Separation Of Bioactive Compounds Research Articles

Green and Efficient Extraction of Phenolic Components from Plants with Supramolecular Solvents: Experimental and Theoretical Studies.

The supramolecular solvent (SUPRAS) has garnered significant attention as an innovative, efficient, and environmentally friendly solvent for the effective extraction and separation of bioactive compounds from natural resources. However, research on the use of a SUPRAS for the extraction of phenolic compounds from plants, which are highly valued in food products due to their exceptional antioxidant properties, remains scarce. The present study developed a green, ultra-sound-assisted SUPRAS method for the simultaneous determination of three phenolic acids in Prunella vulgaris using high-performance liquid chromatography (HPLC). The experimental parameters were meticulously optimized. The efficiency and antioxidant properties of the phenolic compounds obtained using different extraction methods were also compared. Under optimal conditions, the extraction efficiency of the SUPRAS, prepared with octanoic acid reverse micelles dispersed in ethanol-water, significantly exceeded that of conventional organic solvents. Moreover, the SUPRAS method demonstrated greater antioxidant capacity. Confocal laser scanning microscopy (CLSM) images revealed the spherical droplet structure of the SUPRAS, characterized by a well-defined circular fluorescence position, which coincided with the position of the phenolic acids. The phenolic acids were encapsulated within the SUPRAS droplets, indicating their efficient extraction capacity. Furthermore, molecular dynamics simulations combined with CLSM supported the proposed method's mechanism and theoretically demonstrated the superior extraction performance of the SUPRAS. In contrast to conventional methods, the higher extraction efficiency of the SUPRAS can be attributed to the larger solvent contact surface area, the formation of more types of hydrogen bonds between the extractants and the supramolecular solvents, and stronger, more stable interaction forces. The results of the theoretical studies corroborate the experimental outcomes.

DESAIN DAN PEMBUATAN MESIN EKSTRAKSI SUPERKRITIS BERBASIS CO2 CAIR UNTUK MENGEKSTRAKSI SENYAWA BIOAKTIF

Bioactive compounds in animal and plant cells have many benefits for human health, such as antioxidants, antibacterial, anti-inflammatory, and anticancer. Extraction and separation of bioactive compounds from other compounds is an important step, and commonly, conventional methods are used, but these methods have disadvantages, like producing unwanted compounds. Alternative methods can be conducted using supercritical fluid extraction, but this equipment is expensive and has a small capacity. So, this study aims to produce functional and structural designs and manufacture supercritical fluid extraction machines using carbon dioxide solvents (CO2) operating with a semi-continuous system. This research succeeded in designing and manufacturing a supercritical fluid extraction machine using carbon dioxide (CO2) solvent that operates in a semi-continuous system for the extraction of bioactive compounds, with main components including cover frames, supercritical extractor chamber, low and high-pressure CO2 tubes, compressors and boosters, pipelines, direct valves, manometers, heating, cooler, and expanders, result from reservoirs and automatic control. Moreover, the preliminary simulation test studies revealed that the supercritical extractor chamber could withstand an absolute pressure of 1000 bar, a temperature of 300°C, and a work capacity of 1 L. It indicated that the supercritical CO2 fluid extractor system was performing well for the conditioning of the extractor chamber, which is generated using a booster and controlled by a one-way valve. Then, the extract is transferred to the separation chamber to separate the CO2 gas. Then, CO2 gas is returned to the low-pressure CO2 tubes for recycling and reuse for the following process.

Multi‐imaging approach to analyze bioactive compounds in Abelmoschus moschatus and Cinnamomum zeylanicum via planar chromatography hyphenated with effect‐directed assays and high‐resolution mass spectrometry

Citizens in developing countries rely on indigenous knowledge and practices and use locally available medicinal plants for different treatments. Due to limited instrumentation for chemical and biological characterization, most studies investigating the bioactive properties of Sri Lankan medicinal plants rarely progress to the molecular level. While for some plants, the whole plant extracts have been tested, most of the individual active compounds and their effect mechanisms have still not been identified. The separation of bioactive compounds from natural sources is a challenging task. Multi‐imaging high‐performance thin‐layer chromatography (HPTLC‐UV/Vis/FLD) combined with biological/biochemical assays (effect‐ directed analysis, EDA) and high‐resolution mass spectrometry (HRMS) provide the straightforward identification of natural products without prior tedious fractionation and compound isolation. Separating complex plant extracts into individual compounds, and still on the same adsorbent surface, studying their effects highlighted their potential. These findings can be used to substantiate current traditional medicinal knowledge and to evaluate their benefits, risks, and limitations. The developed hyphenated HPTLC‐UV/Vis/FLD‐EDA‐HESI‐ HRMS methods for identification of single compound effects in Sri Lankan Abelmoschus moschatus and Cinnamomum zeylanicum included (1) non‐target screening, (2) assignment of prominent individual bioactive compounds, and (3) comparison of product profiles to check the quality of commercial products. Both studies revealed not only the phytochemical profiles but also prioritized bioactive constituents. Diverse antimicrobials, antioxidants, and inhibitors of glucosidase, tyrosinase, and cholinesterase were detected. Running reference standards in parallel to identified compounds, confirmed the assignments and bioactivities. The sustainable and environmentally friendly technique can be used in developing countries for profiling and valorization of plant‐based preparations.

Advances of responsive deep eutectic solvents and application in extraction and separation of bioactive compounds.

In recent years, it has been found that changing ambient conditions (CO2 /N2 , temperature, pH) can trigger a switchable phase transition of deep eutectic solvents, and such solvents are known as responsive deep eutectic solvents. In this work, we present the development history, properties, and preparation of responsive deep eutectic solvents, followed by the application of responsive deep eutectic solvents in the extraction and separation of bioactive compounds are presented. Importantly, the mechanism of responsive deep eutectic solvents in the extraction of bioactive compounds is discussed. Finally, the challenges and prospects of responsive deep eutectic solvents in the extraction and separation of bioactive compounds are proposed. Responsive deep eutectic solvents are considered green and efficient solvents. Some methods for extraction and separation of bioactive compounds by responsive deep eutectic solvents can increase the possibility of recycling the deep eutectic solvents, and provide higher efficiency in the extraction and separation field. It is hoped that this will provide a reference for the green and sustainable extraction and separation of various bioactive compounds.

Separation of Bio-active Compounds and in vitro Antibacterial Potency of Callistemon linearis Floral Extract with Mechanism of Action

Abstract: Introduction: Infection caused by bacteria is treated with antibiotics which caused several side effects in human and animals. This problem enforced scientist to work out plant originated compounds for treatment of several disease caused by morbific bacteria. Objectives: Present work revealed isolation of bioactive phytocompounds from different fractions of Callistemon linearis flower ethanol extract by chromatographic technique and determination of their antibacterial potency against selected bacteria with mode of action in vitro. Materials and Methods: Antibacterial potency of four different fractions of ethanol extract were done quantitatively via Agar well diffusion method along with determination of Minimum Inhibitory Concentration, and Minimum Bactericidal Concentrations. Further, phytochemical analysis, Fourier Transform Infrared spectroscopic analysis, thin Layer Chromatography separation and Gas Chromatography Mass Spectroscopy analysis of bioactive fractions were done simultaneously to assess the presence of bioactive phytocompounds responsible for the antibacterial potency of the fractions. Time-kill assay, Nucleic acid and protein leakage assay, and FESEM study were employed to determine the mode of activity of the isolated active phytocompounds on the tested bacteria. Results: Maximum antibacterial activity was observed in NHEX of ethanol extract against Escherichia coli (MTCC 739). Chromatographic separation of NHEX detected several bioactive compounds. Assessment of Time-kill assay in treated and untreated bacterial strains determined that bioactive compound of NHEX was bactericidal in nature. Nucleic acid and protein leakage assay of bacterial cells with isolated active compound showed cell damages and FESEM images strongly ascertained this data. Conclusion: The C. linearis floral floral extract possesses antibacterial activities against selected pathogens and causes cellular damages. Keywords: Callistemon linearis, Antibacterial activity, FT-IR, GC-MS analysis, Bioactive compounds, Mechanism of action.

Structural Elucidation and Toxicity Evaluation of Bioactive Compounds from the Leaves and Stem woods of Synadenium glaucescens Pax.

The leaves and stem woods of Synadenium glaucescens Pax. are traditionally used for treatment of various human diseases in Tanzania. However, there is limited information on the structural elucidation and toxicity properties of bioactive compounds responsible for their pharmacological properties. Thus, this study was aimed to elucidate and evaluate toxicity properties of bioactive compounds from these parts. Separation of bioactive compounds was achieved by using column chromatographic method, whereas their structures were determined by GC-MS and NMR data and by comparing with literature spectroscopic data. Toxicity properties of isolated compounds was done by brine shrimp lethality test (BSLT) method. In this study, column chromatographic separation led to isolation of tetracosane (1), hexadecyl palmitate (2), octacosane (3), tetracosanol (4) and β-sitosterol (5) from the leaves and 1-hexacosene (6), campesterol (7), octadecanoic acid (8) and tetracosanoic acid (9) from the stem woods. BSLT of compounds isolated in sufficient amount (1, 2, 3, 4, 5 and 7) were revealed to be non-toxic to brine shrimp larvae (LC50 >100 µg/ml). Except compound 5, this study reports the isolation of other eight (8) compounds for the first time from this plant. Therefore, it can be reasonably concluded that S. glaucescens is a good source of bioactive compounds which justify its traditional uses in treatment of various ailments. Additionally, observed non-toxic effect of the tested compounds indicates safety and it is expected that its traditional use has no toxicity effect.

Simultaneous extraction and separation of compounds from mate (Ilex paraguariensis) leaves by pressurized liquid extraction coupled with solid-phase extraction and in-line UV detection

The in-line coupling of the pressurized liquid extraction with a solid-phase adsorbent and a UV–Vis detector for the simultaneous extraction and separation of bioactive compounds from yerba mate (PLE-SPE-UV) was carried out in two stages. In the first stage, water was used as a solvent, while in the second stage, ethanol was used. For the optimization of the method, different adsorbents (Sepra C18-E, Isolute C18-EC, and Strata-X C18), temperatures (40–80 °C), solvent flow-rate (1–3 mL/min), and pH (4.0 and 8.0) were evaluated. By using a UV–Vis detector on-line, it is possible to monitor the process in real-time. The developed method allowed obtaining similar or higher recoveries of all the compounds classes than other methods, such as ultrasound-assisted extraction, stirring, maceration, and pressurized liquid extraction alone, in addition to separating them into fractions. The developed method could be used as sample preparation for the analysis of different compounds classes from mate.

Application of accelerated solvent extraction coupled with online two-dimensional countercurrent chromatography for continuous extraction and separation of bioactive compounds from Citrus limon peel.

Drug discovery from complex mixtures, like Chinese herbs, is challenging and extensive false positives make it difficult to obtain compounds with anti-Alzheimer's activity. In this study, a continuous method comprised of accelerated solvent extraction coupled with online two-dimensional countercurrent chromatography was developed for the efficient, scaled-up extraction and separation of six bioactive compounds from Citrus limon peels: neoeriocitrin, isonaringin, naringin, hesperidin, neohesperidin, and limonin. These active compounds were isolated and purified from the raw plant materials by two-dimensional countercurrent chromatography separation via two sets of an n-hexane/n-butanol/methanol/water solvent system: 0.23:1.00:0.25:1.13 and 0.47:1.00:0.38:1.46, v/v/v/v. The compounds were collected in yields of 0.22, 0.25, 0.10, 0.31, 0.29, and 0.28mg/g, respectively, with purities of 95.79, 96.47, 97.69, 97.22, 98.11, and 98.82%, respectively. Subsequently, a simple and efficient in vitro method was developed for rapidly evaluating the acetylcholinesterase inhibitory activities of six bioactive components. Furthermore, the PC12 cell model and the in vitro metabolism of cytochromes P450 were employed to verify the monomers obtained from the continuous method. The results demonstrated that these six bioactive extracts from the C. limon peels were strong acetylcholinesterase inhibitors.

Separation of bioactive compounds from epicarp of ‘Hass’ avocado fruit through aqueous two-phase systems

The industrialisation of ‘Hass’ avocado fruit generates residues such as the epicarp that contains bioactive compounds that can be recovered. The objective of the work was to evaluate the extraction of bioactive compounds from ‘Hass’ avocado epicarp through aqueous two-phase systems (ATPS) in order to increase the added value of the fruit. ATPS based on polyethylene glycol 4000 (PEG4000) with sodium citrate (Na3Cit) or magnesium sulphate (MgSO4) were evaluated, both incorporated with lyophilised epicarp of fruit at consumption maturity. Binodal phase diagrams were developed and the salting-out capacity of each system was assessed, which allowed explaining the systems separation behaviour. The extraction of compounds was equivalent to 89.9% in relation to a process based on methanol. Systems with 24.9–14.5% Na3Cit–PEG4000 and 12.2–15.5% MgSO4–PEG4000 were able to recover more than 82% of soluble phenols, flavonoids, and condensable tannins, from avocado peel, with high antioxidant activity. The epicarp of ‘Hass’ avocado fruit had low concentration of anthocyanins. ATPS showed utility in concentrating compounds by a non-thermal procedure.

Design and evaluation of polarity controlled and recyclable deep eutectic solvent based biphasic system for the polarity driven extraction and separation of compounds

Deep eutectic solvent (DES) based extraction methods have promising applications in extracting and separating compounds from plants. In this study, a polarity controlled biphasic extraction system consisting of a hydrophilic DES phase (Hexafluoroisopropanol-choline chloride: HFIP-ChCl) and a hydrophobic DES phase (Menthol-tricaprylylmethylammonium chloride: Menthol-N8881Cl) was constructed. The DES based biphasic system was successful in the simultaneous extraction and separation of high polarity compounds, such as chlorogenic acid (CHA), quercetin (QUE) and anthocyanidins (ANT), and a low polarity compound artemisinin(ART), from Artemisia annua Leaves in a single step by the polarity driven recognition targets. By adjusting the mole ratio of the component in each DES phase, the HFIP-ChCl(1:1)/Menthol-N8881Cl(2:1) biphasic system showed the best extraction and separation performance. A conductor-like screening model for real solvents (COSMO-RS) was used to monitor and analyze the changes in the surface polarity dynamics of each DES. The results showed that the non-polar intensity increased from 11.89 to 21.06 Å2 with increasing proportion of HFIP in HFIP-ChCl and the non-polar intensity weakened from 16.47 to 10.01 Å2 with increasing proportion of Menthol in Menthol-N8881Cl. This model can be efficient for screening suitable DESs in a DES based biphasic extraction system to accommodate compounds with a range of polarities. Supported by the model and experimental data of the DES based biphasic system, 6.21 mg/g ART was transferred to the upper layer (Menthol-N8881Cl with 2:1 mol ratio) and 7.89 mg/g CHA, 5.5 mg/g QUE, and 8.9 mg/g ANT were transferred to the lower layer (HFIP-ChCl with 1:1 mol ratio) under the optimal condition (15/1 solvent/solid ratio, 40 °C extraction temperature, 1:1 vol ratio and 30 min extraction time). Therefore, the polarity controlled DES based biphasic extraction system was used to expand the application of DESs in the extraction and separation of bioactive compounds with various polarities.

Simultaneous extraction and separation of bioactive compounds from apple pomace using pressurized liquids coupled on-line with solid-phase extraction

The objective of this work was the development of an on-line extraction/fractionation method based on the coupling of pressurized liquid extraction and solid-phase extraction for the separation of phenolic compounds from apple pomace. Several variables of the process were evaluated, including the amount of water of the first stage (0–120 mL), temperature (60–80 °C), solid-phase extraction adsorbent (Sepra, Isolute, Strata X and Oasis) and activation/elution solvent (methanol and ethanol). The best results were observed with the adsorbent Sepra. The temperature had a small effect on recovery, but significant differences were observed for phlorizin and a quercetin derivative. Results indicate that ethanol can be used to replace methanol as an activation, extraction/elution solvent. While using mostly green solvents (water, ethanol, and a small amount of methanol that could be reused), the developed method produced higher or similar yields of acids (2.85 ± 0.19 mg/g) and flavonoids (0.97 ± 0.11 mg/g) than conventional methods.

A novel, green cloud point extraction and separation of phenols and flavonoids from pomegranate peel: An optimization study using RCCD

Cloud point extraction (CPE) is one of the novel; environment-friendly; energy; time and cost-effective extraction technique. Therefore, in the present work preconcentration and separation of bioactive compounds from pomegranate peel, a major waste of the pomegranate processing industry was carried out using CPE. CPE works on the principle of entrapping the hydrophobic bioactive compound in the micelle. Surfactant concentration (5–11%), pH (4–8), temperature (30–60 °C), and salt concentration (4–12%) optimization was carried out to separate total phenols and flavonoids with maximum recovery (%R), partition coefficient (Ks/a), fraction concentration (fc), and minimum loss (%L) using the rotatable central composite design (RCCD). The optimum levels were 8.22% surfactant (Triton X-114), 4% salt (NaCl) at temperature of 36.80 °C and pH 4 which resulted in 95% recovery, 14.59 Ks/a, 0.95 fc and 5.27% loss of total phenols with the maximum desirability about 0.846. While in the case of total flavonoids at 8.27% Triton X-114 at pH 5.07/34.30 °C and 4.06% NaCl resulted, 98% recovery, 9.71 Ks/a, 1.01 fc, and 2% loss with 0.842 desirability. The total yield obtained for total phenols was 205.2 mg of GAE/g while for total flavonoids it was 60.05 mg of QE/g of pomegranate peel powder. Therefore, it can be concluded that this method can be successfully applied in the extraction of bioactive from any food systems for clean and green extractives label.

Effects of extraction methods on antibacterial activity of centella asiatica leaves against pathogenic staphylococcus aureus and escherichia coli

Triterpene saponins are remarkable structural and bioactive glycosides in Centella asiatica (pegaga). Since extraction is an important step for separation of bioactive compounds from the plant material, this study was conducted to evaluate the antibacterial activity of C. asiatica leaves extract from different extraction methods, i.e. sonication, soxhlet, magnetic stirring and cold maceration. The efficacy of C. asiatica against Staphylococcus aureus and Escherichia coli by tube dilution and disc diffusion methods at three concentrations, i.e. 25, 50 and 100 mg/mL. Phytochemical screening of the plants showed the presence of alkaloid, tannin, steroid, glycosides and flavonoids. The plant extract with cold maceration method (MIC: 1.56 mg/mL) showed the highest antibacterial activity against S. aureus followed by sonication (MIC: 3.13 mg/mL), magnetic stirring (MIC: 3.13 mg/mL) and soxhlet extraction (MIC: 6.25 mg/mL). Similarly, cold maceration and magnetic stirring method (MIC: 1.56 mg/mL) exhibited the highest antibacterial activity against E. coli. Other extraction methods such as sonication and soxhlet also exhibited promising antibacterial activity against E. coli with MIC value of 3.13 mg/mL. Cold maceration extract had wider zones of inhibition (7.90–8.29 mm) and lowest MIC value (1.56 mg/mL) than other extraction methods. There is a significant difference (P < 0.05) between cold maceration and other extraction methods on the entire test organism, differences exist when compared individually at 100 mg/mL. This study revealed cold maceration method of extraction as the best method in antimicrobial susceptibility assay which could serve as stepping stone to develop C. asiatica as herbal medicine.

Simultaneous HPTLC Densitometric Estimation of KBA and AKBA from Boswellia serrata

Background: Boswellic acids (BAs) are extracted from oleo gum of Boswellia serrata and are utilized as potential anti-inflammatory, hypolipidemic, immunomodulatory and antitumor specialists. The present examination was meant to assess KBA and AKBA in Boswellia serrata separate by High-Performance Thin Layer Chromatography (HPTLC). Methods: The separation of bioactive compounds was performed utilizing mobile phase glacial acetic acid, n-hexane, ethyl acetate and toluene (0.3: 1: 8: 2) (v/v/v/v) and distinguished at wavelength 254 nm. The technique was approved for linearity, precision, accuracy, limit of detection (LOD), limit of quantification (LOQ), and so forth by International Conference on Harmonization guidelines. Results: The calibration range was observed to be 2- 14 μg/band for both the bioactive compounds. KBA was isolated with an Rf estimation of 0.39 ± 0.02 and AKBA with an Rf estimation of 0.42 ± 0.02. The accuracy was seen to be as high as 99.17% and 97.42 for KBA and KBA, respectively. The percentage RSD value for intra-day and between day varieties was under 2%. The system indicated high affectability and specificity. Conclusion: The developed HPTLC method was simple, precise, robust, specific, rapid, and costeffective and could be used for quality control analysis and quantification of KBA and AKBA in different herbal formulations containing the plant species.

BIOACTIVE COMPOUNDS OF RASAMALA (ALTINGIA EXCELSA NORNHA) LEAVES AS C-MYC PROTO ONCOGENE EXPRESSION SUPPRESSOR OF HUMAN TONGUE CANCER CELL IN VITRO

Background : Tongue cancer is a common neoplasm found in oral cavity. It is characterized by aggressive cell growth, poor prognosis and being the cause of mortality. Objectives : to discover bioactive compounds of Rasamala leaves which possess an activity to inhibit SP-C1 human tongue cancer cell proliferation by reducing the expression of c-Myc proto oncogene. Methods : This is an experimental laboratory study using SP-C1 human tongue cancer cell. Separation of bioactive compounds from Rasamala leaves ethyl acetate extract was using various chromatography techniques guided by antiproliferative assay. Results : Two compounds were produced consisting of kaempferol (1) and quercetin (2). Compound 1 and 2 were tested to assess antiproliferative activity of kaempferol and quercetin upon SP-C1tongue cancer cell. IC 50 values obtained from antiproliferative assay of each compound were 0.72 and 0.70 ug/ml respectively. Data analysis using ANCOVA test attained a significant value of α=0.05 and proceeded for probit analysis. The activity of compound 1 and 2 was tested on c-Myc proto oncogene and it was acquired that compound 1 and 2 can suppress c-myc proto oncogene expression. Conclusion : Rasamala compounds consist of kaempferol (1) and quercetin (2) which possess an activity as tongue cancer cell proliferation inhibitor by reducing c-myc proto oncogene expression.

Molecular imprinted polymers based on magnetic chitosan with different deep eutectic solvent monomers for the selective separation of catechins in black tea.

Two types of molecular-imprinted polymers-based magnetic chitosan with facile deep eutectic solvent-functional monomers (Fe3 O4 -CTS@DES-MIPs) were synthesized and applied as adsorbents in magnetic solid-phase extraction (MSPE) for the selective recognition and separation of (+)-catechin, (-)-epicatechin, and (-)-epigallocatechin gallate in black tea. The obtained Fe3 O4 -CTS@DES-MIPs were characterized by Fourier transform infrared spectroscopy and field emission scanning electron microscopy. The selective recognition ability was examined by adsorption experiments. The actual amounts of (+)-catechin, (-)-epicatechin, and (-)-epigallocatechin gallate extracted from black tea using Fe3 O4 -CTS@DES-MIPs by the MSPE method were 13.10, 6.32, and 8.76mg/g, respectively. In addition, the magnetic Fe3 O4 -CTS@DES-MIPs showed outstanding recognition and selectivity. Therefore, it can be used to separate bioactive compounds from black tea. The new-type of DES adopted as the functional monomer in this paper provides a new perspective for the recognition and separation of bioactive compounds.

Enrichment of the Gamma Oryzanol Level from Rice Bran by Addition of Inorganic Salts on Ionic Liquid 1-Butyl-3- Methylimidazolium Hexafluorophosphate ([BMIM] PF6) Extraction

<p style="text-align: justify;"><strong>Objective:</strong> Rice bran oil has many helath benefit. The biologically active compounds of rice bran oil are fatty acids, squalene, tocopherols, phytosterols, tocotrienols, oryzanol, and polyphenols. Gamma oryzanol is a combination of at least 10 components of ferulic acid esters and alcohols triterpene. Gamma oryzanol has pharmacological activity includes cardiovascular disease, antioxidant, anticancer, antidiabetic, antiulcerogenic, neuroprotective and action immunomodulator. Ionic liquids are an environmentally friendly solvent used in the extraction and separation of bioactive compounds from plants. This study was conducted to evaluate the effect of inorganic salts addition on ionic liquid [Bmim]PF₆ in increasing levels of gamma oryzanol from rice bran. <strong>Method:</strong> In the experiments, IL [Bmim]PF₆ - MAE method was used to separated gamma oryzanol compound, and then was partitioned with hexane after the addition of inorganic salts: KH₂PO₄, NaCl, NaNO₃ and Na₂CO₃, respectively. Gamma oryzanol content was determined by HPLC analysis. A mixture of mobile phase (methanol:acetonitrile:isopropan ol (5:4:1) and isocratic conditions at a wavelength of 327 nm. The flow rate is set at 1 ml / min..Results: The highest levels of &gamma;-oryzanol was produced from the addition in KH₂PO₄ + : [Bmim] PF₆ by 0.26 &plusmn; 0.001 mg/g. The mechanism of salting-out effect of the compound due to the interaction of different types of interactions between solutes (ions of inorganic salts and ionic liquid) and solvent. <strong>Conclusion:</strong> The addition of inorganic salt increased the gamma oryzanol level. The addition of KH₂PO₄ on IL-MAE gave the highest level of gamma oryzanol. <p style="text-align: justify;"><strong>Key words:</strong> [BMIM]PF₆, ionic liquid, &gamma;-oryzanol, IL-MAE, inorganic salts.

Simultaneous estimation of genistein and daidzein in Pueraria tuberosa (Willd.) DC by validated high-performance thin-layer chromatography (HPTLC) densitometry method

ABSTRACTThe present study was performed to estimate the concentration of genistein and daidzein in ethanol extract of tubers of Pueraria tuberosa (Indian kudzu or Vidarikanda) and its various fractions (n-hexane, ethyl acetate, n-butanol, and aqueous) by high-performance thin-layer chromatography (HPTLC). The separation of bioactive compounds was performed using mobile phase, toluene:ethyl acetate:acetone:formic acid (20.0:4.0:2.0:1.0) and detected at wavelength 269 nm. The method was validated for linearity, accuracy, precision, limit of detection (LOD), limit of quantification (LOQ), etc. by International Conference on Harmonization guidelines. The calibration range was found to be 100–600 ng/band for both the bioactive compounds. Daidzein was separated with an Rf value of 0.39 ± 0.02 and genistein with an Rf value of 0.54 ± 0.02. Average recovery was 99.96 and 99.90% for genistein and daidzein, respectively. The LOD and LOQ were 14.786 and 44.805 ng, respectively, for genistein, and 9.607 and 29.114 ng, respectively, for daidzein. Both the phytoconstituents were found in ethanol extract and its ethyl acetate fraction only. The developed HPTLC method was simple, precise, robust, specific, rapid, and cost effective and could be used for quality control analysis and quantification of genistein and daidzein in different herbal formulations containing the plant species.

Antimicrobial activity of puffball(Bovistella radicata) and separation of bioactive compounds

BackgroundTo test the antimicrobial activity of different extracts and fermentation broth from puffball(Bovistella radicata), the different extracts and fermentation broth of puffball were prepared, the active fraction was investigated by UPLC–UV–MS and semi-preparative chromatograph.ResultsThrough zones of inhibition (ZOI) and minimum inhibitory concentrations (MIC) tests, the supernatant of fermentation possessed best antimicrobial activity in all extracts whose MIC value is 31.2 μg/ml against T. rubrum, T. mentagrophytes, S. aureus and P. aeruginosa. And ZOI value is 29.01, 21.02, 35.02, 28.01 mm against T. rubrum, T. mentagrophytes, S. aureus and P. aeruginosa. Then we compare the puffball fermentation supernatant with blank contrast by LC–MS. There are the characteristic peaks named PBR-1 and PBR-2 with the puffball fermentation supernatant, the separation of compound PBR-1 and PBR-2 was done on semi-preparative C18 column and the MIC and ZOI of compound PBR-1 and PBR-2 are 15.6 μg/ml and 34 mm with the antifungal test.ConclusionsThe fermentation supernatant and compound PBR-1 and PBR-2 have promising antifungal activity against T. rubrum and T. mentagrophytes.

Separation of Biologically Active Compounds by Membrane Operations.

Bioactive compounds from various natural sources have been attracting more and more attention, owing to their broad diversity of functionalities and availabilities. However, many of the bioactive compounds often exist at an extremely low concentration in a mixture so that massive harvesting is needed to obtain sufficient amounts for their practical usage. Thus, effective fractionation or separation technologies are essential for the screening and production of the bioactive compound products. The applicatons of conventional processes such as extraction, distillation and lyophilisation, etc. may be tedious, have high energy consumption or cause denature or degradation of the bioactive compounds. Membrane separation processes operate at ambient temperature, without the need for heating and therefore with less energy consumption. The "cold" separation technology also prevents the possible degradation of the bioactive compounds. The separation process is mainly physical and both fractions (permeate and retentate) of the membrane processes may be recovered. Thus, using membrane separation technology is a promising approach to concentrate and separate bioactive compounds. A comprehensive survey of membrane operations used for the separation of bioactive compounds is conducted. The available and established membrane separation processes are introduced and reviewed. The most frequently used membrane processes are the pressure driven ones, including microfiltration (MF), ultrafiltration (UF) and nanofiltration (NF). They are applied either individually as a single sieve or in combination as an integrated membrane array to meet the different requirements in the separation of bioactive compounds. Other new membrane processes with multiple functions have also been developed and employed for the separation or fractionation of bioactive compounds. The hybrid electrodialysis (ED)-UF membrane process, for example has been used to provide a solution for the separation of biomolecules with similar molecular weights but different surface electrical properties. In contrast, the affinity membrane technology is shown to have the advantages of increasing the separation efficiency at low operational pressures through selectively adsorbing bioactive compounds during the filtration process. Individual membranes or membrane arrays are effectively used to separate bioactive compounds or achieve multiple fractionation of them with different molecule weights or sizes. Pressure driven membrane processes are highly efficient and widely used. Membrane fouling, especially irreversible organic and biological fouling, is the inevitable problem. Multifunctional membranes and affinity membranes provide the possibility of effectively separating bioactive compounds that are similar in sizes but different in other physical and chemical properties. Surface modification methods are of great potential to increase membrane separation efficiency as well as reduce the problem of membrane fouling. Developing membranes and optimizing the operational parameters specifically for the applications of separation of various bioactive compounds should be taken as an important part of ongoing or future membrane research in this field.