Ascending Paper Chromatography Research Articles

Novel design of low volume mobile phase paper chromatography

A novel design and set up of paper chromatography in the form of radial serpentine arms laying horizontally is proposed. The consumption of mobile phase is reduced to be 1 mL or less as compared to the tens of mL typically used in the conventional set ups for ascending paper chromatography, without any change in the order of the separated components. Greenness measurement based on the ChlorTox value is approximately 25, when comparing to a conventional system that consumes 25 mL of mobile phase. A digital image of the chemical separation makes data analysis more feasible as the image can be kept, and hence, extending the time for data analysis. The Retention Factor (Rf) value from a serpentine shaped stationary phase can be calculated based on measurement of linear distances of the solvent front and those travelled by each component, and converting them to arclengths using the mathematical equation of a catenary wave that produces that particular design. Proof of concept is demonstrated by the separation of food dyes using a two solvent system, where a nonvolatile salt solution is followed by a volatile organic solvent mixture of alcohols and base. Relationships between concentration, color intensity (RGB values), and zone arclength are also demonstrated which indicate possibility of quantitative study on the serpentine design using digital image analysis.

Анализ аминокислотного и элементного состава листьев омелы белой

Objective: comparative study of the amino acid and elemental composition of leaves Viscum album L. and host plants. Materials and methods. The amino acid and elemental composition in raw materials and extracts from leaves Viscum album and the corresponding host plant, obtained by extraction with 50% ethyl alcohol, was determined by capillary electrophoresis. Results. Glycine, alanine, leucine, proline, methionine, valine, asparagine, aspartic and glutamic acids were previously identified in the leaves Viscum album by ascending paper chromatography. By capillary electrophoresis, it was found that the raw materials and extracts contained proline, valine, alanine, threonine and methionine in the greatest amount. It has been established that in a semi-parasitic plant, the content of proline is 16 times higher than in Malus domestica Borkh. leaves and 5 times higher than in Pyrus communis L. leaves In alcohol-water extracts, the proline content is 5 and 1,5 times higher, respectively. It should be noted that in raw materials and extracts from Viscum album, obtained by extraction with 50% ethyl alcohol, potassium ions predominates quantitatively. Compared to leaves Malus domestica, leaves Viscum album accumulate 4,5 times more potassium, and 1,5 times more than in Pyrus communis leaves; in extracts, the content is also higher by 3 and 2 times, respectively. Conclusion. In raw materials and extracts from Viscum album, 13 free amino acids were found, of which 6 are essential (Arg, Phe, Leu, Met, Thr, Val). It was found that potassium predominates quantitatively in raw materials and extracts from Viscum album.

Detection of artificial colors added to cooked faba beans (Ful medames) and microbial examination for samples obtained from street vendors in Egypt

BackgroundCooked dried beans are commonly consumed in many countries and are known in Egypt as Ful medames. Adding synthetic colors to keep the dishes more attractive may have toxic effects on the human body, as it may be unsafe to consume potentially microbial contaminated foods, especially when obtained from street vendors.AimThe aim of this study was to detect the types of artificial colors added to cooked faba bean (Ful medames) by street vendors in Egypt and highlight their health hazards and microbial hazards.Materials and methodsA total of 100 convincing samples were collected from street vendors from four large Egyptian cities. The coloring agents were extracted and purified from samples using the hydrochloric acid extraction method followed by separation using ascending paper chromatography. Total bacterial counts, coliforms, Bacillus cereus, Staphylococcus aureus, and mold and yeast counts were used for microbial tests on 24 samples obtained from Cairo and Giza cities.FindingsArtificial colors were detected in 46% of samples. From the total positive samples, sunset yellow, tartrazine, or both combined were identified in 26.1, 30.4, and 41.3%, respectively. Carmoisine was detected combined with tartrazine in only one sample. Giza and Cairo cities recorded the highest positive samples (64.3 and 42.9%, respectively), followed by Mansoura (10%). No added colors were detected in Mahala samples. Coliforms, mold, yeast, and bacillus were detected in most of the samples that were subjected to microbiological examination, whereas S. aureus was not found in any of the examined samples.ConclusionOverall, 46% of the examined samples from street vendors contained artificial colors, mainly tartrazine and sunset yellow, although the authorities of Egyptian standards do not allow any artificial additives to these types of food. Most positive samples were from more urban cities. Microbial contaminants found in the examined samples may be due to unhygienic approaches in such areas and lack of sanitary practices of street vendors. This also predisposes consumers to the risk of severe public health challenges.RecommendationsThe society should be made aware about the health hazards of the incessant usage of synthetic food colors. The public should be encouraged to add natural food coloring, such as carrots, through home cooking instead. Moreover, strict monitoring of street vendors for food safety regulation should be done by competent authorities.

Detecting Organic Materials in Fly Ash using Chromatographic Method

Abstract: Fly ash, a byproduct of coal-fired power plants, is entirely inorganic and inert, making it unfit for life. In Korba, fly ash is deposited in dykes, which are open embankments. The quest for organic compounds was critical because the humus formation is a revolution in fly ash, and just it will be capable of supporting life forms. This was conducted in the dykes to verify the organic materials source in fly ash, namely, whether they are originated from coal, furnace start-up oil, or animals and plants that grew in the area. Thin layer chromatography and ascending paper chromatography in a liquid medium were used as the method.

Method for Determining the Radiochemical Purity of Radiopharmaceuticals Based on Iodine-125, Iodine-131 and Samarium-153-oxabifor

iopharmaceuticals used in medicine for diagnostics, treatment, and anesthesia in various diseases. Radionuclide 125I was obtained by irradiation with neutrons of the enriched isotope 124Хе in a nuclear reactor. 131I was isolated from irradiated samples of TeO2 enriched in 130Те, and radionuclide 153Sm was obtained by irradiation of samples 152Sm2O3 in a nuclear reactor. The radiochemical purity of sodium iodide preparations labeled with 125I or 131I radionuclides was determined by the electrophoretic method, and the samarium-153-oxabifor preparation was determined by ascending paper chromatography. To determine the distribution of the activity of 125I, 131I, and 153Sm-oxabiphor by electrophoregrams and chromatograms, a γ-radiometric device Ludlum-2200 was used, which makes it possible to determine the radiochemical purity at the level of 99.9%.

Labeled of Irradiated Chitosan with IODIUM-131 Radioisotope

Chitosan is a polysaccharide from the deacetylation process of chitin, a compound derived from crustacean animal exoskeletons. Chitosan has benefits in various fields, one of them is in health field. Chitosan is known to be able to bind with fat so that it can reduce cholesterol levels in the blood, besides chitosan also has activities as an antibacterial and anticancer. Irradiated chitosan is carried out to degrade chitosan to smaller molecular weight and is easily absorbed to the body. As a new drug, many stages of testing are carried out to determine the mechanism and effectiveness of chitosan compound specifically as an anticancer compound. One of the required data is preclinical study data using animal model. Nuclear techniques offers the results of preclinical study that can identify quickly and accurately the distribution of a drug in the body with the radiotracer principle. Therefore it is necessary to make T-chitosan labeled compounds irradiation (131I-chitosan-75kGy) and compared with 131I-chitosan labeled compounds non-radiation (131I-chitosan) as controls. In this study, the optimum formulation of chitosan and chitosan-75kGy with radioisotope iodine-131 with radiochemical purity > 90% was obtained. Chitosan can be labeled by iodine-131 with the formulation of 150 μL (0.5%) chitosan, 20 μl (5 mg / mL) chloramine T, 20 μl (10 mg / mL) sodium metabisulphite, 10 μL Nal3I and 5 minutes incubation time at room temperature. While the optimum formulation of chitosan-75kGy labeled is 100 μL (0.5%) chitosan-75kGy, 10 μl (5 mg / mL) chloramine T, 10 μl (10 mg / mL) sodium metabisulphite, 10 μL Na131I with 1 minute incubation time at room temperature. Radiochemical purity testing was carried out using ascending paper chromatography method with whatman 1 paper as a stationary phase. To separate impurities in the form of I2, chloroform is used as the mobile phase, whereas methanol 90% is used to separate impurities in the form of I−. With this successful in producing labeled chitosan, it is expected that the effectiveness of irradiated chitosan as anticancer compounds can be identified.

Discovery of the canonical Calvin-Benson cycle.

It has been 65years since the Calvin-Benson cycle was first formulated. In this paper, the development of the concepts that are critical to the cycle is traced and the contributions of Calvin, Benson, and Bassham are discussed. Some simplified views often found in text books such as ascending paper chromatography and the use of the "lollipop" for short labeling are discussed and further details given. Key discoveries that underpinned elucidation of the cycle such as the importance of sedoheptulose phosphate and ribulose 1,5-bisphosphate are described. The interchange of ideas between other researchers working on what is now called the pentose phosphate pathway and the development of the ideas of Calvin and Benson are explored while the gluconeogenic aspects of the cycle are emphasized. Concerns raised about anomalies of label distribution in glucose are considered. Other carbon metabolism pathways associated with the Calvin-Benson cycle are also described. Finally, there is a section describing the rift between Calvin and Benson.

131I-trazodone: preparation, quality control and in vivo biodistribution study by intranasal and intravenous routes as a hopeful brain imaging radiopharmaceutical

ObjectivesThe preparation of 131I-trazodone hydrochloride and its biological evaluation as a promising brain imaging radiopharmaceutical using two routes of administration. Material and methodsTrazodone (TZ) was radiolabelled with 131I using direct electrophilic substitution, and different factors affecting labelling yield were studied. Quality control of 131I-TZ was carried out using ascending paper chromatography, paper electrophoresis, and high pressure liquid chromatography (HPLC). In vivo biodistribution of 131I-TZ was evaluated in Swiss albino mice using 3 methods: intravenous 131I-TZ solution (IVS), intranasal 131I-TZ solution (INS), and intranasal 131I-TZ microemulsion (INME). ResultsOptimum labelling yield of 91.23±2.12% was obtained with in vitro stability of 131I-TZ up to 6h at room temperature. The biodistribution results showed a notably higher and sustained brain uptake for INME compared to IVS and INS at all time intervals. In addition, heart and blood uptake levels for INME were lower than those for IV solution which, in turn, could decrease the systemic side effects of trazodone. Also, the 131I-trazodone INME brain uptake of 6.7±0.5%ID/g was higher than that of 99mTc-ECD and 99mTc-HMPAO (radiopharmaceuticals currently used for brain imaging). Conclusion131/123I-trazodone formulated as INME could be used as a promising radiopharmaceutical for brain imaging.

131I-trazodone: preparation, quality control and in vivo biodistribution study by intranasal and intravenous routes as a hopeful brain imaging radiopharmaceutical.

The preparation of 131I-trazodone hydrochloride and its biological evaluation as a promising brain imaging radiopharmaceutical using two routes of administration. Trazodone (TZ) was radiolabelled with 131I using direct electrophilic substitution, and different factors affecting labelling yield were studied. Quality control of 131I-TZ was carried out using ascending paper chromatography, paper electrophoresis, and high pressure liquid chromatography (HPLC). In vivo biodistribution of 131I-TZ was evaluated in Swiss albino mice using 3 methods: intravenous 131I-TZ solution (IVS), intranasal 131I-TZ solution (INS), and intranasal 131I-TZ microemulsion (INME). Optimum labelling yield of 91.23±2.12% was obtained with in vitro stability of 131I-TZ up to 6h at room temperature. The biodistribution results showed a notably higher and sustained brain uptake for INME compared to IVS and INS at all time intervals. In addition, heart and blood uptake levels for INME were lower than those for IV solution which, in turn, could decrease the systemic side effects of trazodone. Also, the 131I-trazodone INME brain uptake of 6.7±0.5%ID/g was higher than that of 99mTc-ECD and 99mTc-HMPAO (radiopharmaceuticals currently used for brain imaging). 131/123I-trazodone formulated as INME could be used as a promising radiopharmaceutical for brain imaging.

Biological Evaluation of 99mTc-Kanamycin for Infection Imaging

Kanamycin antibiotic was radiolabeled successfully with radioisotope technetium-99m for the potential use as radiopharmaceuticals for infection imaging. <sup>99m</sup>Tc-kanamycin complexes was prepared 93 % radiochemical purities by direct labelling using 5 mg kanamycin and 30 µg SnCl2. The reaction occurred at alkaline condition (pH=9) and under room temperature for 30 min to achieve high radiochemical purity. Radiochemical purity and stability of <sup>99m</sup>Tc-kanamycin was determined by ascending paper chromatography using Whatman 3 paper as the stationary phase, and acetone as the mobile phase to separate the radiochemical impurities in the form of <sup>99m</sup>Tc-pertechnetate. While impurities in the form of <sup>99m</sup>Tc-reduced were separated using the stationary phase ITLC-SG and 0.5 N NaOH as mobile phase. This study aimed to determine biological characteristic of <sup>99m</sup>Tc-kanamycin radiopharmaceutical. In vitro cell studies showed that the change of kanamycin structure after labeling with technetium-99m did not give a specific influence to the potency of kanamycin as an antibiotic. In addition on uptake study, a significantly higher uptake of <sup>99m</sup>Tc-kanamycin with S. aureus than E. coli. Biodistribution of <sup>99m</sup>Tc-kanamycin complexes was studied on normal and infection mice at 15, 30, 60 and 120 min post-injections. The biodistribution of <sup>99m</sup>Tc-kanamycin in infection mice showed that the complex accumulated in the infection sites. These results show that <sup>99m</sup>Tc-Kanamycin radiopharmaceutical have a potential application for infection diagnosis.

Radio-iodide uptake by modified poly (glycidyl methacrylate) as anion exchange resin

Abstract Poly(glycidyl methacrylate) (PGMA) microspheres were prepared by radiation induced polymerization of glycidyl methacrylate (GMA) monomer. The factors affecting the degree of polymerization and yield (%) of PGMA such as type of solvent, monomer concentration, and irradiation dose were investigated. It was found that the PGMA yield (%) increases with increasing monomer concentration up to 50% and absorbed dose of 5 kGy. The resulting PGMA containing the epoxy group waschemically modified by hydroxyl amine to act as anion-exchange resin for uptake of 131I− ions. The modified PGMA (MPGMA) was characterized by Fourier transform infra-red (FT-IR) spectrophotometer, thermal gravimetric analysis (TGA) and scanning electron microscopy (SEM). I-131 is produced from the fission of U-235 with low-enrichment uranium (LEU) targets in the Egyptian Second Research Reactor (ETRR-2). Separation of iodide from the radioactive solution by batchwise and column techniques was employed to determine the adsorption capacity of the MPGMA. Quality control of 131I product solution and radiochemical purity was examined by using the ascending paper chromatography method. The uptake behavior of MPGMA towards 131I− ions were studied at different experimental conditions and achieved by X-ray fluorescence (XRF). The synthesized MPGMA showed good results as anion-exchange and an effective adsorbent for uptaking 131I− ions.

Biodistribution and scintigraphic evaluation of 99mTc-Mannan complex.

Technetium-99m (99mTc) is extensively used in nuclear medicine, mostly used to label radiopharmaceuticals and in radio diagnostics. In the present study, we directly radiolabeled mannan with 99mTc by using Tin(II) Chloride Dihydrate (SnCl2·2H2O) as a reducing agent. Mannan, a TLR agonist is a complex carbohydrate identified as a potential modulator of biological effects of ionizing radiation, both in vitro and in vivo, in our laboratory. Under in vivo conditions mannan modulates radiation response when administered through either oral or parenteral routes. The present study aims to understand the pharmacologic biodistribution of the 99mTc-mannan complex in mice (via oral, i.p. and i.v. routes) using non-invasive scintigraphic imaging and invasive radiometry. Qualitative and quantitative analysis of 99mTc-mannan complex was performed by ITLC-SG, ascending paper chromatography. Radio-complexation efficiency of >98% was consistently achieved with hydrolyzed reduced Tc-99m being 1-2%. We confirmed stability of complex in saline and serum up to 24 h at room temperature. Biodistribution studies were performed using the above radiocomplex in BALB/c mice and 99mTc-mannan complex was administered though oral, i.p. and i.v. routes. To our expectations, most of the radioactivity accumulated in stomach and small intestine in mice with oral administration, along with insignificant activity in the remaining studied organs. It suggests that 99mTc-mannan complex did not get absorbed from the gut and was removed as such in the fecal material. On the contrary, i.p. and i.v administration of mannan resulted in significant accumulation of the 99mTc-mannan complex in kidney, liver, intestine, lungs, spleen, bone marrow, blood and heart, at both 1 h and 4 h after i.v. administration. The remaining organs (stomach, testis and muscles) showed lower accumulation of the 99mTc-mannan complex. 99mTc-mannan complex was adminstered (i.v.) in New Zealand white rabbits and it was evident from the scintigraphic images that mannan cleared very rapidly from the administration site and reached into systemic circulation. No activity in the thyroid, salivary gland, or gastric mucosa suggests an insignificant amount of free pertechnetate in the 99mTc-complex preparation, further confirming the in vivo stability of the radiolabeled mannan complex. Significant amount of radioactivity in liver, intestine and kidneys suggests hepatobiliary as well as renal routes of clearance. The bio-availability of the complex varies with the route of administration. An entirely different biodistribution pattern exists when the same molecule is administered through oral or parenteral route. Our study is the first step towards a better understanding of the mechanisms involved in radiation modulation offered by mannan administration, in vivo.

Preparation of <sup>99m</sup>Tc-Kanamycin Using a Direct Labeling Method

Infectious diseases are still the leading cause of death in the world. The accurate technique for early detection and determination of the exact location of infection in the body is still needed. Nuclear techniques are capable for this purpose while other techniques such as MRI, USG and CT-SCAN sometimes cannot be applied. 99m Tc-kanamycin radiopharmaceutical is complex of kanamycin and technetium-99m radionuclide, was used for bacterial detection of infection. The labeling studies of 99m Tc-kanamycin has been carried out by the indirect labeling method using pyrophosphate as a co-ligand with the results of labeling efficiency above 95% . However, the presence of radiochemical impurities in the form of 99m Tc-pyrophosphate in the indirect labeling have to be considered which may interfere the imaging result . This study aimed to determine the optimum labeling conditions of 99m Tc-kanamycin by direct labeling method . Kanamycin was successfully labeled with technetium-99m through direct labeling method. The labeling efficiency was determined by ascending paper chromatography using Whatman 3 paper as the stationary phase, and acetone as the mobile phase to separate the radiochemical impurities in the form of 99m Tc-pertechnetate. While impurities in the form of 99m Tc-reduced were separated using the stationary phase ITLC-SG and 0.5 N NaOH as mobile phase . The experiment result showed that the optimum labeling conditions obtained by using 5 mg kanamycin, 30 µg SnCl 2 .2H 2 O, and pH of labeling was 9. The incubation time of labeling was 30 min at room temperature, provided labeling efficiency of 92.31 ± 1.74 %.The successful of kanamycin labeling with high efficiency makes 99m Tc-kanamycin can potentially be used as a radiopharmaceutical for the early detection of infectious diseases Received: 04 October 2014 ; Revised: 28 June 2015; Accepted: 11 August 2015

Preparation, Biodistribution, and Scintigraphic Evaluation of 99mTc-Clindamycin: an Infection Imaging Agent

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-99m (~380MBq). 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-99m ((99m)Tc) was achieved at pH6-7 while using 2.5-3μg SnCl2 · H2O as a reducing agent and 100μg 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.

Effect of EDTA and PYP as Co-Ligand of Radiolabeled Nanomaterial M41S-NH<sub>2</sub> for Radiosynovectomy

Radiosynovectomy is an innovative medical treatment for rheumatoid arthritis joints. A small amount of beta-emitting radiopharmaceutical is injected into the joint with intra-articullar route of administration. An ideal radionuclide for this treatment is beta emitter whose minimal gamma ray emission, tissue penetration between 5 10 mm, has short half life, high chemical purity and non-toxic properties such as Rhenium-188. For delivering the radionuclide to the specific target site in the synovial tissue and preventing the leakage to the lymph, an ideal carrier as particulate or colloidal compounds is requaired. A silica-based nanomaterial called M41S-NH2 was developed as therapeutic carriers by Polytechnic Institute of Nuclear Technology, Yogyakarta. The advantage of M41S-NH2 is biocompatible and not toxic to human body. As a preliminary experiment, M41S-NH2 was labeled using Technetium-99m which has a similar chemical properties with Rhenium-188. The radiolabelling performed in this system is the indirect method. The purpose of this research is to obtain the highest radiochemical purity of radiolabeled 99mTc-M41S-NH2 by using two co-ligands such as ethylenediaminetetraasetic acid (EDTA) and sodium pyophosphate (PYP). Some parameter such as pH, ratio of SnCl2 to co-ligand, ammount of reductor agent, ammount of M41S-NH2 were optimized in those experiments. Radiochemical purity determination was conducted by ascending paper chromatography method using Whatman 31-ET as stationary phase and physiological saline (NaCl 0,9%) as mobile phase. The optimum condition obtained from the investigation were by using 10 μg PYP as co-ligand, 10 μg SnCl2 as reductor agent, pH 9, and 10 minutes in room temperature as incubation time. More over the highest radiochemical purity obtained from those labeling experiments is 93.42± 0.06 %.

Detection of Hg(II) amidst several heavy and toxic metal ions after their selective separation by chromatography: rationalization of separation factors in terms of Density Functional (hardness) Index

Ascending paper chromatography of Hg+2 has been performed with mixed acids as developing mobile phase. Effects of temperature, concentration, and composition of the mobile phase on Rf values have been investigated. Hg+2 has been selectively separated from several synthetic binary and multicomponent mixtures containing the toxic and heavy metal ions. Often these metal ions remain associated with it in its ores and alloy samples. Hg+2 in trace level has been separated and detected from blood serum containing the congeners Zn+2, Cd+2, Pb+2, Bi+3, Th+4, Tl+3, and Cu+2. On the basis of the difference in the migration of the spots on the paper, twenty difficult separations have been achieved. Several thermodynamic parameters, ΔG, ΔH, and ΔS have been determined by differential variation of radii of the spot with respect to temperature utilizing the thermodynamic relationship. The partition equilibrium constant was also determined. Metal ions were detected by borohydride reduction on the paper strip. Separation factors have been correlated in terms of DesityFunctional (corrected hardness, η∗) Index. From η∗, hydrated radii of the metal ions have been determined. A plausible mechanism for the differential migration of diverse metal ions has been suggested.

99mTc-Daunorubicin a potential brain imaging and theranostic agent: synthesis, quality control, characterization, biodistribution and scintigraphy

Daunorubicin is a chemotherapeutic antibiotic of the anthracycline family used for the treatment of many type of cancers when doxorubicin or other less effective drugs cannot be used. The aim of the present study was labeling of Daunorubicin with (99m)Tc, quality control, characterization, and biodistribution of radiolabeled Daunorubicin. Labeling efficiency was determined by ascending paper chromatography. All the experiments were performed at room temperature (25°C±2°C). More than 96% labeling efficiency with (99m)Tc was achieved at pH 5-6, 2-4 μg stannous chloride and 300 μg of ligand in few minutes. The characterization of the compound was performed by using HPLC, electrophoresis and shake flask assay. Electrophoresis indicates that Tc-99m-Daunorubicin is neutral, HPLC confirms the single specie of the labeled compound, while shake flask assay confirms high lipophilicity. The biodistribution studies of (99m)Tc-Daunorubicin were performed in rats. Significantly higher accumulation of (99m)Tc-Daunorubicin was seen in brain of normal rats. Scintigraphy was also indicating higher accumulation of (99m)Tc-Daunorubicin in brain of normal rabbits.

Tumor Targeting Using Anti–Epidermal Growth Factor Receptor (ior egf/r3) Immunoconjugate with a Tetraaza Macrocyclic Agent (DO3A-EA)

Epidermal growth factor receptor (EGFR) signaling inhibition represents a highly promising arena for the application of molecularly targeted cancer therapies. EGFR conjugated metal chelates have been proposed as potential imaging agents for cancers that overexpress EGFR receptors. Through improved understanding of EGFR biology in human cancers, there is anticipation that more tumor-selective therapy approaches with diminished collateral normal tissue toxicity can be advanced. We report here on the results with a thermodynamically stable chelate, 1,4,7-tris(carboxymethyl)-10-(2-aminoethyl)-1,4,7,10-tetraazacyclododecane (DO3A-EA) and anti-EGFr (ior egf/r3) conjugate to develop immunospecific imaging agent. Conjugation and labelling with anti-EGFr was performed using standard procedure and subjected to purification on size exclusion chromatography. The conjugated antibodies were labeled with a specific activity 20-30 mCi/mg of protein. Labeling efficiencies were measured by ascending paper chromatography on ITLC-SG strips. Radiolabeling of the immunoconjugate was found to be 98.5 ± 0.30%. (99m)Tc-DO3A-EA-EGFr conjugate was studied in athymic mice bearing U-87MG, MDA-MB-468 tumors following intravenous injection. Pharmacokinetic and biodistribution studies confirmed long circulation times (t(1/2)(fast) = 45 min and t1/2(slow) = 4 hours 40 min) and efficient accumulation in tumors. Biodistribution studies in athymic mice grafted with U-87MG human glioblastoma multiforme and Hela human cervical carcinoma tumors revealed significant localization of (99m)Tc-labeled antibodies conjugate in tumors and reduced accumulation in normal organs. This new chelating agent is promising for immunoscintigraphy since good tumour-to-normal organ contrast could be demonstrated. These properties can be exploited for immunospecifc contrast agents in nuclear medicine and SPECT imaging.

Determination of radiochemical and radionuclide purity of a 131I preparation

The radiochemical purity of a 131I preparation (the iodide fraction) was determined by ascending paper chromatography using a mixture of sodium iodide, sodium iodate, and sodium carbonate as a carrier and aqueous methanol as an eluent. The chromatogram was developed with a scanning β-ray spectrometer. The radiochemical purity of 131I determined by this procedure was 99.98%. The radionuclide purity of the 131I preparation was measured on a γ-ray spectrometer with an ultrapure Ge detector. The content of radionuclide impurities estimated from their detection limits in the 131I preparation isolated from TeO2 in 3–4 days after its irradiation in a reactor was no more than 0.03%. The 75Se content in the preparation aged for 6–7 months (which corresponds to a decrease in the 131I activity by a factor of 106–107 owing to decay) corresponds to 2 × 10−6% of the activity by the end of irradiation.

Role of the Carbohydrate Binding Site of the Streptococcus pneumoniae Capsular Polysaccharide Type 3 Synthase in the Transition from Oligosaccharide to Polysaccharide Synthesis

The type 3 synthase catalyzes the formation of the Streptococcus pneumoniae type 3 capsular polysaccharide [-3)-beta-D-GlcUA-(1, 4)-beta-D-Glc-(1-]n. Synthesis is comprised of two distinct catalytic phases separated by a transition step whereby an oligosaccharylphosphatidylglycerol primer becomes tightly bound to the carbohydrate acceptor recognition site of the synthase. Using the recombinant synthase in Escherichia coli membranes, we determined that a critical oligosaccharide length of approximately 8 monosaccharides was required for recognition of the growing chain by the synthase. Upon binding of the oligosaccharide-lipid to the carbohydrate recognition site, the polymerization reaction entered a highly processive phase to produce polymer of high molecular weight. The initial oligosaccharide-synthetic phase also appeared to be processive, the duration of which was enhanced by the concentration of UDP-GlcUA and diminished by an increase in temperature. The overall reaction approached a steady state equilibrium between the polymer- and oligosaccharide-forming phases that was shifted toward the former by higher UDP-GlcUA levels or lower temperatures and toward the latter by lower concentrations of UDP-GlcUA or higher temperatures. The transition step between the two enzymatic phases demonstrated cooperative kinetics, which is predicted to reflect a possible reorientation of the oligosaccharide-lipid in conjunction with the formation of a tight binding complex.