High-pressure Thin-layer Chromatography Research Articles

Fungal Elicitation Enhances Vincristine and Vinblastine Yield in the Embryogenic Tissues of Catharanthus roseus.

Fungal elicitation could improve the secondary metabolite contents of in vitro cultures. Herein, we report the effect of Fusarium oxysporum on vinblastine and vincristine alkaloid yields in Catharanthus roseus embryos. The study revealed increased yields of vinblastine and vincristine in Catharanthus tissues. Different concentrations, i.e., 0.05% (T1), 0.15% (T2), 0.25% (T3), and 0.35% (T4), of an F. oxysporum extract were applied to a solid MS medium in addition to a control (T0). Embryogenic calli were formed from the hypocotyl explants of germinating seedlings, and the tissues were exposed to Fusarium extract elicitation. The administration of the F. oxysporum extract improved the growth of the callus biomass, which later differentiated into embryos, and the maximum induction of somatic embryos was noted T2 concentration (102.69/callus mass). A biochemical analysis revealed extra accumulations of sugar, protein, and proline in the fungus-elicitated cultivating tissues. The somatic embryos germinated into plantlets on full-strength MS medium supplemented with 2.24 µM of BA. The germination rate of the embryos and the shoot and root lengths of the embryos were high at low doses of the Fusarium treatment. The yields of vinblastine and vincristine were measured in different treated tissues via high-pressure thin-layer chromatography (HPTLC). The yield of vinblastine was high in mature (45-day old) embryos (1.229 µg g-1 dry weight), which were further enriched (1.267 µg g-1 dry weight) via the F. oxysporum-elicitated treatment, especially at the T2 concentration. Compared to vinblastine, the vincristine content was low, with a maximum of 0.307 µg g-1 dry weight following the addition of the F. oxysporum treatment. The highest and increased yields of vinblastine and vincristine, 7.88 and 15.50%, were noted in F. oxysporum-amended tissues. The maturated and germinating somatic embryos had high levels of SOD activity, and upon the addition of the fungal extracts, the enzyme's activity was further elevated, indicating that the tissues experienced cellular stress which yielded increased levels of vinblastine and vincristine following the T2/T1 treatments. The improvement in the yields of these alkaloids could augment cancer healthcare treatments, making them easy, accessible, and inexpensive.

Cytotoxic Activities and Fingerprint Analysis of Triterpenes by HPTLC Technique for Distinguishing Ganoderma Species from Vietnam and other Asian Countries

Ganoderma lucidum (Fr.) P. Karst. (Ganodermataceae), commonly called Linhzhi, is traditionally employed in the treatment of human diseases, including hepatitis, liver disorders, hypercholesterolemia, arthritis, bronchitis, and tumorigenic diseases. In this study, the fingerprint profiles of five different strains of G. lucidum originated from Japan, Korea, China, and Vietnam, five samples of G. lucidum growing on Erythrophloeum fordii Oliv. in Vietnam, and five related Linhzhi species (Ganoderma applanatum, Ganoderma australe, Ganoderma clossum, Ganoderma subresinosu, and Ganoderma sp.) were investigated for triterpene derivatives using high-pressure, thin-layer chromatography (HPTLC). The HPTLC fingerprint profiles demonstrated significant differences between G. lucidum and other related Linhzhi species in the presence of triterpene derivatives. Evaluation for the cytotoxicity of these samples against four cancer cell lines, including A549, MCF7, PC3, and HepG2, displayed various levels of cytotoxic effects, with IC50 values of: 15.6-46.3 µg/mL on the A549 cancer cell line, of 18.4-43.6 µg/mL on the MCF7 cancer cell line, of 10.0-32.1 µg/mL on the PC3 cancer cell line, and of 10.6-27.6 µg/mL on the HepG2 cancer cell line. Conclusively, these data contributed to the literature on the cytotoxic activities and fingerprint analysis of triterpenes by the HPTLC technique for distinguishing Ganoderma species from Vietnam and other Asian countries.

Review on Forced Flow Thin Layer Chromatography: An updated Analytical Technique

Chromatography is a method for the isolation of a mixture into separate sections. It is usually based on the principle of partition of solute between two phases. It usually consists of a mobile phase and a stationary phase. The advancement of the mobile process in the planar stationary stage is evoked by different external or internal forces. Capillarity and gravity are the intrinsic force whereas external forces are electric field, pump, and centrifugal forces. Thin layer chromatography and high-pressure thin layer chromatography are based on the adsorption theory. Forced flow thin layer chromatography is also based on the adsorption principle. FFTLC is an authoritative method of separation that uses a solvent pump to extract the sample mixture into a pressurized ultra-micro cavity containing an adsorbent analytical or preparative planer bed. FFTLC needs no or slight sample preparation as in TLC. One relation between FFTLC and high-pressure liquid chromatography is that as in HPLC, the maximum layer length is used for analytical and preparatory separation. FFTLC is a process in which continuous production and continual evaporation of the mobile phase from the end of the chromatography plate produces forced-flow. Here, we focus on FFTLC, an advanced and more sophisticated chromatographic technique.

Gamma ray–induced tissue responses and improved secondary metabolites accumulation in Catharanthus roseus

The present study investigated the impact of gamma ray irradiation on callus biomass growth and the yield of vincristine and vinblastine of in vitro grown tissues of Catharanthus roseus. The biochemical alteration underlying the synthesis of secondary metabolites has also been studied and a comparison of yield was prepared. The embryogenic tissues were exposed to 20, 40, 60, 80, and 100Gy gamma ray doses and the callus biomass fresh weight, the embryogenesis (the embryo numbers, germination, plant regeneration), the alteration of protein, proline, and sugar attributes at different morphogenetic stages were monitored. The callus biomass growth was maximum (1.65g) in 20Gy exposed tissues and was less in 100Gy treatment (0.33g). The gamma-irradiated embryogenic tissues differentiated into embryos but the embryogenesis % and somatic embryo number per culture reduced with increasing doses. It was least in 80Gy where very low numbers of embryos were formed (3.45 and 3.30 mean torpedo and cotyledonary embryo numbers per callus mass, respectively) which later germinated into plantlets. Protein, proline, sugar, and different antioxidant enzymes, i.e., superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT) activities, were investigated as the tissues were exposed to gamma ray elicitation/signaling, evoking cellular stress. Increased 80Gy gamma dose inhibited a 42.73% decrease in protein accumulation at initiation stages of embryogenic tissue. Soluble sugar level also declined gradually being least in 80Gy treated tissues (14.51mg gm-1 FW) compared to control (20.2mg gm-1 FW). Proline content, however, increased with increasing gamma doses, maximum at 80Gy (8.28mg gm-1 FW). The SOD, APX, and CAT activity increased linearly with enhanced level of gamma doses and maximum, i.e., 3.91 EU min-1mg-1, 1.71 EU min-1mg-1, and 4.89 EU min-1mg-1, protein activity was noted for SOD, APX, and CAT, respectively, at 80Gy gamma rays treated tissues. The quantification of vinblastine and vincristine in gamma ray elicitated tissues was made by using high-pressure thin layer chromatography (HPTLC). Somatic embryo-regenerated plant's leaves had the maximum yield of vinblastine (15.13 µgm gm-1 DW) at 40Gy irradiation dose compared to control (13.30 µgm gm-1 DW)-the increased yield % is 13.75. The stem is also rich source producing 11.98 µgm gm-1 DW of vinblastine. Among the various developmental stages of embryos, vinblastine content was highest in germinating stage of embryos (10.14 µgm gm-1 DW) compared to other three, i.e., initiation, proliferation, and maturation embryo stages. Similarly, highest accumulation of vincristine (6.32µg gm-1 DW) was noted at low gamma irradiation dose (20Gy) in leaf tissues. The present study indicates that the synthesis of vinblastine and vincristine was growth- and development-specific and the lower 20-40Gy gamma levels were more effective in enriching alkaloids while higher doses declined yield. KEY POINTS: • Vinblastine and vincristine yield was quantified in in vitro grown tissues and leaves of embryo regenerated Catharanthus roseus after gamma ray treatment. • The accumulation of vinblastine and vincristine was maximum in regenerated leaves; low doses were more efficient in improving yield. • Gamma ray irradiation impacted biochemical profiles, caused cellular stress, and perhaps responsible for improved alkaloid yield.

Chemical profiling, in vitro antioxidant and antidiabetic assessment of flavonoids from the ethanol extract of Hermannia geniculata root

Determination of the in vitro antioxidant and the inhibitory potential of flavonoids from Hermannia geniculata (FHG) roots on diabetes-linked enzymes was carried out. The chemical profiling of FHG roots extract was investigated using High Pressure Thin Layer Chromatography (HPTLC) fingerprint analysis. The reactive oxygen scavenging potential of the extract was analyzed. Starch solution (1%) was reacted with different concentrations of FHG extract to determine the α-amylase inhibitory potential of the extract while α- glucosidase inhibition assay was carried out through incubation of different concentrations of the extract followed by addition of p-ntrophenyl-α-Dglucopyranoside solution. HPTLC results indicated the presence of flavonoids/ phenolcarboxylic acid, and Kaemferol (Rf 0.80) were detected in the extract with retention factor Rf. ranging from 0.08 to 0.95. FHG extract showed commendable antioxidant properties with IC50 values (3.07± 0.12, 2.13± 0.67) µg/mL for 1, 1-diphenyl-2- picrylhydrazyl (DPPH) and 2, 2-azino-bis (3- ethylbenzothiazoline-6-sulphonic) acid (ABTS) radicals which were lower and significantly different (p<0.05) compared to standard silymarin with IC50: (3.55± 0.10, 2.77± 0.75) µg/mL for DPPH and ABTS respectively. The results indicated mild inhibition of α-amylase with IC50: (5.55± 0.37) µg/mL which was higher and significantly different (p<0.05) from acarbose with IC50: (3.81± 0.29) µg/mL. Moreover, the extract showed 73% inhibition of α-glucosidase. Kinetic studies of FHG extract revealed competitive and mixed non-competitive inhibition of α- amylase and α-glucosidase respectively. This study indicated FHG capabilities of scavenging reactive oxygen species and reducing hydrolysis of starch responsible for post-prandial hyperglyceamia seen in type 2 diabetes mellitus.
 Keywords: Antidiabetic, Antioxidant, Flavonoids, Hermannia geniculate, HPTLC

Nutritional Assessment of Indonesian Chilli Landraces (Capsicum chinense Jaqc.)

Chillies (Capsicum spp.) are highly nutritious vegetables and spices and are an important component in the Asian cuisine due to their aroma attributes and health value. To gain a better insight of the health value of chillies in North Toraja, South Sulawesi, Indonesia, the nutrient content, the morphological parameters and the yield of three different chilli landraces of the variety ‘Katokkon’ (Capsicum chinense Jaqc.), i.e. ‘Limbong Sampolo’, ‘Leatung 1’, and ‘Leatung 2’ were investigated. The nutrient parameters included capsaicin, pro-vitamin A, and vitamin C contents. Moreover, those differences were studied in two villages (Buntu Datu and To’pao) for the landrace ‘Leatung 2’. The research was performed with dried chillies using high pressure thin layer chromatography for the determination of capsaicin content, spectrophotometric determination for pro-vitamin A and iodometry for vitamin C content. The results showed that the capsaicin content varied between 7.91 mg/g DM (‘Leatung 1’) to 9.94 mg/g DM (‘Limbong Sampolo’) which corresponds with values on the Scoville heat unit (SHU) ranging from 118,712.677 to 149,074.747. The pro-vitamin A content was found to be highest (1.01 mg/g DM) in ‘Limbong Sampolo’ and lowest (0.60 mg/g DM) in ‘Leatung 2’ from To’pao. Vitamin C was found to be lowest (0.20 mg/g DM) in ‘Leatung 2’ from Buntu Datu and highest (0.26 mg/g DM) in ‘Leatung 1’. The statistical analysis showed that there were no significant differences between the landraces and the villages for ‘Leatung 2’. Compared to the capsaicin values in literature (up to SHU 600,000), the landraces showed lower values. This was also found for the pro-vitamin A contents. In contrast, the vitamin C values were higher than those reported in literature. In conclusion, it can be stated, that the three landraces of ‘Katokkon’ in general showed high nutritious values, irrespectively of the different growing conditions in the two villages.

HPTLC fingerprinting analysis and in vitro pharmacological activities of Hermannia geniculata roots phenols

Abstract Hermannia geniculata is a herb that plays an important role in the treatment of an array of diseases including diabetes, ulcer, and colitis in the South African traditional medicine. The bioactive constituent and medicinal properties in phenols of Hermannia geniculata (PoHG) roots were investigated using high pressure thin layer chromatography (HPTLC). The α-amylase inhibitory potentials of PoHG was determined by reacting different concentration of the plant extract with 1% starch solution containing α-amylase. The inhibitory effect of the extract on α-glucosidase was evaluated by pre-incubating α-glucosidase with varying extract concentrations followed by the addition of ρ -nitrophenylglucopyranoside.. The reactive oxygen and free radical scavenging potentials of the extract were also analyzed. The result showed the presence of phenolic compounds in the extract with retention factor (Rf) values ranging from 0.14 to 095. The extract scavenged DPPH, ABTS+, hydroxyl, and superoxide anion radicals. The extract was able to chelate metallic ions with a lower IC50 value which differs significantly (p≤0.05) from silymarin. Moreover, PoHG extract inhibited the key enzymes (α-glucosidase and α-amylase) involved in carbohydrate catabolism with IC50 values of 1.76 ±0.14 and 7.52 ±0.23 mg/mL respectively while IC50 value reported for acarbose were 7.62 ±0.12 and 4.38 ±0.25 mg/mL for glucosidase and α-amylase, respectively. The α-glucosidase exhibited non-competitive inhibition by PoHG extract while α-amylase showed uncompetitive inhibition. This study confirmed the presence of phenol in PoHG extract and also showed an appreciable antioxidant and antidiabetic activities in vitro. Therefore, PoHG extract may be of nutraceutical importance.

PHYTOCHEMICAL SCREENING OF ETHANOLIC EXTRACT OF WHOLE PLANT OF SIDA GLUTINOSA

Objective: The aim of the present study is to screen for the phytochemical constituents which are of pharmacological importance present in the ethanolic extract of whole plant of Sida glutinosa (SG). 
 Methods: The ethanolic extract of the dried whole plant of SG is subjected to preliminary phytochemical screening which showed the presence of major phytoconstituents such as phenols, flavonoids, and alkaloids. The extract was screened for its antioxidant activity by 2,2-diphenyl-1- picrylhydrazyl, hydroxyl radical, Iron (III) to Iron (II) reducing activity, and nitric oxide scavenging assay. Further, the ethanolic extract was subjected to fingerprinting technique high-pressure thin-layer chromatography (HPTLC). Reverse phase high-pressure liquid chromatography (Rp-HPLC) was performed to estimate the amount of total phenolics, flavonoids, and alkaloids quantitatively in isocratic mode.
 Results: Phytochemical screening of the ethanolic extract of the plant showed the presence of pharmacologically important constituents such as alkaloids, flavonoids, phenolics, and terpenoids. The study also revealed the potential antioxidant activity of the extract with IC50 value. The extract fingerprinting through HPTLC revealed the presence of various phytoconstituents. Rp-HPLC showed 0.35±0.12 μg/ml of total phenolics, 0.0013±0.05μg/ml of alkaloids, and 0.00081±0.08 μg/ml of flavonoids.
 Conclusion: Scientific evaluation of SG which has therapeutic significance was carried out which is an important concept for the standardization of the plant-based drug. Further, there is a need for isolation and characterization of the lead molecules their systemic evaluation for its pharmacological activities.

MARKET SAMPLE SURVEY OF CROCUS SATIVUS LINN. TO ASSESS THE GENUINITY FOR USING ANTIOXIDANT ACTIVITY, TOTAL PHENOLIC CONTENT AND HIGH-PRESSURE THIN LAYER CHROMATOGRAPHY USING DETECTION OF FLAVONOIDS

Objective: Herbalism is a traditional medicine or folk medicine practice based on the use of plants and plant extracts. Many of the drugs used in conventional medicine are dried from herbs. Despite the fluctuation in prices in international markets, saffron was still remained the most expensive spice. The main aim of this study is to examine the antioxidant activity, total phenolic content, high-pressure thin layer chromatography using flavanoid analysis and adulteration detection of saffron. Crocus sativus. Linn is a perennial stemless herb of the Iridaceae family. Saffron stigmas of sample1, sample2, sample3and sample4 are collected from different rates of the market sample from Thrissur district, sample5 collected from the Oushadhi premises, and it is collected from Himachal Pradesh.
 Methods: In this study detecting the antioxidant activity, total phenolic content, high-pressure thin layer chromatography using flavanoid analysis of different samples of saffron stigmas. The extracts were prepared by using ethanol as a solvent.
 Results: Safranal is present only in s5 sample. It is the main essential volatile oil responsible for the saffron characteristic such as odour. Phenolic content is varied in different market samples. The amount of phenolic compounds in the saffron extract was determined using the Folin-ciocalteau reagent. Total phenolic content is the help to detect the pure and fake saffron. The phenolic content is higher in S5. Sample S5 showed 0.737 mg/ml phenolic content. Lowest level of phenolic content in sample S3. Sample S3 showed 0.0887 mg/ml phenolic content. Sample S4 showed 0.564 mg/ml total phenolic content. Sample S1 showed 0.416 mg/ml total phenolic content and sample S2 showed 0.267 mg/ml phenolic content. Antioxidant activity is higher in sample s5. and it is different in different market samples. Sample 5 stigma posses higher antioxidant activity. Sample S5 showing 14.88% antioxidant activity in 100 mg/ml concentration, 7.26% in 80 mg/ml concentration, 2.23% in 60 mg/ml concentration, 2.21% in 40 mg/ml and 1.01% in 20 mg/ml concentration. Sample S3 showed the lower antioxidant activity in 0.1% in 60 mg/ml concentration and 0.1% in 80 mg/ml. Ascorbic acid standard showing 14.89% in 100 mg/ml concentration, 7.26% in 80 mg/ml concentration, 4.56% in 60 mg/ml concentration, and 3.1% in 40 mg/ml concentration, and 1% in 20 mg/ml concentration. Flavonoid content is different in different samples. It is present highly present in sample s1 and s5. sample s3 do not contain the Flavanoid. The quality of the samples depend on the price values.
 Conclusion: The authenticity of saffron is an extremely important matter for the industry and for the consumers in view of security and protection,quality assurance, active properties and last but not least, economic impact. Despite the fluctuation in prices in international markets, saffron was and still remains the most expensive spice. The genuine saffron samples possess higher price value. The fake saffron available in the market with lower price value. The quality of the saffron depends upon the price values. These observations would be of immense value in the botanical identification and standardization of the drug in crude form and would help to distinguish the drug from its other spices.

MARKET SAMPLE SURVEY OF CROCUS SATIVUS LINN. TO ASSESS THE GENUINITY FOR USING ANTIOXIDANT ACTIVITY, TOTAL PHENOLIC CONTENT AND HIGH-PRESSURE THIN LAYER CHROMATOGRAPHY USING DETECTION OF FLAVONOIDS

Objective: Herbalism is a traditional medicine or folk medicine practice based on the use of plants and plant extracts. Many of the drugs used in conventional medicine are dried from herbs. Despite the fluctuation in prices in international markets, saffron was still remained the most expensive spice. The main aim of this study is to examine the antioxidant activity, total phenolic content, high-pressure thin layer chromatography using flavanoid analysis and adulteration detection of saffron. Crocus sativus. Linn is a perennial stemless herb of the Iridaceae family. Saffron stigmas of sample1, sample2, sample3and sample4 are collected from different rates of the market sample from Thrissur district, sample5 collected from the Oushadhi premises, and it is collected from Himachal Pradesh.
 Methods: In this study detecting the antioxidant activity, total phenolic content, high-pressure thin layer chromatography using flavanoid analysis of different samples of saffron stigmas. The extracts were prepared by using ethanol as a solvent.
 Results: Safranal is present only in s5 sample. It is the main essential volatile oil responsible for the saffron characteristic such as odour. Phenolic content is varied in different market samples. The amount of phenolic compounds in the saffron extract was determined using the Folin-ciocalteau reagent. Total phenolic content is the help to detect the pure and fake saffron. The phenolic content is higher in S5. Sample S5 showed 0.737 mg/ml phenolic content. Lowest level of phenolic content in sample S3. Sample S3 showed 0.0887 mg/ml phenolic content. Sample S4 showed 0.564 mg/ml total phenolic content. Sample S1 showed 0.416 mg/ml total phenolic content and sample S2 showed 0.267 mg/ml phenolic content. Antioxidant activity is higher in sample s5. and it is different in different market samples. Sample 5 stigma posses higher antioxidant activity. Sample S5 showing 14.88% antioxidant activity in 100 mg/ml concentration, 7.26% in 80 mg/ml concentration, 2.23% in 60 mg/ml concentration, 2.21% in 40 mg/ml and 1.01% in 20 mg/ml concentration. Sample S3 showed the lower antioxidant activity in 0.1% in 60 mg/ml concentration and 0.1% in 80 mg/ml. Ascorbic acid standard showing 14.89% in 100 mg/ml concentration, 7.26% in 80 mg/ml concentration, 4.56% in 60 mg/ml concentration, and 3.1% in 40 mg/ml concentration, and 1% in 20 mg/ml concentration. Flavonoid content is different in different samples. It is present highly present in sample s1 and s5. sample s3 do not contain the Flavanoid. The quality of the samples depend on the price values.
 Conclusion: The authenticity of saffron is an extremely important matter for the industry and for the consumers in view of security and protection,quality assurance, active properties and last but not least, economic impact. Despite the fluctuation in prices in international markets, saffron was and still remains the most expensive spice. The genuine saffron samples possess higher price value. The fake saffron available in the market with lower price value. The quality of the saffron depends upon the price values. These observations would be of immense value in the botanical identification and standardization of the drug in crude form and would help to distinguish the drug from its other spices.

Inhibitory effects of catechin isolated from Acacia catechu on ovalbumin induced allergic asthma model

Purpose Polyphenols possess anti-allergic activities. Catechin is one of the polyphenols that are abundantly present in the Acacia catechu. In this study, the authors investigated the effect of catechin isolated from A. catechu in an experimental mouse model of ovalbumin (OVA)-induced allergic asthma. Design/methodology/approach Catechin was isolated from A. catechu, and phytochemical analysis was carried out by ultraviolet visible and thin-layer chromatography (TLC), high pressure thin-layer chromatography was used for the determination of an amount of catechin present. In a first set of an experiment, the authors have carried out dose-dependent evaluation of catechin on histamine synthesis in normal rats. In another study, allergic asthma was induced in BALB/c mice by intraperitoneal injection of 50 mg OVA dissolved in 4 mg aluminum hydroxide dissolved in 0.2 ml saline on Days 0 and 14. Catechin was given orally at the dose of 100 mg/kg, once a day from Day 1 to Day 35 and after which various respiratory parameters such as tidal volume, respiratory rate and airflow rate, biochemical parameters such as histamine release from mast cells, bronchoalveolar (BAL) lavage fluid analysis and histopathology of lungs were carried out. Findings Catechin showed significant (p < 0.05) improvement in respiratory parameters such as tidal volume, respiratory rate and airflow rate, as well as biochemical and hematological parameters such as blood histamine, serum bicarbonate and nitric oxide levels as compared to the disease control group. The treatment also showed inhibitory effects on histamine synthesis in rat peritoneal as well as BAL mast cells. Also, a significant (p < 0.05) improvement in lung histopathology was observed with catechin. Originality/value From the present study, the authors can conclude that catechin exhibited potent anti-allergic activity by inhibition of histamine synthesis by inhibition of histidine decarboxylase enzyme. The study suggests that catechin has therapeutic potential for the treatment of allergic inflammatory disease in humans.

Determination of Dipeptidyl Peptidase-4 Inhibitors by Spectrophotometric and Chromatographic Methods

Dipeptidyl peptidase-4 (DPP-4) inhibitors are a novel class of oral anti-diabetic agents used in the treatment of type 2 diabetes mellitus and work to enhance the effect of incretin hormones. There has been significant research on wide range of analytical techniques that could be useful in the estimation of DPP-4 inhibitors in formulations and biological matrices. Analytical methods such as ultraviolet (UV) spectrophotometry, mass spectroscopy, NMR spectroscopy, HPLC, high pressure thin layer chromatography, ultra performance liquid chromatography, liquid chromatography‒mass spectrophotometry and capillary zone electrophoresis have been reported for the estimation of DPP-4 inhibitors in single and/or in combination with other drugs. This comprehensive review covers most of the spectrophotometric and chromatographic methods described for the determination of sitagliptin, alogliptin, vildagliptin, linagliptin, saxagliptin and teneligliptin in pure forms, in different pharmaceutical dosage forms and biological fluids. From the review it can be deduced that a large number of chromatographic methods have been developed, and HPLC-UV methods have been commonly used in the detection and evaluation of DPP-4 inhibitors.

Model Development to Enhance the Solvent Extraction of Polyphenols from Mango Seed Kernel

The aim of this study was to improve the extraction yield of polyphenols from mango seed kernel by solvent extraction. Seven organic solvents were tested to identify the best extraction efficiency, and a ternary solvent mixture of Methanol-Acetone-Water (54: 23:23), developed using the mixture design methodology, allowed to extract the maximum amount of polyphenols with maximum antioxidant activity. The solvent mixture developed was then applied to polyphenols extraction from kernels of 11 mango varieties from the savannah region of Cameroon, and High Pressure Thin Layer Chromatography (HPTLC) used to identify the phenolic compounds present in the extracts. The total polyphenols contents (TPC), flavonoids and Total Reducing Power (TRP) of mango kernels were respectively in the range of 93.38 - 121.01 mg Gallic Acid Equivalent/ g DW, 12.64 - 16.26 mg Catechin Equivalent/g DW and 95.42 - 113.97 mg Ascorbic Acid Equivalent/g DW. Chlorogenic acid, Mangiferin, Gallic acid and Methyl gallate are highly suspected as component molecules of polyphenols from mango seed kernel.

Azilsartan: A Review of Analytical Methods for estimation in Pharmaceutical Formulation

Azilsartan medoxomil was approved by the United States Food and Drug Administration in 2011. AZL is used for the treatment of essential hypertension and has publicized promising results in blood pressure (BP) reduction and tolerability, but has not yet been considered into practice as compared to other angiotensin II receptor blockers (ARBs). AZL lowers blood pressure by blocking the action of angiotensin II at the AT1 receptor, a hormone that contracts blood vessels and reduces water excretion through the kidneys. There have been numerous methods developed so far for quantitative estimation of AZL in bulk and pharmaceutical dosage form. Pharmaceutical analytical methods include UV Visible Spectrophotometry, High Pressure Thin Layer Chromatography, Reversed Phase High Performance Liquid Chromatography and hyphenated techniques like Liquid chromatography-Mass Spectrometry. This review paper includes publications form 2010 to 2018. The chromatographic methods primarily RP-HPLC and LC-MS found more accurate, precise and sensitive compared to other pharmaceutical methods. AZL shows greater extent of degradation up to 40% with hydrogen peroxide. AZL is degraded up to 38% and 31% with acidic and neutral pH respectively while it shows greater photo stability and dry heat.

Estimation of arecoline content of various forms of areca nut preparations by high-pressure thin-layer chromatography

Estimation of arecoline content of various forms of areca nut preparations by high-pressure thin-layer chromatography

Biodegradation and detoxification of textile dye Disperse Red 54 by Brevibacillus laterosporus and determination of its metabolic fate

Bioremediation is one of the milestones achieved by the biotechnological innovations. It is generating superior results in waste management such as removal of textile dyes, which are considered xenobiotic compounds and recalcitrant to biodegradation. In the present bioremedial approach, Brevibacillus laterosporus was used as an effective microbial tool to decolorize disperse dye Disperse Red 54 (DR54). Under optimized conditions (pH 7, 40°C), B. laterosporus led to 100% decolorization of DR54 (at 50 mg L(-1)) within 48 h. Yeast extract and peptone, supplemented in medium enhanced the decolorization efficiency of the bacterium. During the decolorization process, activities of enzymes responsible for decolorization, such as tyrosinase, veratryl alcohol oxidase and NADH--DCIP reductase were induced by 1.32-, 1.51- and 4.37-fold, respectively. The completely different chromatographic/spectroscopic spectrum of metabolites obtained after decolorization confirmed the biodegradation of DR54 as showed by High pressure liquid chromatography, High pressure thin layer chromatography and Fourier transform infrared spectroscopy. Gas chromatography-Mass spectroscopy studies suggested the parent dye was biodegraded into simple final product, N-(1λ(3)-chlorinin-2-yl)acetamide. Phytotoxicity study suggested that the metabolites obtained after biodegradation of DR54 were non-toxic as compared to the untreated dye signifying the detoxification of the DR54 by B. laterosporus.

Degradation of monocrotophos in soil, microbial versus enzymatic method

Aim: The present study has been designed to compare two commonly used methods viz. microbial and enzymatic for the degradation of monocrotophos (MCP) in sandy loam soil of Rajasthan. Methods: Methodology involves the use of molecularly characterized fungal strain Aspergillus niger JQ660373. For the microbial method spore suspension (1 × 108 spores/ml) was used as inoculum. Enzymatic method employ the use isolated, purified extracellular fungal hydrolases as inoculum. %age of MCP degradation was assessed in the form of residual MCP concentration and hence degradation kinetics was calculated. The results were confirmed by using analytical tools, high pressure thin layer chromatography (HPTLC) and Fourier transform infrared spectroscopy (FTIR). Results: The results of this study demonstrated that extracellular fungal hydrolase composed of two different subunits of 33 and 67 Kd. Both subunits synergistically degrade MCP. Residual MCP concentration was found to be 64.94 ± 0.42 and 16.95 ± 0.55 µg ml−1 after 15 days of incubation for microbial and enzymatic method, respectively. Degradation of MCP followed first order kinetics with rate constants of 0.002 and 0.136 day−1 and hence the calculated half-life was found to be 12.64 and 5.14 days, respectively. HPTLC chromatograms clearly indicate the significant decrease in the standard MCP peak (rf 0.19-0.21) with increasing incubation duration. Molecular insight of MCP degradation was studied by FTIR. Degradation proceeded with hydrolytic cleavage of MCP resulting in the formation of inorganic phosphates (−PO4). Conclusion: The study concludes that enzymatic method of degradation was more efficient than the microbial method.

Experimental and DFT studies on the antioxidant activity of a C-glycoside from Rhynchosia capitata

Rhynchosia capitata (=Glycine capitata) Heyne ex roth, was found to possess polyphenolics including flavonoids, which acts as potential antioxidant. The study of ethanolic extract of roots and leaves reveals that the leaves possess high polyphenolics including flavonoids than roots. This was also confirmed by DPPH radical scavenging activity. Leaf powder of the plant was extracted with different solvents by soxhlet apparatus in the order of increasing polarity. The DPPH scavenging activity of methanol fraction was found to be high compared to the crude extract and other fractions. Nitric oxide scavenging activity was dominant in chloroform fraction compared to methanol fraction. Presence of flavonoids especially vitexin, a C-glycoside in methanol and chloroform fractions were confirmed by high pressure thin layer chromatography (HPTLC) analysis. The structural and molecular characteristics of naturally occurring flavonoid, vitexin was investigated in gas phase using density functional theory (DFT) approach with B3LYP/6-311G(d,p) level of theory. Analysis of bond dissociation enthalpy (BDE) reveals that the OH site that requires minimum energy for dissociation is 4′-OH from B-ring. To explore the radical scavenging activity of vitexin, the adiabatic ionization potential, electron affinity, hardness, softness, electronegativity and electrophilic index properties were computed and interpreted. The nonvalidity of Koopman’s theorem has been verified by the computation of Eo and Ev energy magnitudes. Interestingly, from BDE calculations it was observed that BDE for 4′-OH, 5-OH and 7-OH are comparatively low for vitexin than its aglycone apigenin and this may be due to the presence of C-8 glucoside in vitexin. To substantiate this, plot of frontier molecular orbital and spin density distribution analysis for neutral and the corresponding radical species for the compound vitexin have been presented.

High Performance Thin Layer Chromatography (HPTLC) profile of an insecticidal compound of Trochus radiatus and assessment of mucus-associated bacterial extracts

The current study is based on the screening of novel insecticides from new sources that have not being exploited hitherto. The major objectives of this research work were to extract marine molluscs, Lambis lambis, Trochus radiatus and Chicoreus ramosus, from Tuticorin coastal waters using different solvents to test their insecticidal properties and partially purify the active components. The ethyl acetate extracts of L. lambis and T. radiatus showed 100% mortality of Sitophilus oryzae, at a concentration of 200 μg ml−1. The LC50 values for ethyl acetate extracts of L. lambis, T. radiatus and C. ramosus were found to be 67.08, 348.86 and 571.42 μg ml−1, respectively. With regard to bacterial metabolites, at a concentration of 200 μg ml−1 the ethyl acetate extract of A3 was able to elicit an activity of 40% and that of strain A1 – 20%. The LC50 values of the bacterial metabolites were also investigated. Upon chromatographic separation of active ethyl acetate extracts of T. radiatus, the 100% methanol column-purified fraction was found to have an activity of 30% at a concentration of 10 μg against S. oryzae. The purity of the partially pure active compound was observed to be 72.78% on analysis with high pressure thin layer chromatography.

An Improved Commercial Method for Determination of Nicotine in Tobacco

A high-pressure thin-layer chromatography (HPTLC) method for quantitative analysis of nicotine in Nicotiana sp. was developed using a methanol extract of leaves and stems and TLC plates (silica gel 60 GF254) with spot visualization under ultraviolet (UV) light. Scanning at 235 nm in the absorption-reflection mode produced linear calibration curves in the range of 2 to 25 μg, with a correlation coefficient of 0.991. The average recovery rate was 95% (CV % 1.35). From the present study, the lower limit of detection was 0.08 μg spot−1 for the nicotine. The validity of the method was confirmed by comparing the UV spectra of the tobacco plant samples with standards within the same Rf window.