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La cromatografía de Pfeiffer se basa en los principios de la cromatografía en papel, en la actualidad es una herramienta útil y económica para conocer la fertilidad de los suelos. Sin embargo, existe un rezago en la interpretación de los cromatogramas resultantes, lo que dif iculta su uso. El objetivo de este estudio fue generar modelos de predicción para la interpretación de los cromatogramas generados con la técnica de cromatografía de Pfeiffer. Se evaluaron características físicas, químicas y biológicas de muestras de suelo de un huerto de manzano con diferente manejo agronómico mediante la cromatografía de Pfeiffer y análisis convencionales de laboratorio. Los resultados fueron correlacionados estadísticamente, lo que permitió detectar los componentes del cromatograma de mayor interacción para estimar la fertilidad del suelo. Posteriormente a través de modelos de regresión se establecieron patrones numéricos para estimar e interpretar los contenidos de P-extraíble, K+, Cu2+ y materia orgánica, y por medio de categorización para encontrar tendencias de los contenidos de arena, Ca2+, Mg2+, Fe2+, Mn2+ y Zn2+. Los resultados demostraron que, bajo las condiciones en que se realizó esta investigación, la fertilidad del suelo puede estimarse a través de la cromatografía de Pfeiffer y que los patrones generados en este estudio contribuyen en su interpretación.

The use of synthetic stimulants, including designer cathinones, remains a significant concern worldwide. Thus, the detection and identification of synthetic cathinones in biological matrices is of paramount importance for clinical and forensic laboratories. In this study, distribution of mephedrone and its metabolites was investigated in fingerprints. Following a controlled human mephedrone administration (100 mg nasally insufflated), two mass spectrometry-based methods for fingerprint analysis have been evaluated. The samples deposited on triangular pieces of chromatography paper were directly analysed under ambient conditions by paper spray-mass spectrometry (PS-MS) while those deposited on glass cover slips were extracted and analysed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The LC-MS/MS method was 5-6 times more sensitive than PS-MS but required sample preparation and longer analysis time. Mephedrone was detected in 62% and in 38% of all post-administration samples analysed by LC-MS/MS and PS-MS, respectively. Nor-mephedrone was the only metabolite detected in 3.8% of all samples analysed by LC-MS/MS. A large inter- and intra-subject variation was observed for mephedrone which may be due to several factors, such as the applied finger pressure, angle and duration of contact with the deposition surface and inability to control the 'amount' of collected fingerprint deposits. Until these limitations are addressed, we suggest that the sole use of fingerprints can be a useful diagnostic tool in qualitative rather than quantitative analysis, and requires a confirmatory analysis in a different biological matrix.

The adulteration of herbal medicines by dexamethasone or prednisolone is regarded as a serious problem in many communities. Herein, a novel platform for the separation and quantification of both target steroids in herbal medicines based on electrochemical paper-based analytical devices (ePADs) has been created. The ePAD was composed of Whatman SG81 chromatography paper, 3D-printed devices and a commercial screen-printed electrode. Whatman SG81 silica-coated paper was used for the separation of dexamethasone and prednisolone based on the difference in their partition coefficients during the flow of the mobile phase. The optimal mobile phase was composed of 60% ethyl acetate in cyclohexane and required 7 min for separation. The separated steroids on the paper were then quantified by electrochemical detection using differential pulse voltammetry, in which the 3D-printed devices facilitated the measurement. Analytical detection ranges of 10–500 μg mL−1 were obtained for both dexamethasone and prednisolone (r2 = 0.988 and 0.994, respectively). The limits of detection for dexamethasone and prednisolone were 3.59 and 11.98 μg mL−1, respectively, whereas the limits of quantification were 6.00 and 20.02 μg mL−1, respectively. The amounts of both target steroids derived from real herbal medicine samples determined by the proposed method were comparable to those obtained with assays using standard high-performance liquid chromatography. In addition, a simple evaporation step can be used to increase the concentration of the samples before analysis. These ePADs are simple, low-cost, rapid, and very promising for on-site quantitative detection.

This paper describes the development of two microfluidic paper-based analytical devices (μPADs), one well-based and the other based on a lateral flow assay (LFA) configuration, to detect glucose via a colorimetric assay using the solid metal-organic framework (MOF) Zr-PCN-222(Fe), to encapsulate glucose oxidase (GOx). The well-based platform consisted of laminate sheets and multiple layers of wax-printed chromatography paper. Solutions of KI and glucose placed into the well flowed through the device and reacted with the GOx@MOF species sandwiched between the paper layers realizing a yellow-brown color. The LFA platform consisted of chromatography paper between parafilm and polyvinyl acetate (PVA) layers. GOx@MOFs spotted on the paper subjected to solutions of KI and glucose yielded a brown color. The devices were then dried, scanned, and analyzed yielding a correlation between average inverse yellow intensity and glucose concentrations. The development of these devices employing MOFs as biomimetic catalysts should further expand the applications of microfluidic technologies for sensors a variety of analytes.

Diethylenetriamine penta-acetic acid (DTPA), when complexed with a gamma (γ)-emitter radioisotope like 99m Tc, is used for renal function diagnosis and many other diagnostic applications. The main aim of this study was to develop a novel and versatile single-step methodology for the synthesis of a new 177 Lu-labeled radiopharmaceutical with high radiochemical yield, which can be used for diagnostic purposes and therapeutic purposes also. The single and well-defined 177 Lu-DTPA complex was radiochemically characterized by paper chromatography, thin-layer chromatography, high-performance liquid chromatography, and electrophoresis techniques. Dependence of the labeling yield of 177 Lu-DTPA complex on different factors was studied in detail. Biological evaluation was also performed in a normal rabbit by developing images under a γ camera at various time intervals. More than 99% labeling yield was obtained by reacting DTPA with 177 Lu at specific conditions (pH 7.0, 15 minutes reaction time at 100 °C). 177 Lu-DTPA complex showed high stability both at room temperature and in vitro. Biodistribution studies in normal mice indicated the fractional renal uptake of intravenously administered 177 Lu-DTPA complex, which reached in the kidneys within 2-3 minutes. Scintigraphy showed rapid clearance from the body. Based on these results, we propose that 177 Lu-DTPA complex might be used as an ideal candidate for functional evaluation of kidneys and the urinary tract, especially when needed to be transported to long-range consumer sites, because of its suitable half-life.

Researching of the amino acids contents in medicinal plant raw materials is an important task of pharmaceutical science and practice, because complexes of amino acids used for the corrections of hepatobiliary disorders, nervous and cardiovascular systems. The aim of this study was to conduct a qualitative and quantitative analysis of amіno acids composition of these representatives. The article presents the results of the amino acid composition investigation of the above-ground parts of 10 species belonging to Lamiaceae Juss. Family (Ocimum, Hyssopus, Dracocephalum, Lophanthus, Monarda and Satureja genera). By paper chromatography it was identified several amino acids. HPLC analyzes revealed the content and composition of more than 20 amino acids. The highest total amino acid composition was found in the herb of the genus Ocimum. It was identified 10 essential and non-essential 9 amino acids in each herb, and 3 nonproteinogenic amino acids also. It was established that in all plant materials dominate quantitatively nonessential amino acids (asparagine, proline, glutamic and aspartic acid). Among the essential amino acids arginine and threonine predominate.

The water-soluble acidic polysaccharide from Thalassodendron ciliatum (Forss.) den Hartog was successfully extracted, fractionated and purified. The phytochemical profile of the two water-soluble fractions (F1 and F2), were detected using different analytic techniques. GC-MS analysis revealed the presence of 22 saccharide. Acidic polysaccharide, galacturonic and glucuronic acid were the most abundant. Moreover, paper chromatography and electrophoresis also performed as a preliminary chemical characterization of the polymer. The hepatoprotective activity of the fractions against thioacetamide (TAA) induced liver failure; antioxidant potential and preliminary immunomodulatory activity were assigned in-vivo. The results revealed a potent competence to improve the liver function profile (ALT, AST, total bilirubin, total glyceride, etc.) and a remarkable improvement in liver architecture in comparison to the challenged intoxicated groups. Moreover, they showed high anti-oxidative properties and a promising immunomodulatory influence via Il6. These findings provide new insight into the possible role of polysaccharide purified two fractions in the treatment of acute liver injury.

In this work, a low-cost, disposable, and portable lab-on-paper device is proposed to simultaneously quantify total polyphenol content (TPC) and antioxidant capacity (AOC) in 15 min; the assay requires no pre-treatment of the samples. The lab-on-paper device fabrication has been carried out employing a xurography-based benchtop microfabrication technology using low-cost materials as chromatography paper and polymeric sheets. Extra virgin olive oil (EVOO) phenolic compounds' represents a nutritional added value, nevertheless, the high lipidic content hinders their direct and rapid analysis, resulting in an extremely challenging sample. The realized lab-on-paper allows to perform the dual TPC and AOC determination in three simple steps: (i) sample loading, (ii) analytes transport to the analysis spot, and (iii) double colorimetric analysis exploiting the growth of AuNPs and AgNPs on paper mediated by phenolic compounds. Signal acquisition is achieved using a standard digital camera. The dual colorimetric assay is able to detect phenolic compounds in the 25-500 mg L-1 range with limits of detection ≤6 mg L-1 and good reproducibility (RSDs ≤11%). Direct analysis of EVOO samples (n = 30) correlated well (r > 0.92) with conventional spectrophotometric methods for TPC and AOC determination.

DNA methylation serves to mark DNA as either a directed epigenetic signaling modification or in response to DNA lesions. Methods for detecting DNA methylation have become increasingly more specific and sensitive over time. Conventional methods for detecting DNA methylation, ranging from paper chromatography to differential restriction enzyme digestion preference to dot blots, have more recently been supplemented by ultrahigh performance liquid chromatography coupled with mass spectrometry (UHPLC-MS/MS) to accurately quantify specific DNA methylation. Methylated DNA can also be sequenced by either methylated DNA immunoprecipitation followed by sequencing (MeDIP-seq) or single-molecule real-time sequencing (SMRTseq) for identifying genomic locations of DNA methylation. Here we describe a protocol for the detection and quantification of epigenetic signaling DNA methylation modifications including, N6-methyladenine (6mA), N4-methylcytosine (4mC) and C5-methylcytosine (5mC) in genomic DNA by triple quadrupole liquid chromatography coupled with tandem mass spectrometry (QQQ-LC-MS/MS). The high sensitivity of the UHPLC-MS/MS methodology and the use of calibration standards of pure nucleosides allow for the accurate quantification of DNA methylation.

We develop a capillary-paper spray (CPS) ion source which allows for sample separation in the capillary and enables rapid and sensitive paper spray (PS) mass spectrometry (MS) analysis of biofluids. The CPS employs a glass capillary to load liquid analytes, vertically standing at the rear of the PS. To further reduce the matrix effect, a nitrocellulose filter membrane is placed between the glass tube and chromatography paper to absorb proteins and other macromolecules, which is beneficial for the detection of the small molecules. Compared with the normal PS method, the CPS method markedly improves spray stability and prolongs analysis duration, and also generates significantly better signal intensities during the analysis of drugs, thus indicating its potential for clinical use. As a proof of concept, quantitative analysis of drugs (metformin hydrochloride and berberine hydrochloride) in serum is performed.

We describe a method for the differentiation of carboxylate anions on disposable paper supports (common printer paper, filter paper, chromatography paper), based on differential patterns of interactions between carboxylates and a fluorescent sensing system. The sensor was built from commercially available components, namely a polycationic fifth generation amine-terminated poly(amidoamine) dendrimer (PAMAM G5) and a small organic fluorophore (calcein) through non-covalent interactions. The assay's physical dimensions were chosen to conform to the microwell plate standard so detection could be carried out on widely available plate reader instrumentation. The sensing complex was first deposited in spots on a paper support to prepare the sensor strip; a carboxylate solution was then loaded on each spot. Nuanced changes in fluorescence were associated with carboxylate binding to the PAMAM dendrimer, characteristic of the structure and affinity of each carboxylate. Such signal changes, interpreted through Linear Discriminant Analysis (LDA), contained enough information to recognize and successfully discriminate most anions in the panel. Among the substrates we tested, chromatography paper was the most promising. The relationship between the structure of the carboxylates and the patterns giving rise to their differentiation was also discussed. Finally, the long-term stability ("shelf life") of the pre-assembled [calcein·dendrimer] sensing system was found to be excellent when deposited on paper support.

Radioisotope indium-113m ( 113m In) with half-life, T 1/2 = 1.7 hours and gamma energy, Eg = 391 keV is suitable and meets the criteria as radiotracer in industry. 113m In radioisotope was obtained from tin-113 decay ( 113 Sn, T 1 / 2 = 115 days) of 112 Sn (n,g) 113 Sn neutron activation in nuclear reactor. The process of separation of radioisotope 113m In using column chromatography method based on silica gel matrix using 0.05 M HCI solution. Radionuclide and radiochemical purity tests were performed using the gamma-spectrometry method and paper chromatography. The final product specification in the form of 113m InCl 3 is clear solution, pH 2, obtained yield of 81.83%, radionuclide purity of 90.22%, radiochemical purity of 91.61 ± 0,29% and stable for 3 days at room temperature.

The purposes of this study were as follows: (1) to radiolabel ibandronic acid (IBA, a third-generation bisphosphonate) with 177 Lu, investigating optimal labeling conditions, and (2) to analyze biodistribution and imaging properties of intravenous 177 Lu-ibandronate (177 Lu-IBA) administered in animals. 177 Lu-labeled methylene diphosphonate (177 Lu-MDP) served as a comparator agent. Differing proportions of IBA solution and 177 LuCl3 solution were combined to determine an optimal ratio for radiolabeling purposes, varying pH, temperature, and time to establish ideal reactivity conditions. Radiochemical purity of the labeled compounds was then assessed by paper chromatography. In vitro and in vivo stabilities were also measured at specific time intervals. In Kunming mice, biodistributions of 177 Lu-IBA and 177 Lu-MDP and respective agent activities in various organs were monitored by gamma counter, and we performed single photon computed tomography/computed tomography (SPECT/CT) imaging of 177 Lu-IBA in normal New Zealand White rabbits. Radiolabeling yields for 177 Lu-IBA proved to be >97% within 30 minutes at 90°C, and its radiochemical purity ensured stability in vitro and in vivo. Furthermore, we found that 177 Lu-IBA is readily soluble in water, showing higher skeletal uptake than 177 Lu-MDP but lower uptake by liver and spleen. The image quality of 177 Lu-IBA was so clear that even after 6 days, analysis was still feasible.

A colorimetric probe N,N’-bis(2-methoxy-ethyl)-2,3,3-trimethyl-3H-squaraine (MOESQ) with H2O solubility was synthesized to detect Cu2+. MOESQ exhibits good selectivity, high sensitivity and fast UV-Vis response toward Cu2+ over other competing ions in CH3CN. The detection limit of MOESQ for Cu2+ in CH3CN can reach 1.88 × 10−7 molL−1. By adsorbing MOESQ on the chromatography paper, a colorimetric test paper for Cu2+ was prepared, which could detect Cu2+ with the color change from blue to faint yellow even in the limit of detection concentration of 10−6 molL−1.

ObjectiveTo evaluate the effectiveness of two kinds of Arg-Gly-Asp (RGD)-targeted131I-containing nanoliposomes for the treatment of cervical cancer in vitro and in vivo.MethodsThe nanoparticle liposomes designated RGD-131I-tyrosine peptide chain (TPC)-L and131I-RGD-L were prepared. The emulsion solvent evaporation method was used to encapsulate the polypeptide into liposomes. The quantity of entrapped polypeptide was measured using UV spectrophotometry. The labeling rates, radiochemical purities, and total radioactivities were measured using paper chromatography. Cytotoxicity was assessed using the MTS assay and flow cytometry. Therapeutic efficacy was monitored using a mouse xenograft model of cervical cancer.ResultsThe labeling efficiency, radiochemical purity, and specific radioactivity of RGD-131I-TPC-L were greater than those of131I-RGD-L. The cytotoxicity test indicated that late apoptosis of cells treated with RGD-131I-TPC-L and131I-RGD-L was higher than that of cells treated with Na131I. The therapeutic effect of RGD-131I-TPC-L was better than that of31I-RGD-L in the mouse model.ConclusionsThe specific activity of liposome-encapsulated RGD-131I-TPC-L was higher than that of131I-RGD-L, which labeled liposomes directly. Moreover, the RGD-131I-TPC-L liposomes were more effective for killing xenografted tumor cells.

The radiolabeled drug 99m Tc-tazobactam (99m Tc-TZB) was developed and assessed as an infection imaging agent in Pseudomonas aeruginosa and Salmonella enterica infection-induced animal models by comparing with inflammation induced animal models. Radiosynthesis of 99m Tc-TZB was assessed while changing ligand concentration, reducing agent concentration, pH, and reaction time while keeping radioactivity constant (~370 MBq). Percent labeling of the resulting complex was measured using paper chromatography and instant thin layer chromatography. The analysis of the 99m Tc-TZB complex indicated >95% labeling yield and electrophoresis revealed complex is neutral in nature. The biodistribution study also showed predominantly renal excretion; however liver, stomach, and intestine also showed slight tracer agent uptake. The agent significantly accumulated in Pseudomonas aeruginosa and Salmonella enterica infection induced tissues 3.58 ± 0.26% and 2.43 ± 0.42% respectively at 1 hour postinjection. The inflamed tissue failed to uptake noticeable activity at 1 hour time point. The scintigraphic study results were found in accordance with biodistribution pattern. On the basis of our preliminary results, the newly developed 99m Tc-TZB can be used to diagnose bacterial infection and to discriminate between infected and inflamed tissues.

Kanamycin is an antibiotic, isolated from Streptomyces kanamyceticus, which is used to treat serious bacterial infections. The fact that the present radioligand 99m Tc-kanamycin used for diagnosis is short-lived, raised a need to label and study kanamycin with one of the most important beta (β) radiation emitting isotope 177 Lu. Labeling yield of 177 Lu-kanamycin was confirmed by different chromatography techniques such as paper chromatography, TLC, HPLC. Several experiments were performed to optimize labeling with changing reaction conditions such as pH, temperature, amount of ligand, and reaction time. In vitro stability analysis was performed incubation with human serum. Electrophoresis analysis was also conducted to determine the charge on 177 Lu-kanamycin. The biodistribution and scintigraphy were performed in normal mice and rabbit, respectively, at different time intervals of postinjection. 177 Lu-kanamycin was prepared with very high yield (~100%), with excellent stability in vivo and in vitro (>99% 6 hr postprep.), at pH 7. Maximum labeling was achieved at less reaction time (15 min), with maximum conjugation of the ligand (12.5 mg) with 177 Lu. Electrophoresis analysis showed net neutral charge. The radioligand showed rapid clearance from body in biodistribution and scintigraphy studies. The preparation 177 Lu-kanamycin could be used as a radio-pharmaceutical for infection imaging purpose, especially when transporting the radioligand to long-range distances.

The present study aimed to develop a protocol for easy removal of β-ODAP neurotoxin by converting it into its isomer α-ODAP (reported to be less toxic) followed by its separation from the protein fraction in pH dependent manner. Use of β-mercaptoethanol prevented aggregate formation and increased solubility of the prepared Lathyrus sativus protein. Validation of ODAP removal by paper chromatography and mass spectrometry indicated the robustness of the protocol. Removal of ODAP and presence of high antioxidants and homoarginine content can enable Lathyrus sativus to be an alternate source of protein, as well as have other health benefits, including benefits for patients with cardiovascular diseases.

We report the synthesis and purification of a targeting probe for Medullary Thyroid Carcinoma (MTC) by incorporating 131I into PAMAM (G5.0) dendrimers. Both the 131I labeled control dendrimer (131I–PAMAM (G5.0) without attached targeting peptide) and the MTC-targeting dendrimer (131I–PAMAM (G5.0) attached to VTP (vascular targeting peptide)) were labeled with the radioisotope 131I by applying the iodogen method. The resulting G5.0 dendrimers were purified by means of ultracentrifugation. The labelling efficiencies and radiochemical purities vs. time were determined by paper chromatography. The radiolabeling efficiencies of 131I–PAMAM (G5.0) and 131I–PAMAM (G5.0) were 93 ± 1% and 85 ± 2%, respectively. 131I–PAMAM (G5.0) did exhibit small, but significant changes in radiochemical purity as a function of time after labelling. The highest observed highest purity was 82 ± 2%. 131I–PAMAM (G5.0)–VTP did display larger changes in radiochemical purity as a function of time after labelling, maximally 80 ± 2%. The stability of the two probes and their binding behavior to the human medullary thyroid cancer cell line (TT) were observed in vitro. Compared to the negative control group (consisting of Na131I), the TT cell binding rate of 131I–PAMAM (G5.0)–VTP was significantly increased at 48 h and 72 h (P 0.05). These findings have been confirmed by performing MTT assays. These results confirm earlier findings, which demonstrated fast uptake of PAMAM (G5.0) by various cell types.

Human biosynthetic insulin is a polypeptide hormone that plays an important and essential role in control of the level of carbohydrate, protein, and fat metabolism in the blood. Human pancreatic insulin was labeled with 99mTc to form a new radiopharmaceutical with a labeling yield of 99 ± 1% under optimum conditions: 0.1 mL insulin, pH 7, 25 μg stannous chloride, 1 mL (19 mCi) of pertechnetate, room temperature, and 10 min reaction time. The 99mTc–insulin complex was examined using paper chromatography, ITLC, electrophoresis, and HPLC. In addition, in vitro and in vivo study of 99mTc–insulin complex was performed at different time intervals.

Although there are many simple thin-layer chromatography (TLC) separations, many more are complex and involve more than a few components, that means having to use special high-performance TLC (HPTLC) plates or microspotting or banding devices to increase its resolving power if developing in only one direction. However, adding a second development to perform two-dimensional TLC (2D TLC) allows even better resolution of complex samples. This is because different modes of chromatography are being invoked by the use of one stationary phase with two mobile phases, bilayer plates, graft TLC, or multidimensional TLC. This paper is a review of recent applications that have benefitted from using 2D TLC in its various forms. They were chosen for their variety of sample types as well as the unique choices of plates and/or mobile phases made by the researchers to yield improved separations.

Noninvasive brain imaging is a process that allows scientists and physicians to view and monitor the areas of the brain. The aim of this study was to formulate a novel radiopharmaceutical for the detection of brain disorders at early stages in susceptible patients. (99m) Tc-ropinirole was prepared by the direct complexation of ropinirole with technetium-99m. The results showed that the radiochemical yield (99m) Tc-ropinirole was 92 ± 2.87% and the radiochemical yield was evaluated by paper chromatography and HPLC. In vitro studies showed that the formed complex was stable for up to 6 h. In vivo uptake of (99m) Tc-ropinirole in the brain was 4.87 ± 0.15% injected dose/g organ at 30 min post-injection, which cleared from the brain with time till it reaches 2.3% at 2 h post-injection indicating that the brain uptake of (99m) Tc-ropinirole is higher than that of the commercially available (99m) Tc-HMPAO, which is 2.25% at 30 min. Pre-dosing mice with cold ropinirole reduced the brain uptake to 0.26 ± 0.01% injected dose/g organ, so this confirms the high specificity and selectivity of this radiotracer for the assessment of the dopamine receptors.

High-performance affinity chromatography is limited by its high cost and high pressure. Paper is made up of porous fiber networks and has the properties of low cost, ease of fabrication, and biodegradable. Due to these advantages, herein, we immobilized beta2-adrenoceptor (β2-AR) onto the surface of the polytetrafluoroethylene membrane, a paper-based material, and constructed a G protein-coupled receptor (GPCR)-in-paper chromatographic platform. This platform was characterized by Fourier transform infrared spectroscopy, fluorescence analysis, X-ray photoelectron spectroscopy, and chromatographic studies. These morphological and elemental analysis showed that β2-AR was successfully immobilized on the paper surface. The specific drugs have good retentions on the GPCR-in-paper chromatographic platform. The association constants of salbutamol, terbutaline and bambuterol to β2-AR were calculated to be 2.02 × 104 M−1, 1.15 × 104 M−1, 1.75 × 104 M−1 by adsorption energy distribution, which were in good line with the values from frontal analysis, zonal elution and previous literatures. We demonstrated that the GPCR-in-paper platform was cost-effective, easy to be modified for protein immobilization, and applicable in the receptor-drug interaction analysis. We believe such a platform sheds new light on paper chromatography for receptor-drug interaction analysis and other applications.

Herein, we report three-dimensional paper chromatography (3D-PC) as a micro-chromatographic platform. The method was based on applying the origami microfluidic device for separation, coupled by colorimetric methods for simultaneous determination. The microfluidic device fabrication was a facile printing approach. Two azo food dyes, Tartrazine (E102) and Indigo carmine (E132), were selected as a model analyte, while carbonate-bicarbonate buffer was used as the mobile phase. Our micro-chromatographic device is associated with two big advantages including needing very small volume of mobile phase ( ~12 µL) and ultrafast separation time (~35 s). Under the optimal conditions, the method provided acceptable linear ranges of 0. 0 g L 1–18.0 g L 1 (R2 = 0.997) for E102 and 0.070 g L 1–10.0 g L 1 for E132 and the limits of detection (3σ/slope) were evaluated as 0.620 and 0.060 g L 1, respectively. The proposed method was successfully applied in the separation and quantification of these dyes in commercial food products such as jelly, candy, and four kinds of drink samples without any sample preparation prior to analysis. The mean recovery values for the real sample analysis were in the range of 100.14%–102.38% for E132 and E102 respectively. The inter-device relative standard deviations were in the ranges of 1.5%–11.8%. In total, our chromatographic μPAD is small (1.0 cm × 1.0 cm × 0.5 cm), portable, inexpensive, no need of specialized user, requires low volumes of sample (0.5 μL), and can perform separation using 12 μL of aqueous mobile phase in very short time.

Breath analysis is considered to be an effective method for point-of-care diagnosis due to its noninvasiveness, quickness and simplicity. Gas sensors for breath analysis require detection of low-concentration substances. In this paper, we propose that reduction of the background current improves the limit of detection of enzymatic biogas sensors utilizing chromatography paper. After clarifying the cause of the background current, we reduced the background current by improving the fabrication process of the sensors utilizing paper. Finally, we evaluated the limit of detection of the sensor with the sample vapor of ethanol gas. The experiment showed about a 50% reduction of the limit of detection compared to previously-reported sensor. This result presents the possibility of the sensor being applied in diagnosis, such as for diabetes, by further lowering the limit of detection.

β-1,3:1,4-Glucan is a major cell wall component accumulating in endosperm and young tissues in grasses. The mixed linkage glucan is a linear polysaccharide mainly consisting of cellotriosyl and cellotetraosyl units linked through single β-1,3-glucosidic linkages, but it also contains minor structures such as cellobiosyl units. In this study, we examined the action of an endo-β-1,3(4)-glucanase from Trichoderma sp. on a minor structure in barley β-1,3:1,4-glucan. To find the minor structure on which the endo-β-1,3(4)-glucanase acts, we prepared oligosaccharides from barley β-1,3:1,4-glucan by endo-β-1,4-glucanase digestion followed by purification by gel permeation and paper chromatography. The endo-β-1,3(4)-glucanase appeared to hydrolyze an oligosaccharide with degree of polymerization 5, designated C5-b. Based on matrix-assisted laser desorption/ionization (MALDI) time-of-flight (ToF)/ToF-mass spectrometry (MS)/MS analysis, C5-b was identified as β-Glc-1,3-β-Glc-1,4-β-Glc-1,3-β-Glc-1,4-Glc including a cellobiosyl unit. The results indicate that a type of endo-β-1,3(4)-glucanase acts on the cellobiosyl units of barley β-1,3:1,4-glucan in an endo-manner.

Knowledge of the radiochemical purity of radiopharmaceuticals is mandatory and can be evaluated by several methods and techniques. Planar chromatography is the technique normally employed in nuclear medicine since it is simple, rapid and usually of low cost. There is no standard system for the chromatographic technique, but price, separation efficiency and short time for execution must be considered. We have studied an alternative system using common chromatographic stationary phase and alcohol or alcohol:chloroform mixtures as the mobile phase, using the lipophilic radiopharmaceutical [(99m)Tc(MIBI)₆]⁺ as a model. Whatman 1 modified phase paper and absolute ethanol, Whatman 1 paper and methanol:chloroform (25:75), Whatman 3MM paper and ethanol:chloroform (25:75), and the more expensive ITLC-SG and 1-propanol:chloroform (10:90) were suitable systems for the direct determination of radiochemical purity of [(99m)Tc(MIBI)₆]⁺ since impurities such as (99m)Tc-reduced-hydrolyzed (RH), (99m)TcO(4)(-) and [(99m)Tc(cysteine)₂]⁻ complex were completely separated from the radiopharmaceutical, which moved toward the front of chromatographic systems while impurities were retained at the origin. The time required for analysis was 4 to 15 min, which is appropriate for nuclear medicine routines.

A facile and simple paper-based scanometric assay was developed to detect Pb(2+) using GR5-DNAzyme. Magnetic beads (MBs) and gold nanoparticles (AuNPs) were used as a signal collector and a signal indicator, respectively. They were linked together by GR5-DNAzyme, comprising an enzyme and a substrate strand pairing up with each other. In the presence of Pb(2+), the substrate strand is cut into two pieces, resulting in the disassembly of AuNPs from the MBs. These AuNPs were spotted on predefined areas on a chromatography paper, where signal is amplified through silver reduction. This sensing platform exhibits high sensitivity and selectivity toward Pb(2+), giving a detection limit of 0.3 nM and a linear fitting range from 0.1 to 1000 nM. Testing of this biosensor in river water and synthetic urine samples also showed satisfying results. Besides offering simultaneous and multi-sample analysis, this paper-based sensing platform presented here could be potentially applied and served as a general platform for on-site, naked eyes, and low-cost monitoring of other heavy metal ions in environmental and body fluid samples.

We report on the development of an electrochemical probe for the trace analysis of 2,4,6-trinitrotoluene (TNT) in soil samples. The probe is a combination of graphite electrodes, filter paper, with ethylene glycol and choline chloride as the solvent/electrolyte. Square wave chromatovoltammograms show the probes have a sensitivity for TNT of 0.75 nA/ng and a limit of detection of 100 ng. In addition, by taking advantage of the inherent paper chromatography step, TNT can be separated in both time and cathodic peak potential from 4-amino-dinitrotolene co-spotted on the probe or in soil samples with the presence of methyl parathion as a possible interferent.

A simple spectrophotometric method has been developed and validated for quantitative determination of Clarithromycin (CLT) in pharmaceutical formulations. The method is based on derivatization of CLT with Bromotrimethylsilane (TMBS) and spectrophotometric estimation of CLT derivative at 268 nm using double beam spectrophotometer and quartz cells of 1 cm path length. Bear law obeyed over a concentration range of 5.0 to 60 μg mL-1 with specific absorbance (A=1%) of Clarithromycin derivative was 62.4 ± 0.2.0. The method was validated appropriately for accuracy, precision, linearity and specificity, according to the requirements of United State Pharmacopeia and International Conference on Harmonization guidelines for validation of analytical method (category I). The overall recovery of CLT was 100% ± 1% and RSD for precision and intra-day precision was less than 2.0%. The calibration curves were linear with correlation coefficient (R2>0.9998) and there was no interference of placebo of clarithromycin dosage form. The method was successfully used for determination of CLT in dosage form and in vitro dissolution studies.

Sclerotium rolfsii is a ubiquitous soil-borne fungal pathogen known to cause disease on worldwide range of agricultural and horticultural crops. In spite of economic loss caused by this pathogen, very few reports were available on this aspect, biochemical characterization of the S. rolfsii; hence the present study was under taken to study the biochemical characterization of S. rolfsii. Qualitative and quantitative analysis of free and protein-bound amino acids was carried out in mycelial mats and culture filtrate of S. rolfsii. Five amino acids were identified and were common both in free and protein-bound amino acids. Methionine was the most abundantly occurring amino acid in the protein-bound amino acids, whereas asparagine in case of free amino acids. Eleven amino acids were detected in the culture filtrate, but methionine, phenylalanine and leucine/isoleucine were more abundant. Phenolic acids both in culture filtrate and mycelia mat were analyzed by 2-dimensional paper chromatography; seven were detected both in culture filtrate and mycelial mat. Gallic, ferulic, chlorogenic and cinnamic acids were common both in mycelium and culture filtrate of the S. rolfsii. The other biochemical constituents like carbohydrate fractions, phenolic compounds and nitrogen fractions present in both mycelia mat and culture filtrate were also studied. Key words: Sclerotium rolfsii, stem rot, biochemical constituents, amino acids, phenolic acids.

Telomerase is extensively expressed in various cancer cells and recognized as a target for cancer drug discovery. In the present study, a simple and amplification-free fluorescence assay based on polyamidoamine starburst dendrimer (PAMAM dendrimer)-activated paper device is proposed for sensitive detection of telomerase activity through hybridization of Cy5 modified single strand DNA probes with telomerase extension products. The paper substrate is fabricated by hand drawing according to a template, which is low cost, instrument free and easy operation. PAMAM is rich in amino groups on its surface and employed to immobilize the telomerase substrate (TS) primer. Highly sensitive detection of telomerase activity in HeLa cell lysate of 10 cells is achieved since the PAMAM dendrimer-activated paper surface can provide high density of binding sites for immobilization of TS primer. The experimental results also demonstrate that the assay can be employed to evaluate telomerase activity levels of various cell lines and screen telomerase inhibitors.

This paper presents a novel method of fabricating an enzymatic biosensor for breath analysis using chromatography paper as enzyme supporting layer and a liquid phase layer on top of screen printed carbon electrodes. We evaluated the performance with ethanol vapor being one of the breathing ingredients. The experimental results show that our sensor is able to measure the concentration of ethanol vapor within the range of 50 to 500 ppm. These results suggest the ability of detecting breath ethanol, and it can possibly be applied as a generic vapor biosensor to a wide range of diseases.

The materials, techniques, and instruments for the detection and quantification of radiolabeled compounds by planar radiochromatography are described. Included is sample preparation; separations on Instant Thin Layer Chromatography sheets, thin layer plates, and chromatography papers; and detection and quantification by film autoradiography, liquid scintillation counting, storage phosphor imaging, and in situ radioactivity scanning. These methods are widely applied in radiopharmaceutical purity determination, metabolism studies, and many other types of chemical, biochemical, and biological investigations.

Bacterial infection is found to be the cause of death throughout the world. Nuclear medicine imaging with the help of radiopharmaceuticals has great potential for treating infections. In the present work, clindamycin, a lincosamide antibiotic, was labeled with technetium-99 m (~380 MBq). Clindamycin has been proven to be efficient for treating serious infections caused by bacteria such as Staphylococcus aureus. Quality control, characterization, biodistribution, and scintigraphy of radiolabeled clindamycin were done, and labeling efficiency was determined by ascending paper chromatography. More than 95 % labeling efficiency with technetium-99 m ((99m)Tc) was achieved at pH 6-7 while using 2.5-3 μg SnCl2 · H2O as a reducing agent and 100 μg of ligand at room temperature. The characterization of the compound was performed by using electrophoresis, HPLC and shake flask assay. Electrophoresis indicates the neutral behavior of (99m)Tc-clindamycin. HPLC analysis confirms the single specie of the labeled compound, while shake flask assay confirms high lipophilicity. The biodistribution studies of (99m)Tc-clindamycin were performed Sprague Dawley rats bearing bacterial infection. Scintigraphy and biodistribution studies showed a high uptake of (99m)Tc-clindamycin in the liver, heart, lung, and stomach as well as at S. aureus-infected sites in rabbits.

This Letter aimed to develop an efficient method for the determination of cyanide ion (CN − ). A novel colorimetric chemosensor 4-[(1 E )-2-(4-hydroxyphenyl)ethenyl]-1-allylpyridinium bromide (HPEAPB) was synthesized. HPEAPB displayed good selectivity toward CN − over other competing anions in ethanol. A color change from yellow to red was immediately observed upon the addition of CN − and the limit of detection (LOD) was 3.4 × 10 −6 mol L −1 . The sensing mechanism was discussed by UV–vis, 1 H NMR titration, and a comparison study. Colorimetric test paper for CN − was prepared by attaching HPEAPB to a chromatography paper, which could be used to detect CN − in environmental samples as simply as a pH-indicator paper for pH value. The LOD of the test paper for CN − was 2.0 × 10 −4 mol L −1 . This detection method for CN − has potential applications in cyanide ion containing fields by combination of rapid and real-time advantages.

A putative α-amylase gene (accession number, CP000284) of Methylobacillus flagellatus KT ATCC51484 was cloned in Escherichia coli, and its gene product was expressed and characterized. The purified recombinant enzyme (MFAS) displayed a typical amylosucrase (ASase) activity by the demonstration of multiple activities of hydrolysis, isomerization, and polymerization although it was designated as an α-amylase. The optimal reaction temperature and pH for the sucrose hydrolysis activity of MFAS were determined to be 45 °C and pH 8.5, respectively. MFAS has relatively high thermostable characteristics compared with other ASases, as demonstrated by a half-life of 19.3 min at 50 °C. MFAS also showed polymerization activity using sucrose as a sole substrate. Glycerol was transglycosylated by the intermolecular transglycosylation activity of MFAS. Two major products were observed by thin-layer chromatography and isolated by paper chromatography and recycling HPLC. Using (1)H and (13)C NMR, their chemical structures were determined to be (2S)-1-O-α-D-glucosyl-glycerol or (2R)-1-O-α-D-glucosyl-glycerol and 2-O-α-D-glucosyl-glycerol, in which a glucose molecule is linked to glycerol via an α-glycosidic linkage.

The anthocyanins in the brilliant crimson petals from Erythrina crista-galli L. were extracted with 10% AcOH, and the extract was purified by Amberlite XAD-7 column chromatography, paper chromatography, and preparative HPLC. The structure of a major anthocyanin was confirmed as cyanidin 3- O -sophoroside by 1 H and 13 C NMR and FABMS. Then, the catalysis of UDP-glucose: flavonoid 3- O -glucoside glucosyltransferase (3GGT) was predicted at the final step in the anthocyanin biosynthesis. In this study, the 3GGT was purified approximately to 800-fold from E. crista-galli petals by eight-steps purification procedures. The purified enzyme (Ec3GGT) catalyzed only the 2″- O -glucosylation of flavonoid 3- O -glucoside to produce flavonoid 3- O -sophoroside. Ec3GGT showed a pH optimum around 7.5. The molecular weight and isoelectric point were estimated to be about 82 kDa and 6.5, respectively; the molecular weight was larger than known UDP-sugar: flavonoid glycosyltransferases (UFGTs). The enzyme did not require metal cofactors but inhibited by Zn 2+ , Co 2+ and Cu 2+ , and it activity was inhibited by SH-blocking reagents such as NEM and PCMB. Both anthocyanidin 3- O -glucosides and flavonol 3- O -glucosides were favorable acceptor substrates of the enzyme, whereas no activity was shown toward C -glucosylflavones such as isovitexin, which is a minor component in the petals. These results showed that Ec3GGT involved in the last step of biosynthesis of a major anthocyanin in petals.

Labelling of 5-aminolevulinic acid (5-ALA) with 99mTc was achieved by using SnCl2·2H2O as reducing agent. Radiochemical purity and labelling efficiency was determined by instant thin layer chromatography/paper chromatography. Efficiency of labelling was dependent on many parameters such as amount of ligand, reducing agent, pH, and time of incubation. 99mTc labelled 5-ALA remained stable for 24 h in human serum. Tissue biodistribution of 99mTc-5-ALA was evaluated in Sprague–Dawley rats. Biodistribution study (% ID/g) in rats revealed that 99mTc-5-ALA was accumulated significantly in liver, spleen, stomach and intestine after half hour, 4 and 24 h. Significant activity was noted in bladder and urine at 4 h. High liver uptake of 99mTc-5-ALA makes it a promising liver imaging agent.

Radioactivity occurs in trithloroacetic acid (TCA)-soluble and precipitable, cytoplasm and salt-washed walls following germination of Lilium longiflorum, cv. 'Ace' pollen in medium containing [14C]-proline (Pro). Sephadex gel filtration on G-25 through G-100 was employed to determine whether radioactivity in cytoplasm, wall and growth medium from pollen fed [14C]-Pro or [3H]=Pro plus [14C]-arafbinose (Ara) was contained within molecules possessing molecular weights of 5,000 to 100,000 daltones or greater. G-25 elution profiles of a crude cytoplasmic fraction (15,000 X g supernatant) from [14C]-Pro labelled pollen yielded a radioctive void volume peak and a retarded peak. The void volume peak contained hydroxyproline (Hyp), and exhibited a coincidence of [3H]-Pro and [14C] -Ara labelling when pollen was double labelled with the two isotopes. This peak also contained radioactivity when pollen was germinated in 2-[3H]-myo-inositol. Germination in medium supplemented with 100 µM 2,2'-dipyridyl eliminated radioactivity from 2-[3H]-myo-inositol or [14C]-,Pro in the peak. Filtratian on G-25 of aTCA-soluble fraction of a salt-extract of walls from [14C]-Pro labelled pollen resulted in void volume and two retarded peaks. Void volume and two retarded peaks were also obtained upon G-25 filtration of a cellulase-digest of walls from [M]-Pro labeled pollen. The void volume peak contained Hyp, Lys, Gly, Ala, Ser, Glu and Asp acids, Val, Tyr, Leu or lieu and Pro. Sephadex G-90, 75, and 100 elution profiles of cellulasedigests of walls from [3H]-,Pro and [14C]-Ara labelled pollen yielded radioactive retarded and Hyp-containing void volume peaks with a coincidence of [3H] and [14C] labelling. Label in the void volume was obtained when either rhozyme P11- or pepsin-digests of walls from [14C]-Pro labelled pollen were gel filtered on G-50. Paper electrophoresis coupled with paper chromatography of acid hydrolyzates of salt-washed wall fractions demonstrated 15 of the common amino acids. Gel filtration on G-25 of growth medium in which pollen was germinated resulted in two peaks, one of which eluted in the void volume. contained Hyp and excluded during subsequent gel filtration on G-100.

Microcircular planar chromatography (μ-PLC) is possible without any mobile phase tank. It also offers a way to use a quite strong room temperature air or gas flushing mode to remove carefully and completely cleaning liquids, sample solvent traces, focusing liquids from the stationary phase in order to let PLC run under fully controlled mobile and stationary phase conditions. This mode of circular PLC opened a simple way to introduce a third chromatography phase, which is gaseous and can be loaded with selectivity changing substances like acids, bases, or volatile organic substances of any polarity. All phase flow is constant per time. This offers accuracy and precision of analytical data unknown in thin-layer chromatography up to now. The Three Phase chromatography works already with a rather simple instrumentation — comparable to the instrumental mode of μ-PLC [1].

The research on protein deimination and the enzymes responsible for catalyzing this posttranslational modification started from investigations of the hair follicle and myelin in central nerve system. About 60 years ago, in 1958, the presence of protein-bound citrulline was first reported by Dr. George Rogers (Adelaide University, Australia) in the protein of the inner root sheath (IRS) of hair follicles (Rogers 1958). In order to obtain information about the protein composition of the IRS, he conducted a quantitative amino acid analysis of an acid hydrolysate on sufficient amounts of IRS that were dissected from the vibrissae follicle of rats. At that time, the common method for separating amino acids was paper chromatography, and when applied to the IRS hydrolysates, citrulline was discovered as a distinct ninhydrin-positive spot in an area adjacent to the basic amino acids. About 10 years after Rogers’s discovery, Dr. Mario Moscarello (Toronto University, Canada) started an intensive investigation of myelin sheath proteins in the central nerve system. In 1971, he also found the presence of peptide-bound citrulline in myelin basic protein (MBP) using similar methods to Rogers (Finch et al. 1971). Moscarello continued the investigation of MBP until he passed away in 2013, publishing many papers concerning the hyper-deimination of MBP in the pathology of multiple sclerosis. His research career involving the deimination of MBP was described in a eulogy in the first volume of this book series (Nicholas and Bhattacharya 2014).

Fred was still working with active centers of enzymes and with proteins in general. He liked the visual methods of paper chromatography and paper electrophoresis, and he held a certain disdain for work on columns. Protein sequencing, however, had largely gone to column methods because they were more quantitative.

Fusarium solaniThom produced maximum PHB depolymerase by 48 h when grown in BHM containing 0.2%, w/v PHB, pH 8.0 at25°C. Statistical optimization studies using Plackett Burman design of PHB depolymerase production yielded maximum PHB depolymerase activity after 2 days as against 4 days in the unoptimized conditions with a 2-fold increase in activity. Partial purification of the extracellular poly-β-hydroxybutyrate (PHB) depolymerase PHAZFusfromF. solaniThom by two steps using ammonium sulphate (80% saturation) and affinity chromatography using concanavalin-A yielded 162.3-fold purity and 63% recovery of protein. The enzyme composed of a single polypeptide chain of 85 KDa, as determined by SDS-PAGE. The enzyme stained positive for glycoprotein by PAS staining. Optimum enzyme activity was detected at pH 7.0 and55°C. The enzyme was stable at pH 7.0 and 55°C for 24 h with a residual activity of almost 85%. Paper chromatography revealedβ-hydroxybutyrate monomer as the major end product of PHB hydrolysis. Complete inhibition of the enzyme by 1 mM HgCl2(100%) indicated the importance of essential disulfide bonds (cystine residues) for enzyme activity or probably for maintaining the native enzyme structure.

Choline acetyltransferase (ChAT) catalyzes the reaction between choline and acetylcoenzyme A (AcCoA) to form acetylcholine (ACh) in nerve terminals. ACh metabolism has implications in numerous aspects of physiology and varied disease states, such as Alzheimer's disease. Therefore a specific, sensitive, and reliable method for detecting ChAT enzyme activity is of great utility in a number of situations. Using an existing radionuclide-based enzyme activity assay, we have observed detectable ChAT signals from non-cholinergic cells, suggesting a contaminant in the assay producing an artifactual signal. Previous reports have suggested that L-acetylcarnitine (LAC) contaminates many assays of ChAT activity, because of difficulties in separating LAC from ACh by organic extraction. To determine the source of this hypothesized artifact and to rectify the problem, we have developed a paper chromatography-based assay for the detection of acetylcholine and other contaminating reaction products of this assay, including LAC. Our first goal was to develop a simple and economical method for resolving and verifying the identities of various reaction products or contaminants that could be performed in most laboratories without specialized equipment. Our second goal was to apply this separation method in postmortem human brain tissue samples. Our assay successfully detected several contaminants, especially in assays using brain tissue, and allowed the separation of the intended ACh product from these contaminants. We further demonstrate that this assay can be used to measure carnitine acetyltransferase (CrAT) activity in the same samples, and assays comparing ChAT and CrAT show that CrAT is highly active in neuronal tissues and in neuronal cell cultures relative to ChAT. Thus, the simple chromatography-based assay we describe allows the measurement of specific reaction products separated from contaminants using commonly available and inexpensive materials. Further, we show that ChAT activity is significantly reduced in brain extracts from Alzheimer's disease compared to controls.

The lactic acid bacteria to produce GABA and GABA determinations method were studied. Research results show that the strains isolated belong to Lactobacillus delbruckii for GABA with morphological and physiological and biochemical characterization. Paper chromatography for determination of GABA, which suitable conditions for developing agent (N-butanol: acetic acid: water) volume ratio were 4: 1: 3, under 30 min, 80 °C, 510nm detection wavelength respectively.

Momardica charantia L.var. Jhalri (MCJ) is the member of lectins from the family Cucurbitaceae. For further characterization MCJ lectin was analyzed for amino acid identification and sugar analysis by paper chromatography in the present paper. The technique reported the presence of total 18 amino acids with large content of Serine and Glutamic acid. The same technique revealed the presence of glucose, mannose and hexosamine. The total hexose sugar by Orcinol- sulphuric acid method and total hexosamine by Elson and Moragn method was estimated to be 6.81mg and 5.037mg respectively per 100mg of MCJ lectin. The good content of carbohydrate in MCJ lectin being glycoprotein in nature was considered as a major and crucial factor for apparent high mol. Wt. of MCJ lectin.