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Presented are five procedures which permit the effective teaching of chromatography with equipment which is readily available, economical, and simple in design. The first procedure involves a study of solute partition in two immiscible solvents and of countercurrent distribution. The second illustrates the use of unidimensional ascending paper chromatography in separating the same mixture of colored components used in the first procedure. The third procedure illustrates the separation of three common sugars by unidimensional ascending paper chromatography. The fourth procedure demonstrates the separation of amino acids by ascending two-dimensional paper chromatography. The fifth procedure involves the separation of a dye mixture using thin-layer chromatography. (DS)

Describes a method in which glass plates are coated with adsorbent, using a technique which is cheap, rapid, and reliable; avoids the need for expensive, commercially-available thin layer chromatography equipment. (MLH)

New systems for analyzing large numbers of body fluid components can be used in the clinical laboratory. (Author/RH)

Three different problems are given for student investigation in determining amino acid compositions, floral pigments, and water soluble amino acids by using the techniques of Roll Chromotography, DISC Chromotography, Thin Layer, and Paper Chromotography. (BR)

In this second article on thin layer chromatography six kinds of experiments are suggested for science teachers to use in schools. (PS)

Explains the separation of the constituents of mixtures on one piece of chromatography paper. The example presented involves a vitamin C tablet, a disprin tablet, and a glucose tablet. Outlined are two methods for separating the constituents. (GS)

Described is thin-layer chromatography (TLC), a technique that can be conveniently used in the laboratory to generate evidence supporting the principle that degrees of biochemical similarity reflect degrees of evolutionary relatedness among organisms. (Author/PEB)

Describes how to construct thin layer chromatography apparatus by using a sheet of glass, glass strips, epoxy resin and a chromatographic plate. (LS)

This study was conducted to determine the phenolic compounds that may be utilized as chemotaxonomic study to identify some phenolic compounds concentration that are useful for pharmaceutical purposes from 5 taxa belongs to the Brassicaceae family which are; Cardaria draba (L.) Desv., Lepidum aucheri Boiss., Sisymbrium irio L., Sisymbrium officinale (L.) Scop., and Sisymbrium septulatum DC. Leaves powder of these taxa were collected and analyzed by High-performance liquid chromatography (HPLC). The results identified 9 phenolic compounds, chlorogenic acid, cinnamic acid, ferulic acid, genstic acid, kaempferol, p-coumaric acid, protocatechuic acid, sinapic acid, and vanillic acid. The results showed that the genus Lepidium aucheri had the highest concentration of 4 phenolic compounds, cinnamic acid, genstic acid, p-coumaric acid, and sinapic acid with concentration of (37.2, 80.79, 62.91, and 37.10µg/ ml) respectively. On the other hand, the genus Cardaria draba showed the lowest concentration of 3 phenolic compounds, cinnamic acid, p- coumaric acid, and protocatechuic acid, and doesn't have 2 phenolic compounds, genstic acid and kaempferol. This variation in the phenolic concentrations might be due to the genotype and plant growth stage differences. The results of this study provide a source for bioactive compounds to future studies. http://dx.doi.org/10.25130/tjps.23.2018.027

BackgroundEnvironmental water samples are increasingly recognised as an important reservoir of antimicrobial resistance (AMR) genes. Polymerase chain reaction (PCR) and next generation sequencing (NGS) offer a potentially inclusive surveillance platform for a wide range of AMR genes. However, molecular methods are dependent upon the extraction of DNA of high yield and quality. Current options for DNA extraction from complex environmental matrices for downstream molecular applications are either expensive or low yielding. We present here a novel magnetic bead-based DNA extraction method, for the detection of antimicrobial resistance genes (ARGs) from river water in Malawi, named MagnaExtract. MethodsMagnaExtract involves initial filtration of 250ml freshwater, followed by an overnight incubation of the filter in 15ml buffered peptone water (BPW), common procedure in microbiology laboratories. 200{micro}l is then taken for a boil (95{degrees}C) and spin step and mixed with magnetic beads to bind DNA. Following washes with ethanol, the DNA is eluted in nuclease-free water. To determine the effectiveness of this method, 98 freshwater samples were collected from two rivers in Southern Malawi, and DNA was isolated using the MagnaExtract method, two commercial Qiagen (Germany) kits; PowerWater and DNeasy Blood and tissue, alongside a boil and spin of BPW, and a boil and spin from bacterial isolate grown on agar media. All samples were screened with a high-resolution melt (HRM) PCR panel previously validated for the detection of third generation cephalosporin and carbapenem ARGs. We compared the DNA yield obtained using all extraction methods, as well as the identification of each ARG. ResultsDNA yield using MagnaExtract was statistically greater than both boil and spin methods and DNeasy Blood & Tissue (Qiagen, Germany). DNA yield was slightly lower than using PowerWater (Qiagen) but the difference was not statistically significant. MagnaExtract was the only method to identify ARGs in all 98 water samples compared with PowerWater (n=82), DNeasy (n=95) boilate of BPW (n=75) and boilate of bacterial isolate (n=87). The most commonly detected ARG was OXA-48 (n=93). In addition, we found overnight incubation in non-selective enrichment broth (BPW) to promote the growth of bacteria harbouring extended spectrum beta lactamase (ESBL) genes and reduction in the detection of carbapenemase genes. ConclusionThe MagnaExtract approach offers a simple, affordable, high yielding DNA extraction method for the detection of ARGs isolated from river water samples.

The main aim of this study is providing of Mentha longifolia L essential oil emulsion loaded with omega 3 fatty acids and optimization by polyaniline (PANI) nano-fiber/gas chromatography (PANI-F/GC). The head space solid phase microextraction (HS-SPME) method by polyaniline nanofibers was used to extract released essential oils of emulsion. The effects of four factors, including omega 3 percent, surfactant-to-oil ratio (SOR%), surfactant type (TWEEN 20, 80 and caseinate sodium) and storage time on the encapsulation efficiency, and essential oil chromatographic characteristics including total peak number, total peak area and total peak height were studied. The Central Composite Design (CCD) was used to design experiments and study the effects of four factors on the responses and P-value of

Anisodus tanguticus (Maxim.) Pascher is an important Tibetan folk medicine and the source of tropane alkaloids (TAs) grown in Qinghai-Tibet Plateau. There are marked differences in quality of A. tanguticus from geographic areas. The aim of present research was to establish a method for the quantitative analysis of TAs coupled with chemometrics analysis to trace geographical origins. Qualitative analysis of TAs in A. tanguticus was carried out using ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry and quantitative analysis of TAs in different plant organs from different geographical origin was achieved. Contents of TAs were subjected to the principal component analysis, and orthogonal partial least-squares discriminant analysis. The contents of the three marker compounds (anisodamine, anisodine and atropine) in the roots and acrial parts of A. tanguticus were positive correlated and varied significantly from different geographical origins. Principal component analysis, and orthogonal partial least-squares discriminant analysis results showed excellent discrimination between different geographical origin of A. tanguticus. This study could provide comprehensive evaluation and further utilization of A. tanguticus resources.

A validated UHPLC-PDA with an ESI-MS/MS method has been developed for simultaneous estimation of six bioactive alkaloids (magnoflorine, berbamine, columbamine, jatrorrhizine, palmatine and berberine) in the different extracts of the roots of Berberis aristata DC (Family:Berberdiaceae). It is an important medicinal herb native to Northern Himalaya and commonly known as ‘daruharidra’, ‘daruhaldi’, ‘Indian barberry’ or ‘tree turmeric’. An insight into the research literature uncovered reports on isoquinoline alkaloids like magnoflorine, berbamine, columbamine, jatrorrhizine, palmatine, and berberine as major bioactives in B. aristata roots, possessing different pharmacological and therapeutic effects. In the present study, these aforementioned alkaloids were separated on Phenomenex Luna®, 5 µm-C8 analytical column. The HPLC-MS analysis was performed at a flow rate of 0.90 mL min−1. Each alkaloid that is resolved was characterized by precursor ions and fragment ions with electrospray ionization (ESI) source in both positive and negative ionization using scan mode. The limit of detections (LODs) were 0.087, 0.727, 0.035, 0.124, 0.782 and 0.794 μg mL−1 for magnoflorine, berbamine, columbamine, jatrorrhizine, palmatine and berberine, respectively. The proposed UHPLC-PDA method was fully validated according to international (ICH) guidelines and was found to be selective, sensitive and highly accurate for the concomitant estimation of the aforementioned symbolic bio-markers of B. aristata roots.

Antibody-drug conjugates (ADCs) are commonly heterogeneous and require extensive assessment of exposure-efficacy and exposure-safety relationships in preclinical and clinical studies. In this study, we report the generation of a monoclonal antibody against monomethyl auristatin E (MMAE) and the development, validation, and application of sensitive and high-throughput enzyme-linked immunosorbent assays (ELISA) to measure the concentrations of MMAE-conjugated ADCs and total antibodies (tAb, antibodies in ADC plus unconjugated antibodies) in cynomolgus monkey sera. These assays were successfully applied to in vitro plasma stability and pharmacokinetic (PK) studies of SMADC001, an MMAE-conjugated ADC against trophoblast cell surface antigen 2 (TROP-2). The plasma stability of SMADC001 was better than that of similar ADCs coupled with PEG4-Val-Cit, Lys (m-dPEG24)-Cit, and Val-Cit linkers. The developed ELISA methods for the calibration standards of ADC and tAb revealed a correlation between serum concentrations and the OD450 values, with R2 at 1.000, and the dynamic range was 0.3–35.0 ng/mL and 0.2–22.0 ng/mL, respectively; the intra- and inter-assay accuracy bias% ranged from −12.2% to −5.2%, precision ranged from −12.4% to −1.4%, and the relative standard deviation (RSD) was less than 6.6% and 8.7%, respectively. The total error was less than 20.4%. The development and validation steps of these two assays fulfilled the acceptance criteria for all addressed validation parameters, which suggested that these can be applied to quantify MMAE-conjugated ADCs, as well as in PK studies. Furthermore, these assays can be easily adopted for development of other similar immunoassays.

Background: The fruit pulp decoction of Crescentia cujete, commonly known as calabash, is traditionally used for the treatment of several respiratory diseases and is available as syrup formulations. Unfortunately, there is no detailed investigation on analytical methods for warranting the quality of these products. Aim and objective: To develop and validate an appropriate analytical method for the simultaneous quantification of transcinnamic acid, 4-hydroxybenzoic acid, verbascoside and 6-epi-aucubin in the decoction and commercial cough syrups of Crescentia cujete fruit. Materials and methods: A reversed-phase ultra-high-performance liquid chromatographic method coupled to a diode array detector (UPLC-DAD) was validated following the ICH guidelines. The chromatographic analysis was performed using a C18 column, the mobile phase system consisted of water and acetonitrile containing 0.1% formic acid, and UV chromatograms were recorded from 200 to 400 nm. Results: A new UPLC-DAD method was validated for the simultaneous quantification of trans-cinnamic acid, 4- hydroxybenzoic acid, verbascoside and 6-epi-aucubin in calabash-derived products. After successful validation, this method was applied for the quantification of the selected chemical markers in an in-house decoction and three commercial cough syrups. Among the selected chemical markers, 6-epi-aucubin was the main compound in the calabash decoction, while trans-cinnamic acid and 4-hydroxybenzoic acid were the major compounds in the commercial products. Verbascoside and 6-epi-aucubin were below the limit of quantification in all syrup samples. Conclusion: The proposed method was successfully applied for the analysis of three commercial syrup formulations and can be useful for standardization and quality control of raw and pharmaceutical calabash preparations.

Background: Polyhydroxyalkanoates (PHAs) are bacteria-synthesized biopolymers under imbalanced growth conditions. These biopolymers are acknowledged as potential biomaterials for future applications because of their characteristics of biocompatibility and biodegradability, and ability to be produced rapidly, and strong functionality of mechanical resistance. This article aims to perform microbial fermentation using the Pseudomonas putida strain to identify the quantity of biopolymers, particularly of the medium-chain-length (mcl-PHA) polyhydroxyalkanoates, based on the type and quantity of the added precursors (glucose and fatty acids). Methods: To understand the microbial interaction and the mechanism involved in PHA biosynthesis, several methods were employed and microbial biomass was obtained using the Pseudomonas putida strain capable of producing PHA. The polymer production by acetone extraction was analyzed using the Soxhlet method, while the biopolymer purification was done via the methanol-ethanol treatment, after which the biomass estimation was done through spectrophotometric analysis. This was followed by measuring the dry weight of the cells and quantification of the biopolymer produced using the gas chromatography method (GC). Results: The highest PHA yield was obtained using the octanoic (17 mL in 2000 mL medium) and hexanoic acids (14 mL in 2000 mL medium) as the precursors. As a result, the octanoic acid - octanoic acid, heptanoic acid – nonanoic acid, and octanoic acid - hexanoic acid were identified as the different precursors that supported the quantity of PHA obtained. Conclusion: Among the 4 types of structurally related substrates, the Pseudomonas putida ICCF 319 strain showed a preference for the C8 sublayer for the biosynthesis of the elastomeric PHAs composed predominantly of more C8 monomers than the C6 and C10.

Plants secondary metabolites undergoes qualitative and quantitative variation due to environmental and growth factors. It is a crucial factor to select the proper time for collection of medicinal plants to assure maximum content of active components reflected as maximum efficacy. Olive leaves (Olea europaea L.) are known traditionally for their antidiabetic effect. The secoiridoid glycoside oleuropein is the main active component of Olive leaves responsible for the biological activity. The current study was conducted to monitor the seasonal variation of oleuropein in Olives leaves collected from the same location. To achieve this goal a validated HPLC method following the ICH guidelines was established. Separation was conducted using RP18 column and a mobile phase consisted of ultrapure water containing 20% acetonitrile and 1% acetic acid. Detection was performed at 254 nm with 1 mL/min flow rate. The method was simple, linear, accurate, precise, specific and robust. The analyses revealed considerable variations in the level of oleuropein throughout the year. This variation cannot be explained by temperature variation during the year. Two points of high levels of oleuropein were detected prior to flowering stage and ripening of the fruits. The levels of growth regulators most likely is responsible for the increased production of oleuropein. It is recommended that leaves intended for medicinal use to be collected during the fruiting stage prior to fruit ripening.

Rapid screening of foodborne pathogens is of great significance to ensure food safety. A microfluidic biosensor based on immunomagnetic separation, enzyme catalysis and electrochemical impedance analysis was developed for rapid and sensitive detection of Salmonella typhimurium. First, the bacterial sample, the magnetic nanoparticles (MNPs) modified with capture antibodies, and the enzymatic probes modified with detection antibodies and glucose oxidase (GOx) were simultaneously injected into the microfluidic chip, followed by mixing and incubation to form MNP-bacteria-probe sandwich complexes. Then, glucose with high impedance was injected into the chip and catalyzed by the GOx on the complexes into hydrogen peroxide with high impedance and gluconic acid with low impedance, which was finally measured using the low-cost interdigitated microelectrode and the electrochemical impedance analyzer to determine the target bacteria. Under the optimal conditions, this biosensor could quantitatively detect Salmonella typhimurium at the concentrations from 1.6 × 102 CFU/mL to 1.6 × 106 CFU/mL in 1 h with the low detection limit of 73 CFU/mL. Besides, this biosensor was demonstrated with good feasibility for practical applications by detecting the Salmonella typhimurium spiked chicken meat samples.

Background: The marine algae are considered a diverse source of bioactive compounds. Many active compounds have been isolated from algae and show good biological activities. Materials and Methods: The aim of this study is to detect the antiviral and anticancer activities in some extracts of marine algae. Extraction, purification and identification of some marine algae common in Egypt were conducted. Extraction of Ulva lactuca, Sargassum dentifolium, and Cystoseiara myrica was conducted. A sequence of extractions, including extraction by ethanol, boiling water, hydrochloric acid and sodium hydroxide were carried out. The obtained extracts were evaluated for their antitumour and antiviral activities against liver tumour cells, brain tumour cell lines, measles virus, mumps virus and hepatitis B virus (HBV). The extracts of the best activities were subjected for purification by size exclusion chromatography and anion exchange chromatography for ethanolic extracts or precipitation by cetylpyridinium chloride (CPC) then by size exclusion chromatography and anion exchange chromatography for aqueous extracts. Separation by GLS/MS was performed. The structures of the active compounds have been identified through different chemical analyses, including sugar analysis, configurational analysis, High-Performance Liquid Chromatography (HPLC), infrared spectroscopy (IR), Gas-Liquid Chromatography-Mass Spectroscopy (GLC-MS) and 1H,13C nuclear magnetic resonance (NMR) at ZV. Results: The active compounds from the water extracts have been identified mainly as polysaccharides and sulphated polysaccharides. The antitumour and the antiviral activities of ethanolic extracts are attributable to compound identified as Ethyl Palmitate. These natural compounds did not show cytotoxic effect. Conclusion: These outputs could be preliminary for further biological studies aiming to therapeutic application.