHL. In addition, higher recoveries of PHL following administration of TAPHL indicated that TAPHL might reduce the excretion of PHL from the body by reducing the first pass effect.
Authors:U. Rakibe, R. Tiwari, V. Rane and P. Wakte
The aim of this work was to simultaneously separate, identify, and characterize all the degradation products (DPs) of atorvastatin (AT) and olmesartan (OM) formed under different stress conditions as per International Conference on Harmonization (ICH) Q1A(R2) guideline. AT showed labile behavior in acidic, basic, neutral, and oxidative stress and led to the formation of two DPs, while OM degraded under acidic, basic, and neutral and resulted in the formation of four DPs. All the stressed samples of AT and OM were resolved on a C-18 column in single run on a gradient liquid chromatographic (LC) mode. A complete mass fragmentation pathway of both the drugs was established with the help of tandem mass spectrometry (MS/MS) studies. The fragmentation was further supported by MSn studies, and for AT, it was carried out up to MS6, while for OM, it was up to MS5. Then, the stressed samples were analyzed by LC–MS/MS to get the fragmentation patterns of DPs. LC–MS/MS data helped to propose chemical structure of all the DPs. Based on this entire information, degradation pathway of both the drugs was established. The developed method has shown excellent linearity over the range of 10 to 150 μg/mL of OM and AT. The correlation coefficient (r2) for OM and AT is 0.999 and 0.998, respectively. The main recovery value of OM and AT ranged from 99.97% to 100.54%, while the limit of detection (LOD) for OM and AT was 0.018 and 0.021 μg/mL, and limit of quantitation (LOQ) was found to be 0.051 and 0.063 μg/mL. Finally, the in-silico carcinogenicity, mutagenicity, and hepatotoxicity predictions of AT, OM, and all the DPs were performed by using toxicity prediction softwares, viz., TOPKAT, LAZAR, and Discovery Studio ADMET, respectively.
Scentless chamomile (Tripleurospermum inodorum = M. inodora) is a plant belonging to Anthemideae tribe of Asteraceae family, with phenotype similar to the common chamomile, a plant used in human consumption in the form of herbal tea infusion. In order to be able to understand possible health-promoting properties and adverse effects of the scentless chamomile's consumption, it is of essence to examine its chemical composition. The aim of the study was to perform phenolic profiling using high-performance liquid chromatography–tandem mass spectroscopy (HPLC–MS/MS), in comparison to the common chamomile. In the investigated extracts, qualitative and quantitative analyses enabled the identification of 66 compounds based on their retention times, mass (MS/MS) spectra, and analysis of their characteristic fragmentation patterns in MS/MS Product Ion Scan experiments. A new HPLC–MS/MS method for quantitation of common plant metabolites was hereby developed, enabling quantitation of 47 compounds. All examined M. inodora samples have relatively high combined phenolic and flavonoid contents (25.2–51.9 mg/g). Apigenin, apigenin-7-O-glucoside, luteolin, luteolin-7-O-glucoside, quinic acid, and 5-O-caffeoyl quinic acid were the compounds with highest concentration in both inodorous and common chamomile. The results obtained hereby represent the first and most detailed chemical profile of scentless chamomile so far.
Authors:Khudbudin B. Mulani, Brahmanand M. Kamble, Vijay R. Chandegaonkar and Hemantini A. Deshpande
In the present study, we developed a simple, sensitive, and selective thin-layer chromatographic method for the detection and identification of amitraz using chloranil as chromogenic spray. In the first step, amitraz was hydrolized by using 10% sodium hydroxide solution in water to give dimethylaniline (DMA) and N-2,4-dimethylphenyl- N-methyl formamidine (DMPMF). The dimethylaniline further treated with chloranil in acetone gives blue spot. This spray reagent does not react with other organophosphorous, organochloro, carbamate, and synthetic pyrethroid insecticides. This reagent is sensitive and selective only to amitraz. The constituents of viscera (amino acids, peptides, and proteins) do not interfere with the test. The detection limit for amitraz is about 0.7 µg.
Authors:Aradhana Sharma, Chetan Dhal and Richa Srivastava
The separation and estimation of natural products by chromatographic techniques such as high-performance liquid chromatography (HPLC) and high-performance thin-layer chromatography (HPTLC) are widely preferred. The present work reveals the quantification of morphine (a regulated narcotic agent) in the Ayurvedic formulation ̀Kamini Vidrawan Raś using reversed-phase HPLC (RP-HPLC). The results obtained herein were compared with the results of earlier reported work using HPTLC. The HPTLC separation of morphine was performed on an aluminum-backed layer of silica gel 60 F254 using ethyl acetate–methanol–ammonia solution as the mobile phase, while RP-HPLC was performed on Kromasil C8-column (150 mm × 4.6 mm, 5 µm) using a mobile phase comprising of N-heptane sulfonate sodium salt–acetonitrile (70:30, v/v) at a flow rate of 1.5 mL min-1. The International Conference on Harmonisation (ICH) guidelines were followed for validation of both the chromatographic methods. Both the developed chromatographic methods are simple, rapid, accurate, and sensitive.
Centella asiatica (L.) Urban, Apiaceae, is a medicinal plant, rooted in the Ayurvedic medicine with wide and potent health beneficial activity, and is of great interest to researchers concerning phytotherapy due to its numerous pharmacological activities. In this paper, we have developed a valid thin-layer chromatography (TLC)–densitometric method for the simultaneous determination of the four main C. asiatica active compounds. The additional goal was to achieve the conditions of determination, which would not require spraying with the derivative reagent and subsequent heating ? the factors that greatly deteriorate the validation parameters. In order to do so, we have tested diff erent sorbents (alumina, silica gel, and high-performance thin-layer chromatography [HPTLC] silica gel) and a number of eluents. The best results were obtained with HPTLC silica gel plates eluted with a mixture of CHCl3–MeOH–H2O (10:8:2, v/v) and analysed with a densitometer at λ = 202 nm. The chromatographic method was validated for linearity, limit of detection, limit of quantification, repeatability, intermediate precision, and recovery.
Authors:Pankaj Kumar Jain, Sonika Jain, Swapnil Sharma and Sarvesh Paliwal
The present study was aimed to assess the scientific appraisal of Phoenix sylvestris in the course of pharmacognostical properties and phytochemical parameters, as this has not been done yet. Pharmacognostic study mainly covered the macroscopic and microscopic features of the roots including powder microscopy and revealed the presence of pitted and spiral xylem vessels, lignified xylem fiber, and brown-colored stone cells. Physicochemical constants such as ash value, extractive value, and fluorescence analysis were assessed in the preliminary physicochemical screening. Qualitative analysis revealed the existence of certain chemical constituents such as flavonoids, tannins, steroids, and phenolic compounds. The crude extract of the root was subjected to high-performance thin-layer chromatography (HPTLC) for the separation of components. HPTLC analysis was carried out on the methanolic extract of the root by using toluene–ethyl acetate–formic acid acid (5:4:1, v/v) as the mobile phase at 254-nm detection to quantify the amount of quercetin. The calibration range was 50–250 ng per band (r2 = 0.994). The method was accurate in triplicate results at diff erent standard addition levels with average recovery of 99.40% for quercetin. Limits of detection and quantification were calculated as 20.94 ng per band and 43.90 ng per band, respectively. The quantity of quercetin in the methanolic extract was found to be as 3.13 mg g-1. The developed and validated HPTLC method can be used as a tool for standardization of roots in diff erent formulations using quercetin as the marker.
Authors:Monika Dąbrowska, Kinga Sokalska, Paweł Gumułka, Żaneta Binert-Kusztal and Małgorzata Starek
The effect of omega–3 fatty acids in the human body is multidirectional. They are used in cancer and cardiovascular disease prevention, to stabilize blood pressure, to improve immune function, and in the treatment of concentration disorders. Omega-3 acids, especially eicosapentaenoic and docosahexaenoic acids, are needed for proper human growth. There are several groups of pharmaceutical products containing these compounds, available in pharmacies and markets. Due to a high accessibility of various consumable products such as dietary supplements or food, their quality monitoring is advisable. Therefore, analytical procedures are needed, which will allow the quality of these products to be fast and accurately defined. In our work, thin-layer chromatography (TLC) with densitometric detection was designed for the identification and quantification of the omega–3 fatty acids. The stationary phase constituted TLC silica gel plates activated by AgNO3 solution. As the mobile phase, a mixture of toluene–n-propanol–acetic acid (20:2:0.1, v/v) was used. Detection was performed at 520 nm, after staining with iodine vapor. The method was validated according to the International Conference on Harmonisation (ICH) guidelines. The correctness of the results guarantees good precision and satisfactory recovery (96.59–103.20%). The presented analytical method was used for the determination of omega-3 fatty acids in selected dietary supplements and cooking oils giving satisfactory results.
Authors:Dipshi Singh, Poonam Malik and Ravi Bhushan
The scientific development in the area of enantioseparation during the last few decades has centered on the production of new chiral stationary phases (CSPs) and new chiral derivatizing reagents (CDRs) for use in liquid chromatography, and in particular high-performance liquid chromatography (HPLC) only. Both CSPs and CDRs have several limitations which, in general, are ignored. Little attention has been paid to thin-layer chromatography (TLC) despite its many advantages compared to HPLC in pharmaceutical and drug analysis and the areas of natural products chemistry and organic synthesis, particularly enantioselective synthesis in purification of the product prior to establishing enantiomeric ratio by different method(s). TLC provides a rapid, easy, aff ordable, and simple approach in all these situations. The demonstrated capability and effi- ciency of TLC in direct resolution of the racemate clearly establish its superiority, and the methodology should allow its application in the resolution of several other racemates, irrespective of the functional group, in a very short time along with the recovery of native enantiomers (for further use).
A simple, rapid, quantitative, and validated high-performance thin-layer chromatographic (HPTLC) method has been developed for the determination of ayapanin in the extract of Ayapana triplinervis leaves. Densitometric analysis of ayapanin was carried out in the absorbance mode at 254 nm. The method gave spot at RF = 0.56, corresponding to ayapanin. The limit of detection (54.32 ng) and limit of quantification (168.85 ng) per spot were confirmed with the mobile phase petroleum ether and ethyl acetate in a ratio of 60:40 v/v. Linear regression analysis data for the calibration plot for ayapanin showed good linear relationship with a correlation coefficient (r) of 0.997 in the concentration range of 200 ng to 1000 ng per spot. The method was validated for sensitivity, linearity, accuracy, precision, and specificity as per the International Conference on Harmonization (ICH) guidelines. After development and validation, the method was applied for the isolation of ayapanin from the extract. The isolated ayapanin was characterized by Fourier-transform infrared, nuclear magnetic resonance, and mass spectroscopy techniques.
Authors:Bellahsen Naoufal, Kertész Szabolcs, Pásztory Zoltán and Hodúr Cecilia
Nutrient removal has become one of the key challenges for wastewater treatment facilities all over the world due to the harmful effect of these pollutants on water bodies and ecosystems known by eutrophication, however, most of the currently used technologies are not focused on nutrients recovery from wastewater. Recently, using agricultural waste/by-products for adsorption of nutrients acquired more interest because of their abundant availability, low-cost, high efficiency and eco-friendly advantages and this method may become more environmentally sustainable through maximizing removal while delivering nutrient and energy recovery technologies with economically attractive return on investment.
This review investigates the application of agricultural waste/by-products as bio-sorbent for phosphate, ammonium and nitrate removal with a focus on the modification methods and the process mechanism including influent parameters, kinetics and isotherms.
Authors:Pintu B. Prajapati, Kunjan B. Bodiwala and Shailesh A. Shah
A robust and eco-friendly stability-indicating high-performance thin-layer chromatography (HPTLC) method was developed for the stability study of thiocolchicoside using analytical quality-by-design approach. Full factorial design was used for screening potential variables affecting method development. Box‒Behnken design was used subsequently for investigation of the main, interactive, and quadratic effects of these variables on response. Four potential variables were selected on the basis of scientific knowledge for the development of a method for the stability study of thiocolchicoside. The selected potential factors, namely, volume of water (mL), saturation time (min), migration distance (mm), and volume of mobile phase (mL) were screened by 24 full factorial designs by selecting resolution as a critical method attribute. Pareto chart analysis showed that 3 variables, namely, volume of water (mL), saturation time (mm), and migration distance (mm), out of 4 potential variables were significantly affecting the response variable (resolution). Optimization with response surface methodology further clarified the relationship between critical variables and resolution using Box–Behnken design. The experimental design model was found to be quadratic, and the design space was developed on the basis of suggested model for optimization of critical method variables for maximum desirable resolution and for the development of a control strategy of the HPTLC method for the stability study of thiocolchicoside. The developed method was validated for linearity, range, specificity, precision, accuracy, limit of detection and limit of quantification as per the International Conference on Harmonization guidelines (ICH) Q2 (R1). The developed method was applied for the estimation of thiocolchicoside in its pharmaceutical dosage forms. The degradation products formed in acidic and alkaline media were isolated and characterized by their infrared (IR), nuclear magnetic resonance (NMR), and mass spectral data.
A WRB diagnosztikai elemeit és a felszínborítási adatokat kombinálva a talajokat ért antropogén hatás mértéke szerint négy csoportot alkottunk: 1. nincs talaj, 2. antropogén eredetű talaj, 3. természetes talaj, de lényeges antropogén bélyegekkel, illetve 4. természetes talaj. A négy csoport valamelyikéhez egyértelműen hozzárendelhető a felszínborítási osztályok mindegyike. Az általunk kidolgozott módszer segítségével értékeltük Magyarország talajtakarójának természetességét. Az ország területének 2%-án nem számolhatunk a FAO által definiált értelemben talaj létével, 6%-án antropogén talajok várhatók (Anthrosol, vagy Technosol), 66%-án a természetes talajok antropogén átalakítottsága eléri a WRB diagnosztikai határértékeit, és mindössze 26% azon talajok aránya, amelyekben antropogén hatások a diagnosztikában nem jelennek meg, azaz természetes vagy természetközeli állapotúként értékelhetők. Talajtípusok tekintetében legnagyobb mértékű emberi hatással a csernozjomok, réti és öntés talajok esetében számolhatunk, míg természetközeli állapotú talajok legnagyobb kiterjedésben a kőzethatású és váztalajokon maradtak fenn. A területi különbségek is jelentősek: míg legnagyobb arányban a Hajdúságon és a Körös-Maros közén találunk antropogén hatásokkal érintett talajokat, addig a természetközeli állapotú talajok aránya az Északi-középhegység egyes hegyvidéki területein a legnagyobb.
Módszerünk csak becslésre alkalmas, mégis jó áttekintést ad a hazai talajok antropogén átalakítottságának mértékéről, az emberi tevékenység, mint hatodik talajképző tényező jelentőségéről, intenzitásának térbeli eloszlásáról, amely a hazai talajtani adottságoknak egy eddig kevéssé vizsgált aspektusa.
Authors:Gyöngyi Vastag, Suzana Apostolov, Biljana Kaurinovic and Ljubica Grbovic
It has been established that the selected acetamide derivatives fulfill Veber’s and Pardridge’s rule of good bioavailability. The lipophilicity as a crucial molecular descriptor of biological activity for acetamide derivatives was determined by reversed-phase thinlayer chromatography (RP-TLC) in different mixtures of water and organic modifiers (ethanol and tetrahydrofuran). Also, lipophilicity was examined mathematically, by using relevant software packages. The effects of the substituent on the lipophilicity of acetamide derivatives were analyzed. By applying linear regression analysis and 2 multivariate methods (cluster analysis and principal component analysis), the obtained chromatographic parameters, RM0 and m, of the examined acetamides were correlated with the standard measure of lipophilicity, log P, important pharmacokinetic predictors, and selected toxicity parameters. The obtained results confirmed that chromatographic parameters, RM0 and m, determined by RP-TLC, could be successfully used for describing the lipophilicity and estimation of pharmacokinetics and the toxic effects of the studied acetamide derivatives.
A talajok szélerózióval szembeni érzékenységének egyik gyakran használt mérőszáma az erodálhatósági faktor, amelynek mérését többféle módszerrel is el lehet végezni. Mivel e mérőszám hazai használhatóságának vizsgálata eddig még nem történt meg, ezért munkánk alapvető célja annak megválaszolása volt, hogy hazai talajok esetében mennyire használható ez az érték, valamint, hogy a vizsgált talajtulajdonságok milyen mértékben és milyen irányban hatnak az EF-index értékének változására. Vizsgáltuk továbbá azt is, hogy az amerikai talajokra kidolgozott, az EF- index becslését lehetővé tevő egyenlet adaptálható-e hazai talajok esetén.
Az EF-index meghatározásához szitarázó gépet, a kapott adatok feldolgozásához, valamint a vizsgált paraméterek közötti kapcsolatok feltárásához statisztikai módszereket használtunk.
A mérések eredményei alapján elmondhatjuk, hogy a vizsgált nyírségi talajminták kb. 50%-a az erősen veszélyeztetett kategóriába tartozik. Az EF értékét legerőteljesebben a talajok mechanikai összetétele, kisebb mértékben pedig a szervesanyag- és CaCO3-tartalma befolyásolta. A kapcsolat minden esetben szignifikáns volt.
Elemzéseinkkel az is kimutatható volt, hogy a vizsgált nyírségi talajok esetében a FRYREAR et al., által kidolgozott egyenlet nem alkalmas arra, hogy kielégítően becsülje a talajokra jellemző EF- index értékét.
Authors:M. Dąbrowska, Ł. Komsta, W. Opoka and M. Starek
The specific aim of this investigation was to study the kinetics of the degradation of cefazolin, cefaclor, cefuroxime axetil, and cefepime in aqueous solution, in the presence (or absence) of various redox agents (iodine solution, potassium permanganate, hydrogen peroxide, sodium thiosulfate, and ascorbic acid) as a function of temperature. Various factors, such as concentration of the analyzed compounds and redox agents, storage time, and temperature, were analyzed. The degradation process of chosen antibiotics was observed chromatographically and fitted to the kinetic models, obtaining model parameters (k, t0.1, t0.5). Principal component analysis (PCA), parallel factor analysis (PARAFAC), and hierarchical cluster analysis (HCA) methods were carried out to interpret the dependencies between these factors on the drug stability.
Authors:Renuka S. Joshi, Shubhada A. Tamhankar and Anuradha S. Upadhye
An ingredient of ‘Dasamoola’ and ‘Laghupanchamoola’ group of drugs, the source of ‘Brihati’ has been controversial. Although the dried root of Solanum anguivi is considered as the source of the drug ‘Brihati’ according to the Ayurvedic Pharmacopoeia of India, closely related and morphologically similar few species like Solanum torvum, Solanum melongena, Solanum incanum, and Solanum insanum are known as its substitutes. In the present study, a high-performance thin-layer chromatography (HPTLC) method was developed and validated for the chemoprofiling and quantitative estimation of glycoalkaloid solamargine from 5 species of the genus Solanum as well as market samples. The developed method was precise, accurate, robust, specific, and linear. The results showed that S. incanum has the highest content of solamargine, followed by S. insanum. Out of the 9 market samples analyzed, solamargine was detected only in 3 samples. Unsupervised pattern recognition techniques, such as principal component analysis and hierarchical cluster analysis, were used to analyze the complex fingerprint patterns and to predict the grouping of samples. The method clearly segregated the field and market samples. Our study is the first attempt to evaluate the drug ‘Brihati’ and the market samples using HPTLC.
Authors:Mohamed S. Abdel-Aziz, Mosad A. Ghareeb, Amal M. Saad, Laila A. Refahy and Ahmed A. Hamed
Eight compounds were isolated and identified from the soil-inhabiting fungus Aspergillus fumigatus 3T-EGY, namely, stearic acid (1), α-linolenic acid (2), physcion (3), di-(2-ethylhexyl) phthalate (4), 2,4,5,17-tetramethoxy pradimicin lactone (5), 3,5-dihydroxy-7-O-α-rhamnopyranoyl-2H-chromen-2-one (6), juglanthraquinone A-5-O-d-rhodosamine-(4′→1″)-2-deoxy-d-glucose (4″→1″′)-cinerulose B (7), and micropeptin (8). Their structures were determined on the basis of one-dimensional (1D-) and two-dimensional nuclear magnetic resonance (2D-NMR) [1H-, 13C-NMR, 1H-1H COSY (COrrelated SpectroscopY), and 1H-13C HMBC (Heteronuclear Multiple Bond Correlation) spectroscopy]. Compound 7 showed moderate in vitro antimicrobial activity against three pathogenic strains with inhibition zones values were ranged from 9.0 to 10.66 mm compared to neomycin as a positive control with inhibition zones values were ranged from 14.0 to 19.0 mm.
Authors:Sally Nasser, Ismail Salama, Samia M. Mostafa and Mohamed Saleh Elgawish
The discovery of potent antidiabetic drugs is of necessity owing to the rapid prevalence of diabetes worldwide. The investigation of a new separation method for the simultaneous determination of combined antidiabetic drugs is thus an essential issue to cover the usual demands of simple analytical methods for the routine analysis. Therefore, herein, simple and fast chromatographic methods were established for synchronized determination of metformin (MET) and dapagliflozin (DAP), a mixture approved recently by the Food and Drug Administration (FDA) for diabetes therapy. In high-performance liquid chromatography (HPLC), a Water-pak C18 column was used as the stationary phase with an isocratic mobile phase composed of 10 mM NaH2PO4 (pH 3.5 adjusted using ortho-phosphoric acid)‒acetonitrile (65:35, v/v) containing 0.1% trimethylamine at a flow rate of 1.2 mL min−1 and a wavelength detector set at 225 nm. In high-performance thin-layer chromatography (HPTLC), separation was achieved on pre-coated silica gel 60 F254 aluminum plates using acetonitrile‒ammonium acetate 10%‒acetic acid (9:0.9:0.1, v/v). The proposed methods were validated in the light of the International Conference on Harmonization (ICH) guidelines, and it was found that the two chromatographic methods are accurate, precise, and linear in the range of 2–20 μg mL−1 and 1–10 μg per spot for DAP and 20–400 μg mL−1 and 10–100 μg per spot for MET by HPLC and HPTLC, respectively. The methods achieved a reasonable sensitivity as shown by the low limit of detection ranging from 0.58 to 6.1 μg mL−1 and 0.314 to 3.1 μg per spot for HPLC and HPTLC, respectively. The validated methods succeeded in detecting the cited drugs in pharmaceutical formulation without interference of excipients. Although HPLC method is the most applicable method, HPTLC method exhibits a superior sensitivity and is cheap and fast allowing the determination of a large number of samples in due time.
Authors:Nafiu Aminu, Siok-Yee Chan, Nasir Hayat Khan and Seok-Ming Toh
A new high-performance liquid chromatographic (HPLC) method for determination of triclosan (TCS) and flurbiprofen (FBP) was successfully developed and validated at a single wavelength. The method involves extraction of the targeted drugs from nanogels and simulated saliva by using methanol as the extractant. The Agilent ZORBAX SB-C18 column (5 μm, 4.6 × 250 mm) was used for the chromatographic separations. The effects of various parameters were extensively evaluated and optimized. The optimal HPLC conditions were acetonitrile and 0.001 M citric acid (90:10, v/v) with a pH of 3.24 as the mobile phase, at a 0.3 mL/min flow rate under isocratic elution mode. Excellent sensitivity and specificity were achieved by ultraviolet (UV) detection at 242 nm. The method also demonstrated excellent linearity within the test range of 10–100 μg/mL with the correlation coefficient (R2) of 0.9998 for both the analytes. The practical applicability of the method was demonstrated by recovering TCS and FBP from nanogels and simulated saliva. The recovery of the analytes from the nanogels and the spiked simulated saliva samples was in the range of 97–98% and 96–99%, respectively, and their respective relative standard deviation (RSD) was less than 0.9% in both cases. System suitability parameters were found to be within acceptable limits. The method is simple, specific, and precise, and to the best of our knowledge, it is the first reported validated quantitative HPLC method for the concurrent determination of TCS and FBP in a pharmaceutical dental product. The method can be useful in the routine quality control analysis of dental formulations with TCS and FBP contents or products with a similar composition.
In the present study, a design-of-experiment (DoE) approach Was used to determine optimized mobile-phase compositions for the development of a high-performance thin-layer chromatography (HPTLC) method for the simultaneous estimation of berberine chloride (BER-H) and galangin (GAL) in Tinospora cordifolia M. and Alpinia galanga L., respectively, and their formulations. A Box‒Behnken design (BBD) was used to optimize the compositional parameters and evaluate the main effect, interaction effects and quadratic effects of the mobile-phase compositions on the retardation factor (RF) of both drugs. HPTLC separation was performed on aluminum plates pre-coated with silica gel 60 F254 as the stationary phase, using toluene‒ethyl acetate‒formic acid (3:6:1, v/v) as the mobile phase at a wavelength of 267 nm. A sharp and well-resolved peak was obtained for BER-H and GAL at RF values of 0.17 ± 0.01 and 0.82 ± 0.01, respectively. The calibration curve was in the range of 200–1200 ng per band for both BER-H and GAL, with r2 = 0.984 and r2 = 0.980, respectively. Statistical insight was achieved with analysis of variance (ANOVA). The method was validated for linearity, accuracy, precision, limit of detection, limit of quantification, robustness, and specificity. To provide a better visualization of the statistically significant factors derived from the statistical analysis, the perturbation plot and response surface plot for the effect of independent variables on the RF of BER-H and GAL were evaluated. The developed HPTLC method was found to be simple, accurate, precise, sensitive, and specific for the simultaneous quantification of berberine chloride and galangin in Tinospora cordifolia M. and Alpinia galanga L., respectively, and their
Authors:Swaha Satpathy, Arjun Patra and Bharti Ahirwar
The present study was undertaken to develop and validate a novel high-performance thin-layer chromatography (HPTLC) method for the quantification of apigenin and luteolin in Hygrophila spinosa. Separation was carried out on HPTLC plates using the mobile phase toluene‒ethyl acetate‒formic acid (6.0:4.0:1.0, v/v), and detection was achieved at 349 nm. The method was validated by the International Conference on Harmonization (ICH) guidelines. The calibration range was 80–560 ng per band (R2 = 0.997) for apigenin and 40–280 ng per band (R2 = 0.998) for luteolin. The method was accurate in triplicate results at different standard addition levels with average recoveries of 99.94 and 100.01% for apigenin and luteolin, respectively. The limits of detection and quantification were 6.25 and 18.95 ng, respectively, for apigenin, and 2.36 and 7.55 ng, respectively, for luteolin. The quantity of apigenin was 8.84 and 15.67 mg g−1 in the alcoholic extract and its ethyl acetate fraction, respectively. The luteolin contents in the alcoholic extract and ethyl acetate fraction were 0.172 and 0.464 mg g−1, respectively. Further, luteolin was found to be present only in the flower (6.87 mg g−1) extract, but apigenin was detected in the husk (2.38 mg g−1), flower (7.97 mg g−1), spine (1.01 mg g−1), fruits (0.60 mg g−1), and leaf (4.28 mg g−1). The developed method could be used for the quality control analysis and quantification of apigenin and luteolin in herbal preparations containing H. spinosa.
Authors:Asmaa A. El Zaher, Ehab F. Elkady, Ola M. El Houssini and Hind E. El Ghwas
New stability-indicating thin-layer chromatography (TLC)‒densitometry and reversed-phase liquid chromatography (RP-LC) methods for the determination of dabigatran etexilate mesylate (DEM) in the presence of its risky degradation products were developed and validated. The first method was a simple economic TLC‒densitometric method, where separation was achieved on aluminum plates pre-coated with silica gel gel 60 F254 using methanol‒ethyl acetate‒benzene (15:20:20, v/v) as the mobile phase. The method gave a compact and well-resolved spot for DEM. Quantification was performed at 225 nm over a concentration range of 0.3–3 μg per spot with a mean percentage recovery of 98.93 ± 0.80. The second method was an RP-LC method. Short analysis time and good efficacy were achieved on a BetaSil C18 (4.6 mm × 250 mm, 5 μm) column using gradient elution. The flow rate was 1.5 mL min−1, and ultraviolet (UV) detection was carried out at 310 and 225 nm using time programming. The responses were linear over a concentration range of 5–100 μg mL−1 with a mean percentage recovery of 99.25 ± 0.74. The limit of quantification and limit of detection were 0.10 and 0.03 μg per spot for the TLC and 0.287 and 0.086 μg mL−1 for the LC method. The proposed methods were used for the identification and quantitative determination of DEM in bulk powder, pharmaceutical dosage forms, and in the presence of degradation products. The International Conference on Harmonization (ICH) guidelines were applied for validation of the methods.
Authors:L. Velkoska-Markovska, B. Petanovska-Ilievska, M. S. Jankulovska and U. Ilievski
This study presents the development and validation of a new reversed-phase high-performance liquid chromatography (RP-HPLC) method for simultaneous determination of captan, folpet, and metalaxyl residues in table grape samples with ultraviolet–diode array detection (UV–DAD). Successful separation and quantitative determination of analytes was carried out on LiChrospher 60 RP-select B (250 × 4 mm, 5 μm) analytical column. Mixture of acetonitrile–0.1% formic acid in water (65:35, v/v) was used as a mobile phase, with flow rate of 1 mL/min, constant column temperature at 25 °C, and UV detection at 220 nm. The target residues were extracted with acetone by ultrasonication, followed by a cleanup using liquid–liquid extraction (LLE) and solid-phase extraction (SPE). The obtained values for multiple correlation coefficients (R2 > 0.90), relative standard deviation (RSD) of retention times, peak areas and heights (RSD ≤ 2.25%), and recoveries ranging from 90.55% to 105.40%, with RSD of 0.02% to 5.37%, revealed that the developed method has a good linearity, precision, and accuracy for all analytes. Hence, it is suitable for routine determination of investigated fungicides in table grape samples.
Authors:Peiwu Geng, Xinhua Luo, Xiufa Peng, Zixia Lin, Wenhao Chen, Jin Zhang, Congcong Wen, Lufeng Hu and Siyi Hu
Eupatilin, mainly derived from Artemisia asiatica (Asteraceae), is an O-methylated flavone with various bioactivities. In the present study, a validated ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method was established for the quantification of eupatilin in rat plasma with the internal standard (IS) of tussilagone and the protein precipitation of plasma samples was performed using acetonitrile–methanol (9:1, v/v). The eupatilin and IS were eluted separately on a UPLC BEH C18 column (2.1 mm × 100 mm, 1.7 μm) with the gradient mobile phase consisted of 0.1% formic acid and acetonitrile. The protonated analytes were quantified by multiple reactions monitoring (MRM) mode with an electrospray ionization (ESI) source operated in positive ion mode. The calibration plots were found to be linear over the range from 2 to 1000 ng/mL for eupatilin in rat plasma. Both of the intra-day and inter-day precision variations (RSDs) were ≤13%. The recoveries of eupatilin in rat plasma were between 83.7% and 94.6%, and the accuracy of the method ranged from 95.8% to 107.6%. In addition, the validated method was applied to pharmacokinetic study of eupatilin after an intravenous dose of 2 mg/kg to rats.
The present paper reports a validated high-performance thin-layer chromatography (HPTLC)‒densitometric method for the simultaneous quantification of phenolic (ferulic acid and caffeic acid) and terpenoid (β-sitosterol and lupeol) markers in Convolvulus pluricaulis Choisy. According to Ayurveda, it is commonly known as ‘Shankhpushpi’ due to its ‘Conch’ or ‘Shankh’-shape flower. The plant species, viz., Clitoria ternatea L., Evolvulus alsinoides (L.) L., and Tephrosia purpurea (L.) Pers., also having similar flowers are reported as its adulterants/substitutes. This creates a problem in its quality and efficacy in the commercial drug market of India. Therefore, a HPTLCmethod was performed on a pre-coated silica gel 60 F254 plate with the aforesaid markers. The solvent system toluene–ethyl acetate–formic acid (8.5:1.5:0.1) was determined to be the best system for the simultaneous separation of caffeic acid, ferulic acid, β-sitosterol, and lupeol at RF values of 0.14, 0.29, 0.48, and 0.63, respectively. A densitometric scanning profile of all the samples at 580 nm showed peaks for all the four markers of varying heights in the samples, except the absence of caffeic acid in Tephrosia purpurea. The developed method was standardized and validated for the quantification of active principal-based quality-control markers in terms of precision, accuracy, linearity, recovery, and repeatability. It will help to maintain batch-to-batch consistency and identification of adulterants/substitutes in raw materials during production of drug in the pharmaceutical units.
A model process, previously developed in a series of studies, allows for the transfer of thin-layer chromatography (TLC) methods for qualitative screening of counterfeit drug products published in the Global Pharma Health Fund (GPHF) Minilab manual and US Food and Drug Administration (FDA) Compendium of Unofficial Methods for Screening of Pharmaceuticals by TLC to quantitative high-performance TLC (HPTLC)–densitometry methods. This article describes HPTLC–densitometry methods developed and validated according to this model process for pharmaceutical products of amiodarone HCl, carvedilol, doxylamine succinate, magnesium salicylate, metoprolol succinate, nebivolol HCl, and salicylamide, for which qualitative screening methods have not been published in the Minilab manual or FDA Compendium. These methods use relatively inexpensive and nontoxic “green solvents” for sample and standard solution and mobile phase preparation, Merck Premium Purity silica gel 60 F254 plates, automated standard and sample solution bandwise application, and automated densitometry for the assessment of peak purity and identity and quantification. Corresponding to the quantitative HPTLC–densitometry methods, qualitative TLC screening methods for these drug products were developed and posted online with open access as supplements to the FDA Compendium.
Authors:Hayun Hayun, Rina Rahmawati, Yahdiana Harahap and Santi Purna Sari
A specific, very rapid, and sensitive ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method for quantitative analysis of curcumin in human plasma has been developed and validated. Diazepam was used as internal standard (IS). The analytes were isolated using liquid–liquid extraction method with the mixture of ethyl acetate–methanol (95:5). The organic solvents were evaporated, reconstituted in mobile phase, and injected to UPLC completed with UPLC BEH C18 column 1.7 μm, 2.1 × 100 mm Acquity® Waters as stationary phase, mixture of 0.15% formic acid–acetonitril (50:50, v/v) as mobile phase, and flow rate of 0.5 mL/min and detected in positive ionization mode tandem mass spectrometer operated in multiple reaction monitoring (MRM). The MS/MS ion transitions monitored were m/z 369.05 → 176.95 and 284.95 → 193 for curcumin and IS, respectively. The retention times for curcumin and IS were 1.7 and 1.4 min, respectively, and the linearity range was 1–100 ng/mL with a coefficient correlation (r) of 0.999 and lower limit of quantitation (LLOQ) of 1 ng/mL. The relative standard deviation (RSD) values of the intra- and inter-assay precisions of the method were below 8.3% and 12.7%, respectively, while the accuracy ranged from 89.5 to 98.7% and the extraction recovery of curcumin and IS was up to 86.6%. The data presented show that the method provides specific, very rapid, sensitive, precise, and accurate measurements of curcumin concentrations in human plasma.
The wind drift and evaporation losses (WDEL) are high in arid, semi-arid and windward areas, and reduce the efficiency of the sprinkler irrigation system; therefore, this study was carried out in order to achieve a practical criterion and provide a relationship for accurate estimation of the wind drift and evaporation losses in different atmospheric conditions. The experiments were done at the Meteorological Station of the Faculty of Agriculture of Ferdowsi University of Mashhad using a line-source sprinkler irrigation system based on the single sprinkler installation method. To achieve the objectives of this plan, factorial experiment was performed on PGP sprinkler with regard to the two factors, the pressure of the sprinkler function (with three levels 1.6, 2.5 and 3.4 bar) and the diameter of the nozzle (with three levels of 4, 5 and 6 mm) with three replications (morning, noon and night). Assessing the result of the data variance analysis showed that the effects of pressure, aperture diameter, and time on the wind drift and evaporation losses are not significant. Investigating the main effects of these factors showed that the effect of aperture diameter on irrigation losses is significant at the level of the 1%. In order to further investigate, the comparison of mean losses data in three aperture diameter was done using Duncan′s test. The results indicated that aperture 4 with the losses of 44% had a significant difference with other diameters. This result suggests an increase in losses for smaller diameters due to the small droplets and the increase in wind drift. Also, the comparison of the mean losses data in three times showed that irrigation at noon with the losses of 44% had a significant difference compared to other times due to a significant increase in temperature and radiation of the sun and saturation vapor pressure deficit, and there is no significant difference between morning and evening irrigation. Also, analysis of variance showed that the effect of water pressure change between 1.6 and 4.3 bar does not have a significant effect on the WDEL in this sprinkler. In general, the results showed that increasing wind speed increases the losses of evaporation and wind. Also, this study suggested that changing the irrigation time in areas with hot and dry days, from day to night in summer, leads to a significant reduction of the wind drift and evaporation losses.
In practice, there is a demand for quick characterization of rheological properties of food materials. The exact model calculation requires complex and long-term mathematical process. In this work, a simple, quick linearization method – the Peleg linearization – is discussed and is compared with the Prony series method. In the Peleg linearization only two constants are used, one of them gives the initial rate of relaxation or creep and the second one gives the equilibrium value of relaxing force or of creeping strain. The Prony series approach the relaxation and creep with the sum of two or more exponential functions and equilibrium values. Both methods give the same equilibrium values for both the relaxation and creep of wine gums and apple. The initial increasing rate of creep is higher by the Peleg linearization and lower by the Prony series. At relaxation the initial decreasing rate is lower by the Peleg linearization and higher by the Prony series.
A high-performance thin-layer chromatography (HPTLC) method has been developed for the simultaneous estimation of abacavir sulfate (ABC), lamivudine hydrochloride (LAM), and dolutegravir sodium (DTG) in an in-house physical mixture and it was validated as per the International Conference on Harmonization (ICH) guidelines. A 23 full factorial design was utilized to aid in method development and optimization. Effective chromatographic separation was achieved using pre-coated silica gel aluminum HPTLC plate 60 F254 as the stationary phase and ethyl acetate‒ethanol‒ acetone‒ammonia (4.478:0.740:0.50:0.15, v/v) as the mobile phase. The optimized chamber saturation time was kept at 30 min. Densitometric scanning was performed in the absorbance–reflectance mode at a detection wavelength of 267 nm. Quality by design approach was applied to evaluate the effect of all three factors, i.e., volume of ethyl acetate, volume of ethanol, and volume of acetone, on the chromatographic response retardation factor (RF) of each drug. The RF values of ABC, LAM, and DTG were found to be 0.65, 0.34, and 0.26, respectively. The calibration curves were found to be linear by taking independently weighed standard solutions with concentrations of 4.8, 7.2, 9.6, 12.0, and 14.4 μg per band for ABC, 2.4, 3.6, 4.8, 6.0, and 7.2 μg per band for LAM, and 0.4, 0.6, 0.8, 1.0, and 1.2 μg per band for DTG. The limit of detection and limit of quantification values were found to be 0.9972 μg per band and 3.0218 μg per band for ABC, 0.2544 μg per band and 0.7711 μg per band for LAM, and 0.1004 μg per band and 0.3043 μg per band for DTG, respectively. The % recoveries of the drugs by the developed method were found in the range of 98.0955–100.9813% for ABC, 98.2616–99.9900% for LAM, and 98.4666–101.3000% for DTG, respectively. The proposed method was found to be novel, simple, accurate, reproducible, and robust. The developed HPTLC method was successfully applied for the quantitative determination of ABC, LAM, and DTG in an in-house physical mixture.
Authors:Meghna Patel, Mihir Raval, Mansi Joshi and Jalpa Sanandia
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 RF 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 (r2 = 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.
The aim of this study was to identify and determine by means of gas chromatography–flame ionization detector (GC–FID) and gas chromatography–mass spectrometry (GC–MS) method the volatile compounds of essential oils obtained from three varieties of narrow-leaved lavender grown in the field and in in vitro cultures. The essential oils were isolated by hydrodistillation in Deryng apparatus. It was found that the analyzed essential oils varied in terms of chemical composition depending on the variety and conditions of growth. Sixty-four to 87 different compounds were identified in the oils. Essential oils of all 3 varieties obtained in in vitro cultures contained large amounts of borneol (13–32%). This compound was also dominant in plants obtained from in vivo conditions in varieties Ellagance Purple (11%) and Blue River (13%), and in the Munstead variety, the dominant compound was linalool (13%). High concentration of epi-α-cadinol (10–20%) was found in essential oils obtained from in vitro cultured plants. Globulol was found in high concentration (10%) in the Munstead variety grown in in vitro conditions. However, significant quantitative and qualitative differences were found with respect to composition of essential oils obtained from plants grown in the field and in vitro conditions. There was a lack of (E)-β-ocimene, 3-octyn-2-one, 1-octen-3-yl acetate, sabina ketone, pinocarvone, trans-carveol, nerol, epi-longipinanol, or humulene epoxide II. In comparison to oils obtained from field-grown plants, the oils isolated from plants grown in in vitro conditions contained the less volatile compounds identified in the final stage of GC–FID and GC–MS analysis, i.e., thymol, carvacrol, γ-gurjunene, trans-calamene, α-calacorene, khusinol, and 8-cedren-13-ol.
Authors:Gergely Ujvári, Andrea Borsodi, Júlia Margit Aszalós, Melinda Megyes, Márton Mucsi, Attila Szabó and Károly Márialigeti
E tanulmány célja egy martonvásári hosszútávú tartamkísérlet trágyázás nélkül művelt kukorica monokultúra talajában fellelhető baktériumközösségek filogenetikai diverzitásának és anyagcsere potenciáljának a felmérése volt. A kutatás során NGS és MicroResp™ technikával vizsgáltuk a művelt és a természeteshez közeli állapotú talajok mikrobiális jellemzőit.
Az NGS adatai alapján a kukorica monokultúra szántott rétegének mintáinak baktériumközösség szerkezete nagyfokú hasonlóságot mutatott egymással, és elkülönült a löszpusztagyep A és C rétegéből formálódó csoporttól, míg a kukorica monokultúra C szintjéből származó minta élesen elvált a többitől. A gyepek talajában nem találtunk nagyobb bakteriális taxonómiai diverzitást, mint a művelt talajokban.
A MicroResp™ mérés alapján megállapítottuk, hogy a természeteshez közeli állapotú talajok felszínhez közeli (A) rétegében kiugró a mikrobiális aktivitás mértéke. A kukorica monokultúrából származó A szint minták mikrobiális aktivitási mintázata egymáshoz hasonló volt, a C rétegből származó minták külön csoportot képeztek.
Eredményeink alapján tehát a hosszú távú tartamkísérletbe vont művelt talajok baktériumközösségeinek filogenetikai diverzitása és metabolikus potenciálja jelentősen eltért a löszpusztagyep mintákétól.
Authors:M. Y. Guida, F. E. Laghchioua and A. Hannioui
This article deals with fast pyrolysis of brown algae, such as Bifurcaria Bifurcata at the range of temperature 300–800 °C in a stainless steel tubular reactor. After a literature review on algae and its importance in renewable sector, a case study was done on pyrolysis of brown algae especially, Bifurcaria Bifurcata. The aim was to experimentally investigate how the temperature, the particle size, the nitrogen flow rate (N2) and the heating rate affect bio-oil, bio-char and gaseous products. These parameters were varied in the ranges of 5–50 °C/min, below 0.2–1 mm and 20–200 mL. min–1, respectively. The maximum bio-oil yield of 41.3wt% was obtained at a pyrolysis temperature of 600 °C, particle size between 0.2–0.5 mm, nitrogen flow rate (N2) of 100 mL. min–1 and heating rate of 5 °C/min. Liquid product obtained under the most suitable and optimal condition was characterized by elemental analysis, 1H-NMR, FT-IR and GC-MS. The analysis of bio-oil showed that bio-oil from Bifurcaria Bifurcata could be a potential source of renewable fuel production and value added chemicals.
In this reported study, a direct high-performance thin-layer chromatographic (HPTLC) method was developed to qualitatively detect and quantitatively determine glycerol in Antarctic krill for the first time. This procedure was based on the extraction of glycerol by ultrasonic solvent extraction with anhydrous ethanol, silica-gel column chromatographic separation, HPTLC detection and quantification using methylene chloride–methanol (5:1, v/v) as the developing solvent and alkaline potassium permanganate as chromogenic agent. The content of glycerol was 1.3725 ± 0.218 mg/g in freeze-dried Antarctic krill. The structure of glycerol in the Antarctic krill was subsequently determined by gas chromatography–mass spectrometry (GC–MS) which verified the presence of the material in the krill. The HPTLC method exhibited excellent accuracy with a recovery of 90.1–103.3% and good precision with a relative standard deviation (RSD) of 1.59–4.84%. The results clearly exhibited the applicability of the proposed for quantifying glycerol in Antarctic krill.
Authors:Márta Birkás, Igor Ðekemati, Zoltán Kende, Zoltán Radics and András Szemők
Jelen dolgozatban két célt jelöltünk ki. Az első cél a talajművelés hazai fejlődését nehezítő és elősegítő körülmények elemzése, egyúttal a karcagi kutatási eredmények kiemelése volt. A fejlődést az ún. sokszántásos művelés alkalmazásától (1800-as évektől) napjainkig kísértük nyomon. A második cél négy jeles előd munkásságának felidézésével példát kívántunk adni a hazai talajművelés fejlesztéséért tett erőfeszítésekről.
A talajművelés fejlődésének voltak akadályai úgy, mint egyes ökológiai körülmények – nehéz művelésű talajok, időjárási szélsőségek –, körülményekhez alkalmas vonóerő- és művelőgép hiány, háborúk, a szaktudás jelentős elmaradása a Nyugat-európaihoz hasonlítva, továbbá a talajok minőségét veszélyeztető művelési szokások kialakulása és tartós fennmaradása. A fejlődést előmozdították az európai kitekintésű szakírók, a szaksajtó a segítő cikkek, szakmai viták és az újdonságok közlésével, a kísérleti intézetek létrejötte, a kísérletezés megindulása. A tudományok előrehaladása a talajok és a művelés egzakt leírását, új, hazai termőhelyekre alkalmas módszerek kidolgozását tette lehetővé.
A Karcagi Kutatóintézet megalakulásának kezdetétől nem csak bekapcsolódott a talajművelés és kapcsolódó tudomány területek fejlesztésébe, hanem tevékenységével új, nagy térségben hasznosítható témákat munkált ki. A vonatkozó publikációk elemzése során megállapítható, hogy
- A Karcagi Kutatóintézetben kimunkált eredmények a tudomány előrehaladását és a gyakorlat szemléletváltását az ország más termőhelyein működő intézményekkel – Mosonmagyaróvár, Keszthely, Martonvásár, Kompolt, Gödöllő, Nyíregyháza, Debrecen, Szeged – harmóniában, gyakran együttműködésben segítették elő. Ezzel együtt a térségi talajok jobbítása az első évektől napjainkig kiemelt feladat maradt.
- A térségben akut művelési és talajjavítási feladatok megoldásával párhuzamosan folyt a csernozjom, réti és szikes talajok tulajdonságainak megismerése, valamint adott művelési beavatkozások hatásainak értelmezése.
- A nehéz művelésű talajok állapotának javítására kidolgozott periódusos mélyművelési rendszer országos elterjedését előbb állami támogatás, majd az alkalmazók kedvező tapasztalatai mozdították elő.
- A talajállapot javulása a művelési rendszerek ésszerűsítését, a művelési mélység okszerű – ökonómiai szempontból is kedvező – váltogatását tette lehetővé.
- A nagy agyagtartalmú és a kémiailag hibás talajokon bizonyítást nyert a mélyítő művelés, a trágyázás és talajjavítás együttes alkalmazásának hasznossága.
- A mélyművelési módszerek, valamint a magágy minőség változatok vizsgálatainak eredményei között a talajvédelmi ajánlások napjainkban is megállják a helyüket.
- A fenntartható fejlődéssel harmóniában lévő talajhasználati, talaj- és környezetvédelmi, vízgazdálkodási kutatások eddig elért eredményei a további feladatokat alapozzák, és kiszélesedésüket segítik.
- A Karcagi Kutatóintézet fejlődésre ösztönző környezetében bontakozott ki Sipos Sándor és Nyiri László tudós szakírók munkássága. A gyakorlatot segítő tudományos eredményeik, cikkeik és könyveik mellett számos olyan tanítványt neveltek ki, kik ma a talajművelés és a kapcsolódó tudományterületek meghatározó személyiségei.
Authors:Imre Cseresnyés, Tünde Takács, Ramóna Kovács, Anna Füzy and Kálmán Rajkai
Tenyészedény-kísérletben vizsgáltuk, hogy a szárazságstressz és az arbuszkuláris mikorrhiza gomba (AMF) oltás milyen változást okoz a búza gyökérnövekedésében, és ez hogyan követhető nyomon a gyökér–talaj rendszer elektromos kapacitásának (CR) in situ mérésével.
A kísérletet randomizált blokk elrendezésben végeztük két búzafajtával (Mv. Hombár őszi és TC 33 tavaszi), kétféle öntözéssel (optimális és szárazságstressz) és kétféle oltással (oltatlan és AM-gombával oltott), 12 ismétlésben. A tenyészidőszakban monitoroztuk a CR-t, valamint mértük a sztómakonduktanciát és a levelek klorofilltartalmát (SPAD-értékben). A kísérlet végén TTC-teszttel vizsgáltuk a gyökerek életképességét, mikroszkópos vizsgálattal becsültük az AM gomba gyökérkolonizációját, valamint meghatároztuk a gyökér- és hajtástömeget.
A vízhiány szignifikánsan (9–35%-kal) csökkentette a búzafajták gyökértömegét, mely a mért CR-értékekben is tükröződött. A szárazság okozta CRés biomassza-csökkenés jelentősebb volt a TC 33, mint az Mv. Hombár esetében. A CR monitorozásával kimutattuk a növények stressz utáni regenerációját és a fajták eltérő gyökérnövekedési dinamikáját. Az AMF oltás csökkentette a CR-t és a biomassza-produkciót (29–42%-kal), vélhetően az intenzív (84–87%-os) gyökérkolonizáció és a növénynevelés körülményei (erős szárazságstressz) következtében. Az oltás optimális öntözés mellett növelte a sztómakonduktanciát és a gyökér vitalitását. A vízhiány azonban csökkentette a gyökér életképességét. A klorofilltartalom leginkább a búzafajták között mutatott eltérést az Mv. Hombár nagyobb SPAD-értékével. A gyökértömeg és -kapacitás között szoros lineáris korrelációt (R2 = 0,792–0,865) találtunk. A TC 33 fajta regressziós egyenesének nagyobb meredeksége a nagyobb hajtástömegből eredő nagyobb fajlagos vízfelvételre vezethető vissza.
Eredményeink alapján a CR-mérés alkalmas a gyökérnövekedési dinamika monitorozására és a környezeti hatások detektálására. A roncsolásmentes eljárás egyéb növénymorfológiai és -élettani vizsgálómódszerek hasznos kiegészítője lehet.
Authors:Włodzimierz Opoka, Katarzyna Kała, Remigiusz Krężałek, Katarzyna Sułkowska-Ziaja, Anna Maślanka and Bożena Muszyńska
Agaricus bisporus and Imleria in vitro cultures were cultivated on modified Oddoux medium, and Oddoux medium was enriched with serine or anthranilic acid. Serine or anthranilic acid was used at the concentrations of 0.1, 0.25, 0.5, and 0.75 g/L of medium. Determination of indole compounds in the obtained biomass was carried out using thin-layer chromatography (TLC) with densitometric detection. In every analyzed sample, presence of serine or anthranilic acid was studied. Comparison of the results obtained for the treatment and control samples allowed us to determine the optimum concentration of serine or anthranilic acid in the medium in order to obtain biomass with increased content of indole compounds. A. bisporus with addition of anthranilic acid or serine to the medium at the concentration of 0.5 g/L was the most beneficial. In the case of Imleria badia, anthranilic acid at the concentration of 0.5 g/L was the most optimal. This is the first report demonstrating the content of indole derivatives in biomass affected by their precursors (serine or anthranilic acid). The study indicates that modification of the medium can provide satisfactory results, and it is worth to search for its new, improved compositions.
Authors:Chen Cheng, Nie Cun-Xi, Liang Jing, Wang Yong-Qiang, Liu Yan-Feng, Ge Wen-Xia and Zhang Wen-Ju
A validated high-performance liquid chromatography (HPLC) method has been developed to analyze the (±)-gossypol in the selection of strains of Candida tropicalis culture. Since gossypol was easily degraded and oxidized, the addition of antioxidant NADPH-Na4 and acetone extraction was chosen to prevent gossypol degradation and gradient elution assay was applied to obtain gossypol resolution. Concentrations of gossypol in C. tropicalis ZD-3 culture 20 μg/mL were determined, and concentration–time profiles were observed. Linearity of the gossypol standard curve by HPLC area method was ranged from 0.1 to 20 μg/mL with Y = 26.954 × X − 29.547, R2 = 0.9991, and n = 3, with limit of detection (LOD) of 50 ng/mL and lower limit of quantification (LLOQ) of 500 ng/mL. The recovery rate is dose-dependent and ranged from 85.3% to 103.5%. It is a rapid and reliable HPLC method for gossypol quantization in microorganism culture which could be applied in solid fermentation in the feed industry.
Authors:V. T. Gawande, K. G. Bothara and C. O. Satija
Stability-indicating High-Performance Thin-Layer Chromatography (HPTLC) method for simultaneous estimation of cefixime trihydrate and azithromycin dihydrate was developed. Both the drugs were subjected to different stress conditions recommended by International Conference on Harmonization (ICH) guideline Q1A (R2). Forced degradation was carried out for hydrolytic, oxidative, photolytic, and thermal degradation conditions. Cefixime was susceptible for degradation under all stress conditions showing four degradation products (CI–IV). However, azithromycin formed only one degradation product (AI) under acid hydrolysis. Aluminum plates precoated with silica gel 60F254 were used as the stationary phase while mixture of ethyl acetate–methanol–acetone–toluene–ammonia (1:5:7:0.5:0.5, v/v) was used as mobile phase. Detection wavelength used was 235 nm for CEFI and CI–IV. AZI and AI were detected by post development derivatization, spraying with sulfuric acid–ethanol (1:4, v/v) followed by heating at 100 °C for 5 min. Degradation products were isolated by preparative HPTLC and characterized by MS/MS. The developed method was validated for linearity, precision, accuracy, specificity, and robustness and has been successfully applied in the analysis of these drugs in tablet dosage form.