Thin Layer Chromatography Chromatogram Research Articles

A novel thin layer chromatography-flame ionization detection method for saturated, asphaltenes, resins, and asphaltenes group-type composition analysis with reverse order of chromatogram development.

The methodologies of asphaltenes-containing petroleum materials: saturated, aromatics, resins, asphaltenes group-type composition analysis are performed with the use of column adsorption-desorption or thin layer chromatography (TLC)-flame ionization detection under normal phase conditions with silica gel as the adsorbent. In a three-step procedure, the TLC chromatogram is developed within a decreasing distance by the mobile phase with increasing elution strength (polarity). The n-alkane used in the first step does not dissolve asphaltenes, which leads to the occlusion effect and an underestimation of the percentage of saturated hydrocarbons. In this article, the reverse order of the subsequent elution steps was proposed: the solvent polarity is simultaneously reduced and the chromatogram development distance is increased in the order dichloromethane:methanol 95:5 v/v, 3cm; toluene, 6cm; and n-hexane, 10cm. It was also intentional to reduce the weight of the applied sample to 5μg for bitumen and 2μg for asphaltene purity testing. It should be the rule that in stepwise TLC chromatogram development, the first mobile phase is a good solvent for all testing components. The IP 469 procedure should be corrected.

Cytotoxicity, Phytochemical Screening and Genetic analysis of Ginger (Zingiber officinale Rosc.) Callus and Rhizome

• Ginger rhizome extracts displayed strong cytotoxic effect against HT29, HT116 and MCF-7 cancer cells lines with highest activity against HT29. • Ginger callus extracts recorded non-cytotoxic effect against all examined cancer cells with IC50 >100 µg/mL. • Ginger rhizome extracts contained more phenols and terpenes than the callus ones. • The principal components of ginger; 6-gingerol and 6-shogaol were not detected in callus extracts. • Incidence of genetic changes in DNA of ginger callus during in vitro culture in 2,4-D containing medium. • Most polymorphic bands (11 out of 12) were generated in callus during in vitro culture with total polymorphism 23.07% between callus and plant. Ginger ( Zingiber officinale Rosc.) is well known as distinctive spice with considered therapeutic values. In the current work we aimed to investigate cytotoxic effect, phytochemical screening and genetic analysis of ginger callus and their rhizome. Cytotoxicity of ginger rhizome and callus extracts was examined against some human cancer cell lines HT29, HT116 and MCF-7 by MTT method. Phytochemicals were screened by thin layer chromatography (TLC) and genetic analysis was performed by ISSR markers. Ginger rhizome principle components, 6-gingerol and 6-shogoal were used as standards to detect their presence in callus. Results showed that, rhizome extracts displayed a strong cytotoxicity towards all tested cancer cells with the highest activity (IC 50 20.4 ± 3.0 µg/mL; P< 0.05) against HT29. IC 50 of ginger callus extracts were >100 µg/mL in all examined cancer cell lines. TLC chromatograms of rhizome and callus extracts revealed the presence of phenols and terpenes, whereas 6-gingerol and 6-shogaol were only detected in rhizome extracts. Genetic analysis by ISSR markers revealed occurrence some genetic changes in DNA of callus compared to that of rhizome with total polymorphism 23.07%. In conclusion, ginger cytotoxicity to HT29, HT116 and MCF-7 cancer cells was mainly associated with the presence of 6-shogaol and 6-gingerol in extracts, which appeared as strong cytotoxic effect in rhizome compared to non-cytotoxic effect was evaluated in the callus extracts. Therefore, further studies on the callus elicitation and precursors feeding to induce biosynthesis of ginger bioactive molecules are recommended.

Volatile Bioactive Compounds from Lasiodiplodia pseudotheobromae IBRL OS-64, an Endophytic Fungus Residing in the Leaf of Ocimum sanctum

Endophytic fungi are known as potential novel compound producers with promising antimicrobial activities. Hence, the present study aimed to investigate the possible bioactive compounds present in the ethyl acetate extract of Lasiodiplodia pseudotheobromae IBRL OS-64. The ethyl acetate extract exhibited significant antibacterial activity against both Gram-positive and Gram-negative bacteria in disc diffusion assay. Thin-layer chromatography (TLC) was performed with chloroform, acetone and ethyl acetate (1:2:1, respectively) used as a solvent system and nine spots with diverse polarities were obtained. The TLC chromatogram with the active spot was localized with bioautography assay and the finding revealed that the dark spot with an Rf value of 0.5882 showed good antibacterial activity against all test bacteria. The fraction F5 exhibited promising antibacterial activity upon partial purification of dark spot via column chromatography and the GC-MS analysis of fraction F5 resulted in the detection of a major compound, 2-Benzenedicarboxylic acid, mono (2-ethylhexyl) ester with 90% matching factor. Thus, this compound may greatly contribute to the antibacterial activity of the fraction and has the potential to be developed as an antibiotic. The findings indirectly indicate that fungal endophytes from the medicinal plant could be a potential candidate for bioactive compounds with pharmaceutical properties.

Polarity Difference and the Presence of Phytoestrogen Compounds Affecting Estrogenic Activity of Peperomia pellucida Extracts

Peperomia pellucida (L.) Kunth has been studied as an anti-osteoporotic agent. However, there is no report about its estrogenic activity, which is important for its anti-osteoporotic activity. Thus, the aim of this research was to study the estrogenic potency of P. pellucida extracts. The estrogenic activity of P. pellucida extracts (n-hexane, ethyl acetate, ethanol, and water extracts) was studied using E-screen assay and confirmed with a molecular docking simulations. Further, the presence of phytoestrogen compounds was identified using thin layer chromatography (TLC), TLC densitometry, and high performance liquid chromatography. The n-hexane, ethyl acetate, and ethanol extracts at a concentration of 0.1 μg mL-1 exhibited a partial agonist effect, whereas the water extract showed full agonisteffect at the similar concentration. This activity was produced through a classical ligand-dependent mechanism similar to estradiol. N-hexane and ethyl acetate extracts showed antiestrogenic activity. The TLC chromatogram evidently depicted the presence of quercetin and stigmasterol in the n-hexane and ethyl acetate extracts. Apigenin and apigetrin at concentrations of 0.239±0.076 and 1.063±0.156 μg mg-1 extract, respectively, were present in the water extract. A docking study on estrogen receptors confirmed that apigetrin prefer to produce estrogenic activity, whereas the other compounds can produce both estrogenic and antiestrogenic activity. Hence, we suggest that the bioactive compounds in the water extract are flavonoids, such as apigenin and apigetrin. In summary, the water extract is recommended to be used as an estrogenic agent.

Qualitative and Quantitative Analysis of Linoleic Acid in Polygonati Rhizoma

ObjectiveTo explore the major compound in Polygonati Rhizoma (Huang Jing, 黄精) for quality control. MethodsThe major compound was isolated and analyzed by liquid chromatography-mass spectrometry (LC-MS), and subsequently further identified by nuclear magnetic resonance (NMR). Thin layer chromatography (TLC) was optimized based on the previous methods reported in the Chinese Pharmacopeia (2015 edition). ResultsThe major compound was isolated from the natural material and identified as linoleic acid. A high performance liquid chromatography (HPLC) method with robust linearity (R2 = 0.999 7), specificity, precision, stability, repeatability and recovery was developed for linoleic acid determination. TLC chromatogram was improved significantly after optimization for qualitative analysis. ConclusionsThe optimized TLC method is practical and can be adopted for quality control of Polygonati Rhizoma (Huang Jing, 黄 精 ). The levels of linoleic acid vary between species of Polygonati Rhizoma (Huang Jing, 黄 精 ), with Polygonatum cyrtonema Hua (Jiang Xing Huang Jing, 姜型黄精) showing the highest contents. This study provides valuable information for quality control of Polygonati Rhizoma (Huang Jing, 黄精).

TLC-Based Bioassay to Isolate Kairomones from Tea Tree Essential Oil That Attract Male Mediterranean Fruit Flies, Ceratitis capitata (Wiedemann).

The Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) poses a major threat to fruit and vegetable production in the United States and throughout the world. New attractants and detection methods could improve control strategies for this invasive pest. In this study, we developed a method that combined thin-layer chromatography (TLC) of tea tree essential oil (TTO) (Melaleuca alternifolia) with short-range bioassays to isolate attractive kairomones for male C. capitata. After development, the TLC chromatogram indicated that TTO separated into five major spots, designated as zones 1 to 5. When the TLC plate was exposed to flies, zones 1 and 3 were strongly attractive to male C. capitata. To confirm activity, the developed TLC plate was cut into five zones which were then tested in short-range bioassays. Again, flies were observed to aggregate around zones 1 and 3, which corresponded with Rf values of 0.93 and 0.59. In addition, zones 1 to 5 were separated using preparative-TLC, and olfactory responses to volatile emissions from the five fractions were quantified by electroantennography (EAG). Highest amplitude EAG responses were recorded with fractions 1 and 3, further supporting the bioactivity of these samples. In conclusion, a TLC-based bioassay system can provide an effective, rapid screening protocol for initial isolation of insect kairomones from complex mixtures such as essential oils or plant extracts. Further analysis of TTO fractions 1 and 3 is needed to identify the specific constituents attractive to male C. capitata.

Inhibitory Effect of Roselle Aqueous Extracts-HPMC 6000 Gel on the Growth of Staphylococcus Aureus ATCC 25923.

Roselle (Hibiscus sabdariffa L.) is a medicinal plant commonly used as a beverage and herbal medicine. Complex compounds in the aqueous extracts have provided good antibacterial activity by which the growth of gram-negative and -positive bacteria is inhibited. The aims of this research were to formulate hydroxypropyl methylcellulose (HPMC) 6000 gel containing the extract and investigate the inhibitory activity of the extract and its gel formula against Staphylococcus aureus ATCC 25923. Thin layer chromatography (TLC) on silica gel GF254 was used for analyzing flavonoids and polyphenols using butanol: acetic acid: water (4:1:5) and chloroforms: ethyl acetate: formic acid (0.5:9:0.5) as eluent, respectively. A serial dilution of aqueous extract powder in citrate buffer was made to obtain 0.50, 0.25, 0.10, 0.05, and 0.02 mg/mL solution. The roselle aqueous extract (3%) was formulated as a component of gel containing HPMC 6000 in various concentrations (2%, 3%, and 4%). A diffusion agar method on two layers of nutrient agar media was applied using Staphylococcus aureus ATCC 25923 and gentamicin 25 ppm as bacterial test and standard, respectively. After incubation for 24 h at 37°C, the inhibitory effect was denoted by a clear zone around the hole and the inhibitory activity was measured as minimum inhibitory concentration (MIC). The aqueous extract of Hibiscus sabdariffa L. contained flavonoid and polyphenol compounds based on the TLC chromatogram profile. It was found that the gel formula containing 3% HPMC 6000 and 3% aqueous extract gave a good physical characteristic and the lowest MIC (6.0 mg/mL), equivalent to 7.58 ppm of gentamicin standard at 12.0 mg/mL concentration. The HPMC 6000 at 3% (w/w) concentration in roselle aqueous extract gel preparation gave good physical characteristics. The gel preparation exhibited inhibitory activity against Staphylococcus aureus ATCC 25923 shown by MIC 6.0 mg/mL. Formula 2 is recommended and should be further investigated for implementation in topical preparations.

Quality Control Of Crude Drugs And Capsules Of Turmeric (Curcuma longa L.)

Turmeric (Curcuma longa L.) is a medicinal plant commonly used in Thai Traditional Medicine (TTM), both in single use and formula. It has been known for its antioxidant activity and applied for management of many diseases. In this article, we reported some aspects of quality control of crude drugs and capsules of turmeric according to the standards set in Thai Herbal Pharmacopeia (THP). The results of our evaluations demonstrated that turmeric crude drugs met the criteria set in THP for microscopical identification, foreign matter (1.31%), loss on drying (6.89±0.174%), ethanol-soluble extractive (13.56%), water-soluble extractive (15.17%), and the profile of Thin Layer Chromatography (TLC) chromatogram. However, its volatile oil content (5.95%) was below the minimum value set in THP. The turmeric capsules met the criteria set in THP for loss on drying (8.64±0.093%), ethanol-soluble extractive (18.07%), water-soluble extractive (14.95%), and profile of TLC chromatogram.

Antibacterial Activity of Leaves and Rhizome of Curcuma xanthorrhiza Essential Oils on Different Distillation Time

This research aims to determine the antibacterial activity of Curcuma xanthorrhiza leaves and rhizomes essential oil with different distillation time of 2 hours, 4 hours and 6 hours against Escherichia coli and Staphylococcus aureus. The essential oils produced then separated by thin layer chromatography (TLC) with n-hexane: toluene: dichloromethane 1:8:1 as the mobile phase. The result shows that on the first two hours distillation gave the highest yield compared to the next two hours or four hours. Based on TLC chromatograms, the highest content of compounds found at 4-6 hours distillation time. TLC bioautography contact was used for antibacterial activity tested on chromatogram and the clear zone appears on it indicates the spots were active as antibacterial activity against both bacteria with Rf values of 0.35 and 0.49 for rhizomes, rhizome essential oils 4-6 hours distillation time as the most significance active.

Characterization of a yellow pigmented coral-associated bacterium exhibiting anti-Bacterial Activity Against Multidrug Resistant (MDR) Organism

Multidrug resistant (MDR) infections have been a world health issue for many decades, and therefore exploration of new antibiotics to overcome this issue is urgently needed. Regarding antimicrobial properties in the coral holobiont, coral-associated bacteria are suggested as potential producer of new antibacterial agents against MDR infections. The purposes of this study include isolation and identification of pigmented coral-associated bacteria, preparation of pigmented extract and evaluation of its antibacterial property, as well as the characterization of the pigmented extract using thin layer chromatography (TLC). Nine bacteria candidates were isolated from an unidentified stony coral collected from Tanjung Gelam, Karimunjawa National Park, Jepara Region, Central Java Province, Indonesia. Bacterial pigment and other metabolites were extracted using 1-butanol, ethyl acetate and acetone. Silica gel-based TLC was applied to detect β-carotene and characterize the bacterial crude extracts. Antibacterial activity was evaluated against the extended spectrum β-lactamase (ESBL) E. coli, Klebsiella pneumoniae, and MRSA (methicillin-resistant Staphylococcus aureus). A yellow pigmented bacterium was isolated and identified as Vibrio owensii TNKJ.CR.24-7. Nonribosomal peptide-synthetase Peptide-Synthetase NRPS genes fragments were detected. All bacterial extracts from 1-butanol, ethyl acetate and acetone contained the yellow pigment that was identified as β-carotene. Only the crude extract from ethyl acetate inhibited all of MDR bacteria. TLC chromatogram showed that there were 7 bands from 1-butanol extract and 6 bands from ethyl acetate extract. However, crude extract from acetone was not well separated using silica-based TLC. Bands at Rf 0.64 and 0.81 were found both in 1-butanol and ethyl acetate extracts.

Ekstraksi Kayu Artocarpus lakoocha: Pengaruh Metode Dan Rasio Bahan Tumbuhan-Penyari Terhadap Rendemen Ekstraksi

Artocarpus lakoocha Roxb. (Ma-haad) is a medicinal plant commonly used in Thai Traditional Medicine (TTM) for a wide array of indications. In this study, we compared yield of extraction and the Thin Layer Chromatography (TLC) profile of the heartwood of A. lakoocha extracted with the different methods and different material-solvent ratios. Extractions were carried out by maceration and reflux extraction with ethanol as the solvent, in material-solvent ratio of 1:5 and 1:3. The TLC profile was obtained from the separation of the extracts with methylene chloride/methanol (85:15) on silica gel F254 plate. The result exhibited that reflux extraction produced the highest yield of extraction and the material-solvent ratio of 1:3 generated more yield than that of 1:5. Nevertheless, the profile of TLC chromatogram of those extracts were similar.

Anti-Tuberculosis Activity of Extract Ethyl Acetate Kenikir Leaves (Cosmos caudatus H.B.K) and Sendok Leaves (Plantago Major L.) By In Vitro Test

The increasing therapy problem including multidrug-resistant tuberculosis (TB) has made it important to discover a new anti-TB drug candidate. The aim of this study was to acknowledge the activity of ethyl acetate extracts of kenikir (Cosmos caudatus H.B.K) and sendok (Plantago major L.) leaves against Mycobacterium tuberculosis (M. tuberculosis) H37Rv. This research used Middlebrook (MB) 7H9 media and observed the growth of M. tuberculosis using Lowenstein Jensen (LJ) media. The concentration of extracts were 0.25 mg/ml, 0.50 mg/ml, and 1.00 mg/ml. The result of this study showed that ethyl acetate extracts exhibited anti-TB activity in 1000 µg/ml of both extracts. The active compound group was detected by thin layer chromatography (TLC) and the separation of compounds was shown by retardation factor (Rf) and the color of the spots. Based on TLC chromatograms, it is known that there are types of compounds, such as ortho-dihydroxy compounds, phenolic compounds, and compound leads to terpenoids for both extracts.

Chromatographic fingerprinting and free-radical scavenging activity of ethanol extracts of Muntingia calabura L. leaves and stems

ObjectiveTo determine the thin-layer chromatography (TLC) fingerprint profiles and to evaluate the in vitro antioxidant activity of ethanol extracts of Muntingia calabura (M. calabura) leaves and stems. MethodsThe leaves and stems were extracted using ethanol as solvent. The TLC separation of the phytochemical constituents of the leaf and ethanol extracts was carried out in ethyl acetate: n-hexane and chloroform: ethyl acetate mobile phase systems. Distinct spots were visualized under visible light, UV 254 nm, UV 366 nm and after spraying with vanillin-sulfuric acid. The 2,2-diphenyl-1-picrylhydrazyl free-radical scavenging assay was used to evaluate the antioxidant activity of the extracts. ResultsBoth the leaf and stem ethanol extracts at 4 mg/mL exhibited 2,2-diphenyl-1-picrylhydrazyl inhibition of more than 90%, relative to gallic acid. The results of TLC showed that the degree of resolution between the constituent spots was comparable between the two mobile phase systems using the different visualization wavelengths. Under the 254 nm visualization, few spots were observed in leaf and stem extracts. Visualization at 366 nm yielded the greatest number of observable spots of various colors in both leaf and stem extracts. More spots were visualized upon post-derivatization with vanillin-sulfuric acid in the TLC chromatograms using chloroform: ethyl acetate mobile phase, compared to those in ethyl acetate: n-hexane mobile phase. ConclusionsM. calabura exhibited very high antioxidant activity in its leaves and stems ethanol extracts, both of which are used in traditional medicine. The TLC results demonstrated the presence of diverse secondary metabolites in the leaf and stem ethanol extracts, indicating that the antioxidant activity, including other bioactivities may be attributed to these phytochemical constituents. This paper has reported for the first time the TLC fingerprinting of M. calabura using visible light, UV 254 nm, UV 366 and post-derivatization with vanillin-spray to visualize separate spots on TLC plates.

Morphological Characteristics, Phenolic and Terpenoid Profiles in Garden &lt;i&gt;Chrysanthemum&lt;/i&gt; Grown in Different Nutritional Conditions

Chrysanthemum indicum L. (Dendranthema indicum (L.) Des Moul) became useful as herbs and curatives in different parts of the world. In the current study, the chemical profile of outdoor cultivated Chrysanthemum indicum ‘Avalone Red’ was investigated for polyphenolic acid derivatives, flavonoids, alantolactone and ursolic acid using thin layer chromatography(TLC). Total phenolic content was determined through a classical method and semiquantitative analyses of alantolactone were performed through the HPLC method. Morphological and yield parameters and the chemical profile of plants were registered in two different nutritional statuses (local and enriched soil conditions). The results indicate that the studiedcultivar is a potent source of phenolics and that the fertilisation increases the plant biosynthetic capacity for polyphenols (159.74 mg/100 g in leaves and 79.82 mg/100 g in flowers, in unfertilised plants, and 388.54 mg/100 g and 144.86 mg/100 g in leaves and flowers of fertilised plants, respectively, expressed as gallic acid equivalent value). The studied cultivar contains hyperoside (and other derivatives) and four main polyphenol carboxylic acids (including chlorogenic acid). Ursolic acid wasnot detected. The high pressure liquid chromatography (HPLC) analyses confirmed the presence of alantolactone identified in TLC chromatogram. Moreover, the fertilised plant samples contain only traces of alantolactone compared to the unfertilised plants. Alantolactone is present in a small amount (less than 0.1 mg %) but its presence alerts to the potential allergenic effect of the plant. Besides their ornamental value, chrysanthemums can have a wide array of uses due to their high amounts of bioactive compounds.

Identifying Type of Refined Petroleum Products in Environmental Media: Thin-Layer Chromatography (TLC) as a Quick Methodology

The ultimate goal of our study is to establish thin-layer chromatography (TLC) as a quick and simple method for identifying the type of refined petroleum products present in the environmental media. As a pre- liminary step, TLC chromatograms of different petro- leum products, including gasoline, kerosene, and diesel, were characterized and compared. Methanol was deter- mined as the optimum carrier solution in TLC analysis. The spherical-shaped TLC chromatogram of gasoline showed the longest migration distance, and thus the highest retardation factor (Rf )o f 0.91. This was follow- ed by that of kerosene (0.63) with an elliptical-shaped, and diesel (0.24) with an elongated trapezoid-shaped chromatogram. Rf of kerosene and diesel increased with the dilution factor, while gasoline showed a constant value. Additionally, it was observed that the TLC chro- matograms of oils produced the same peak pattern with the corresponding petroleum products in gas chroma- tography (GC). A mixed sample of kerosene and diesel presented a triangular shaped chromatogram, underlining the need to consider the shape of chromato- gram in addition to the Rf value, as an indicator of the petroleum type. The findings indicate that TLC has a huge potential to be used as a quick and reliable method for identifying the type of refined petroleum products in the environmental media.

Bioemulsification activity assessment of an indigenous strain of halotolerant Planococcus and partial characterization of produced biosurfactants

Halotolerant bacteria are regarded as effective oil-scavengers in the polluted saltern and seawater. In this regard, a halotolerant Planococcus was isolated from oil-contaminated area of Dezful north springs, Iran, due to its capacity in biosurfactant (BS) production. To facilitate hydrocarbons degradation, in the current study, the efficiency of BS production as function of growth rate of the halotolerant Planococcus was investigated in the vicinity of heavy crude oil by emulsification index (E24). Subsequently, the BS characterization was made by thin-layer chromatography (TLC), gas chromatography (GC) and infrared spectra analysis, and the stability was determined by E24 value measurement over a certain pH (5–9), temperature (20–100 °C) and salt concentration (0–10 % w/v) ranges. The BS production was found to be growth-associated. Detection of a unique band on TLC and GC chromatogram showed the extensive refining capacity of the BS purification, using the medium supernatant under acetone alkaline precipitation followed by oil dissolution from the sediment by carbon tetrachloride. Accordingly, it was clarified that the BS ultimately accumulated outside the cells. The glycolipid quality of the BS was further determined by the routine chemical characterization on TLC and by IR spectra analysis. Moreover, there was no protein detected by lowery total protein assay. Finally, the optimal temperature, pH and NaCl concentration to reach highest E24 values (85.7, 77.0, and 79.0 %) were found at respective 40 °C, pH = 9 and 0 % w/v. Our results revealed the practically potential of strain Dezful Isolate for BS large-scale production as environmentally friendly oil-eliminating agents.

Anti-Inflammatory Effect and Chemical Composition of Bupleurum chinense and Bupleurum kaoi

Bupleurum species represent one of the most successful and widely used herbal drugs for treatment of many diseases over the past 2000 years. Bupleurum chinense (B. chinense) is officially listed in China and Bupleurum kaoi (B. kaoi) is the only indigenous Bupleurum plant in Taiwan. Therefore, the aim of our study is to investigate the chemical components, anti-inflammatory, and antioxidants of B. kaoi and B. chinense. We report highly sensitive and specific assay for anti-inflammatory with a TNF-α luciferase plasmid vector. In preliminary experiments, we found that B. kaoi have stronger anti-inflammatory effects than B. chinense. However, it was no significant difference in the free radical scavenging activities of B. kaoi and B. chinense. We also demonstrated that saikosaponins (SS) a, SSc and SSd are the major compounds of B. kaoi and B. chinense in thin layer chromatography (TLC) analysis. The SSa, SSc and SSd content in B. kaoi has been reported to be higher than that in B. chinense. We combined TLC chromatograms with hierarchical clustering analysis(HCA)to classify into B. chinense and B. kaoi. The major impact feature were Rf 0.58(SSd) and Rf 0.44(SSc) .Theses studies showed that TLC chromatograms with HCA could be as tool in to differentiation and identify different species of herbal medicine in the future.

Enzymatic Decolorization of Malachite Green Dye by a Newly Isolated Bacillus Cereus Strain wwcp

Enzymatic decolorization of Malachite Green (MG) dye was studied using crude enzyme from a newly isolated Bacillus cereus strain wwcp1. 98% decolorization efficiency was achieved within 24 hours using an initial dye concentration of 1.0 x 10 -5 M. Batch experimental results revealed that the decolorization process was highly dependent on contact time, initial MG concentration, aqueous solution temperature and pH. Biodegradation of MG dye was monitored spectrophotometrically and metabolites confirmed by thin layer chromatography (TLC). The comparison of TLC chromatograms before and after decolorization confirmed that crude protease enzyme had the ability to degrade MG dye. The results provide evidence that the crude enzyme from Bacillus cereus strain wwcp1 is an effective and potential candidate for industrial wastewater treatment.

Isolation and Partial Characterization of Bioactive Fucoxanthin fromHimanthalia elongataBrown Seaweed: A TLC-Based Approach

Seaweeds are important sources of carotenoids, and numerous studies have shown the beneficial effects of these pigments on human health. In the present study, Himanthalia elongata brown seaweed was extracted with a mixture of low polarity solvents, and the crude extract was separated using analytical thin-layer chromatography (TLC). The separated compounds were tested for their potential antioxidant capacity and antimicrobial activity against Listeria monocytogenes bacteria using TLC bioautography approach. For bio-autography, the coloured band on TLC chromatogram was visualized after spraying with DPPH and triphenyl-tetrazolium chloride reagents which screen antioxidant and antimicrobial compounds, respectively, and only one active compound was screened on the TLC plate. Preliminary identification of this active compound was done by comparing its colour and R f (retention factor) value with the authentic fucoxanthin standard. Further, the active compound was purified using preparative TLC. This purified compound showed a strong antioxidant (EC50: 14.8 ± 1.27 µg/mL) and antimicrobial (inhibition zone: 10.27 mm, 25 µg compound/disc) activities, which were examined by DPPH scavenging and agar disc-diffusion bioassay, respectively. The bioactivity shown by the purified compound was almost similar to the fucoxanthin standard. The characteristic UV-visible and FT-IR spectra of the purified active compound completely matched with the standard. Hence, the main active compound in H. elongata was identified as fucoxanthin.

Quantification of Phenylpropanoids in Commercial Echinacea Products Using TLC with Video Densitometry as Detection Technique and ANN for Data Modelling

Echinacea preparations are among the most popular herbal remedies worldwide. Although it is generally assigned immune enhancement activities, the effectiveness of Echinacea is highly dependent on the Echinacea species, part of the plant used, the age of the plant, its location and the method of extraction. The aim of this study was to investigate the capacity of an artificial neural network (ANN) to analyse thin-layer chromatography (TLC) chromatograms as fingerprint patterns for quantitative estimation of three phenylpropanoid markers (chicoric acid, chlorogenic acid and echinacoside) in commercial Echinacea products. By applying samples with different weight ratios of marker compounds to the system, a database of chromatograms was constructed. One hundred and one signal intensities in each of the TLC chromatograms were correlated to the amounts of applied echinacoside, chlorogenic acid and chicoric acid using an ANN. The developed ANN correlation was used to quantify the amounts of three marker compounds in Echinacea commercial formulations. The minimum quantifiable level of 63, 154 and 98 ng and the limit of detection of 19, 46 and 29 ng were established for echinacoside, chlorogenic acid and chicoric acid respectively. A novel method for quality control of herbal products, based on TLC separation, high-resolution digital plate imaging and ANN data analysis has been developed. The method proposed can be adopted for routine evaluation of the phytochemical variability in Echinacea formulations available in the market.