The determination of the specific activity of plasma glycerol

1982 ◽  
Vol 60 (6) ◽  
pp. 856-858 ◽  
Author(s):  
Clément Gauthier ◽  
Ross Layberry
Keyword(s):  
Column Chromatography ◽  
Turnover Rate ◽  
Specific Activity ◽  
Anionic Exchange ◽  
Removal Of Contaminants ◽  
The One ◽  
Exchange Resins

A method for the determination of the specific activity of plasma glycerol is described. Anionic contaminants are first removed from deproteinized plasma by anionic exchange resins (treated plasma). Glycerol in treated plasma is then quantitatively converted to glycerol-3-phosphate (G3P), which is isolated by column chromatography and counted for 14C radioactivity. The specific activity thus calculated was 100.1 ± 2.9% of a standard of known specific activity. When the specific-activity of glycerol is determined from plasma without prior removal of anionic contaminants (untreated plasma), the calculated specific activity is 1.99 ± 0.15 times higher than the one calculated after their removal. Omission of the removal of contaminants leads to a near 100% error in the calculation of the turnover rate of glycerol.not available

Improvements in the determination of the turnover rate of plasma thyroxine in avian species: application to the Japanese quail

1987 ◽  
Vol 114 (2) ◽  
pp. 191-198 ◽  
Author(s):  
E. J. Cookson ◽  
J. Glover
Keyword(s):  
Japanese Quail ◽  
Half Life ◽  
Turnover Rate ◽  
Specific Activity ◽  
Simple Procedure ◽  
High Specific Activity ◽  
Future Application ◽  
Sodium Thiocyanate ◽  
Plasma Samples

ABSTRACT The disappearance of the thyroid hormone thyroxine (T4) from plasma in fully grown male Japanese quail can be described as a first order process with a rate constant of 0·178 ± 0·013/h (mean±s.e.m., n = 8), which represents a half-life of 3·90 h. A small amount of [125I]T4 in relation to total circulating T4 was injected i.v. into Japanese quail and plasma samples were taken at appropriate time-intervals for the determination of residual plasma radioactivity. The rate of disappearance of [125I]T4 was subsequently equated to the turnover rate of the endogenous hormone. Previous methods were modified to overcome problems arising from possible disturbance of plasma T4 metabolism, recirculation of radiolabelled iodide, and to purify the [125I]T4 from the plasma samples. By using labelled T4 of very high specific activity, the amount of [125I]T4 administered was kept much smaller than has been used in previous studies on Japanese quail, thus limiting any interference with plasma T4 dynamics. To minimize any disturbance of plasma T4 metabolism, only four blood samples were taken, at three-hourly intervals after the injection of [125I]T4. The rapid turnover of T4 produced a large amount of labelled inorganic iodide, the re-entry of which into the plasma T4 pool was inhibited by s.c. administration of sodium thiocyanate 1 h before injection of[125I]T4. Assay of the true [125I]T4 turnover was significantly improved over that used in previous studies by purifying the [125I]T4 from the plasma samples chromatographically. The samples were applied to small Sephadex G-25 columns with sodium hydroxide (0·1 mol/l) as the eluant. This simple procedure clearly separated the [125I]T4 from the other radioiodinated plasma components such as free iodide, non-hormonal iodinated proteins and triiodothyronine (T3), thus enabling a more accurate assessment of the residual labelled T4 concentration in the plasma and hence the T4 half-life. The future application of this method to the study of plasma T4 turnover under various experimental conditions is discussed and the possible involvement of T4 turnover studies in the assessment of T4 to T3 conversion is considered. J. Endocr. (1987) 114, 191–198

Determination of the role of elevated B2 esterase in insecticide resistance in Culex quinquefasciatus (Diptera: Culicidae) from studies on the purified enzyme

1994 ◽  
Vol 84 (1) ◽  
pp. 39-43 ◽  
Author(s):  
K.G.I. Jayawardena ◽  
S.H.P.P. Karunaratne ◽  
A.J. Ketterman ◽  
J. Hemingway
Keyword(s):  
Culex Quinquefasciatus ◽  
Turnover Rate ◽  
Specific Activity ◽  
Sri Lankan ◽  
Organophosphorus Insecticides ◽  
Purification And Characterization ◽  
Sds Page

AbstractA method for the purification of elevated esterase B2 from the Sri Lankan Pel RR strain of Culex quinquefasciatus (Say) has been developed, using sequential column chromatography. The pure enzyme is stable, with no decrease in specific activity after several months, if stored in the presence of 25 mM dithiothreitol and 50% glycerol at –20°C. The enzyme has a pl of 5 and an estimated molecular weight for the monomeric enzyme of 63 kD from SDS-PAGE. In all Culex strains with elevated B2, another esterase (A2) is always co-elevated. The strains with these esterases are resistant to a broad range of organophosphorus insecticides. The previous purification and characterization of A2 from Pel RR indicated that the role of this enzyme was primarily to sequester rather than metabolize organophosphates. Purified B2, rapidly binds malaoxon, the activated form of malathion, but not malathion itself. The turnover rate of bound malaoxon is slow, suggesting that the role of this esterase in malathion resistance is also sequestration. The similar bimolecular rate constants for both A2 and B2 with malaoxon suggest both esterases are important in resistance, which may explain the almost complete linkage disequilibrium that the elevated forms of these two esterases are maintained in when under pesticide selection pressure.

Estimation of cyclic AMP turnover in normal and methylprednisolone-treated dogs: effect of catecholamines

1975 ◽  
Vol 229 (2) ◽  
pp. 291-297 ◽  
Author(s):  
TB Issekutz
Keyword(s):  
Plasma Level ◽  
Turnover Rate ◽  
Specific Activity ◽  
Specific Binding ◽  
Mass Action ◽  
Glucocorticoid Treatment ◽  
Norepinephrine Infusion ◽  
Specific Binding Protein ◽  
Plasma Camp

The turnover rate of plasma cAMP was measured in unanesthetized normal and methylprednisolone (MP)-treated dogs with [8-3H]cAMP. Determination of specific activity was based on saturation of the specific binding protein, thus bypassing the need to know the plasma level of cAMP. In normal dogs the infusion of epinephrine increased plasma glucose by 33% and both the plasma level and rate of appearance (Ra) of cAMP 2.5-fold. Treatment with MP decreased the concentration as well as the turnover rate of cAMP by about 50%. It greatly potentiated not only the hyperglycemic effect (4-fold) of epinephrine but also the rise of plasma cAMP (8.5-fold) and that of Ra (6.8-fold). Norepinephrine infusion increased cAMP only by 50%. This effect was not potentiated by MP treatment and not altered by Na nicotinate. There was a linear correlation between plasma cAMP concentration and the rate of disappearance (Rd) of cAMP. It is concluded that: a) the increase of hepatic glycogenolysis is accompanied by a rise of cAMP output, whereas that of lipolysis is not; b) glucocorticoid treatment affects cAMP turnover; and c) the plasma level of cAMP is controlled by Ra, whereas Rd is the result of the mass-action effect of the concentration.

Two-Stage Procedure for the Quantitative Determination of Autoprothrombin III Concentration and Some Applications

1967 ◽  
Vol 18 (01/02) ◽  
pp. 198-210 ◽  
Author(s):  
Ronald S Reno ◽  
Walter H Seegers
Keyword(s):  
Prothrombin Time ◽  
Factor Vii ◽  
Two Stage ◽  
Pronounced Increase ◽  
One Stage ◽  
Assay Procedure ◽  
Bovine Plasma ◽  
The One ◽  
Autoprothrombin C

SummaryA two-stage assay procedure was developed for the determination of the autoprothrombin C titre which can be developed from prothrombin or autoprothrombin III containing solutions. The proenzyme is activated by Russell’s viper venom and the autoprothrombin C activity that appears is measured by its ability to shorten the partial thromboplastin time of bovine plasma.Using the assay, the autoprothrombin C titre was determined in the plasma of several species, as well as the percentage of it remaining in the serum from blood clotted in glass test tubes. Much autoprothrombin III remains in human serum. With sufficient thromboplastin it was completely utilized. Plasma from selected patients with coagulation disorders was assayed and only Stuart plasma was abnormal. In so-called factor VII, IX, and P.T.A. deficiency the autoprothrombin C titre and thrombin titre that could be developed was normal. In one case (prethrombin irregularity) practically no thrombin titre developed but the amount of autoprothrombin C which generated was in the normal range.Dogs were treated with Dicumarol and the autoprothrombin C titre that could be developed from their plasmas decreased until only traces could be detected. This coincided with a lowering of the thrombin titre that could be developed and a prolongation of the one-stage prothrombin time. While the Dicumarol was acting, the dogs were given an infusion of purified bovine prothrombin and the levels of autoprothrombin C, thrombin and one-stage prothrombin time were followed for several hours. The tests became normal immediately after the infusion and then went back to preinfusion levels over a period of 24 hrs.In other dogs the effect of Dicumarol was reversed by giving vitamin K1 intravenously. The effect of the vitamin was noticed as early as 20 min after administration.In response to vitamin K the most pronounced increase was with that portion of the prothrombin molecule which yields thrombin. The proportion of that protein with respect to the precursor of autoprothrombin C increased during the first hour and then started to go down and after 3 hrs was equal to the proportion normally found in plasma.

An Enzyme Linked Immunosorbent Assay for Determination of Tissue Plasminogen Activator Applied to Patients with Thromboembolic Disease

1983 ◽  
Vol 50 (03) ◽  
pp. 740-744 ◽  
Author(s):  
Nils Bergsdorf ◽  
Torbjörn Nilsson ◽  
Per Wallén
Keyword(s):  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Specific Activity ◽  
Thromboembolic Disease ◽  
Tissue Type ◽  
Enzyme Linked Immunosorbent Assay ◽  
Type Plasminogen Activator ◽  
Tissue Plasminogen ◽  
Normal Human

SummaryUtilizing the immunoglobulin fraction from a goat antiserum against human uterine tissue plasminogen activator, an enzyme- linked immunoassay for tissue-type plasminogen activator in human plasma has been developed. With the new method, the concentration of t-PA in normal human acidified plasma is found to be 4.0 ± 1.8 (SD) ng/ml. It increases to 12 ng/ml after a tomiquet test, and to 14 ng/ml after strenous physical exercise. In a group of patients with idiopathic thromboembolic disease, the resting t-PA concentration was 5 ng/ml and the post-occlusion value 16 ng/ml. Furthermore, the patients also exhibited a normal post-occlusion rise in the concentration of plasmin-α2-antiplasmin complex. However, in 37% of the post-occlusion patient plasmas, virtually no increase in t-PA could be detected by a specific activity assay. The results indicate that the reason for a defective post-occlusion fibrinolytic activity in a majority of cases may be the presence of increased concentrations of a fast-acting specific t-PA inhibitor.

SIMPLIFIED DEXAMETHASONE SUPPRESSION TEST

1969 ◽  
Vol 61 (2) ◽  
pp. 219-231 ◽  
Author(s):  
V. H. Asfeldt
Keyword(s):  
Cushing’S Syndrome ◽  
Normal Subjects ◽  
Cushing's Syndrome ◽  
Dexamethasone Suppression Test ◽  
Clinical Value ◽  
Suppression Test ◽  
The One ◽  
Steroid Excretion ◽  
Dexamethasone Suppression

ABSTRACT This is an investigation of the practical clinical value of the one mg dexamethasone suppression test of Nugent et al. (1963). The results, evaluated from the decrease in fluorimetrically determined plasma corticosteroids in normal subjects, as well as in cases of exogenous obesity, hirsutism and in Cushing's syndrome, confirm the findings reported in previous studies. Plasma corticosteroid reduction after one mg of dexamethasone in cases of stable diabetes was not significantly different from that observed in control subjects, but in one third of the insulin-treated diabetics only a partial response was observed, indicating a slight hypercorticism in these patients. An insufficient decrease in plasma corticosteroids was observed in certain other conditions (anorexia nervosa, pituitary adenoma, patients receiving contraceptive or anticonvulsive treatment) with no hypercorticism. The physiological significance of these findings is discussed. It is concluded that the test, together with a determination of the basal urinary 17-ketogenic steroid excretion, is suitable as the first diagnostic test in patients in whom Cushing's syndrome is suspected. In cases of insufficient suppression of plasma corticosteroids, further studies, including the suppression test of Liddle (1960), must be carried out.

Determination of Specific Activity of Cholera Chemical Vaccine Components using Cell Culture

2020 ◽  
Vol 36 (3) ◽  
pp. 82-89
Author(s):  
O.V. Gromova ◽  
O.S. Durakova ◽  
S.V. Generalov ◽  
L.F. Livanova ◽  
O.A. Volokh
Keyword(s):  
Cell Culture ◽  
Vibrio Cholerae ◽  
Production Control ◽  
Specific Activity ◽  
Methodological Approach ◽  
Toxin Production ◽  
Submerged Cultivation ◽  
Vaccine Production ◽  
Antigenic Activity

Том 36(2020) №3 стр. 82-89; DOI 10.21519/0234-2758-2020-36-3-82-89А.В. Гаева1*, О.В. Громова1, О.С. Дуракова1, С.В. Генералов1, Л.Ф. Ливанова1, О.А. Волох1 Определение специфической активности компонентов холерной химической вакцины с использованием культуры клеток 1ФКУЗ «Российский научно-исследовательский противочумный институт «Микроб»» Федеральной службы по надзору в сфере защиты прав потребителей и благополучия человека, Саратов 410005 *[email protected] Поступила - 2019-11-26; После доработки - 2020-03-16; Принята к публикации - 2020-05-15 Список литературы Описаны методы определения динамики продукции токсинов штаммом Vibrio cholerae 569B при глубинном культивировании в биореакторе и антигенной активности специфической фракции холерогена-анатоксина по анатоксинсвязыванию с использованием клеточных культур. Показана высокая степень соответствия результатов, полученных методами, применяемыми для контроля этапов производства холерной химической вакцины и рассмотренными в данной работе. Отмечено, что применение клеточной линии СНО-К1 наиболее перспективно для замены биомоделей на промежуточных этапах контроля активных компонентов холерной химической вакцины. Разработанный методический подход впервые предлагается использовать на этапах производства холерной бивалентной химической вакцины. культура клеток, Vibrio cholerae, холерная химическая вакцина, контроль производства, холера. Vol 36(2020) N 3 p. 82-89; DOI 10.21519/0234-2758-2020-36-3-82-89A.V. Gaeva1*, O.V. Gromova1, O.S. Durakova1, S.V. Generalov1, L.F. Livanova1, O.A. Volokh1 Determination of Specific Activity of Cholera Chemical Vaccine Components using Cell Culture 1Russian Research Anti-Plague Institute «Microbe» of the Federal Service for Surveillance on Consumer Rights Protection and Human Wellbeing, Saratov, 410005 *[email protected] Received - 26.11.2019; Accepted - 15.05.2020 References The methods has been described to determine the dynamics of toxin production by the Vibrio cholerae 569B strain during submerged cultivation in bioreactor and of the antigenic activity of specific choleragen anatoxin fraction by anatoxin binding levels using cell cultures. High degree of consistency was observed between the results obtained via the method under consideration and those obtained via control methods at different stages of cholera chemical vaccine production. It was shown that the CHO-K1 cell line is the most promising substitute for biomodels at the intermediate stages of control of active cholera chemical vaccine components. The developed methodological approach was first proposed for use at the stages of cholera chemical bivalent vaccine manufacturing. cell culture, Vibrio cholerae, cholera chemical vaccine, production control, cholera.

A Validated 2D-LC-UV Method for Simultaneous Determination of Imatinib and N-demethylimatinib in Plasma and Its Clinical Application for Therapeutic Drug Monitoring with GIST Patients

2020 ◽  
Vol 17 ◽  
Author(s):  
Houli Li ◽  
Di Zhang ◽  
Xiaoliang Cheng ◽  
Qiaowei Zheng ◽  
Kai Cheng ◽  
...  
Keyword(s):  
Therapeutic Drug Monitoring ◽  
One Dimension ◽  
Therapeutic Drug ◽  
Plasma Samples ◽  
Column Temperature ◽  
Injection Volume ◽  
Target Analytes ◽  
The One ◽  
Middle Column

Background: The trough concentration (Cmin) of Imatinib (IM) is closely related to the treatment outcomes and adverse reactions of patients with gastrointestinal stromal tumors (GIST). However, the drug plasma level has great interand intra-individual variability, and therapeutic drug monitoring (TDM) is highly recommended. Objective: To develop a novel, simple, and economical two-dimensional liquid chromatography method with ultraviolet detector (2D-LC-UV) for simultaneous determination of IM and its major active metabolite, N-demethyl imatinib (NDIM) in human plasma, and then apply the method for TDM of the drug. Method: Sample was processed by simple protein precipitation. Two target analytes were separated on the one-dimension column, captured on the middle column, and then transferred to the two-dimension column for further analysis. The detection was performed at 264 nm. The column temperature was maintained at 40˚C and the injection volume was 500 μL. Totally 32 plasma samples were obtained from patients with GIST who were receiving IM. Method: Sample was processed by simple protein precipitation. Two target analytes were separated on the one-dimension column, captured on the middle column, and then transferred to the two-dimension column for further analysis. The detection was performed at 264 nm. The column temperature was maintained at 40˚C and the injection volume was 500 μL. Totally 32 plasma samples were obtained from patients with GIST who were receiving IM. Conclusion: The novel 2D-LC-UV method is simple, stable, highly automated and independent of specialized technicians, which greatly increases the real-time capability of routine TDM for IM in hospital.

The Natural Radioactivity in Food: A Comparison Between Different Feeding Regimes

2019 ◽  
Vol 15 (5) ◽  
pp. 493-499 ◽  
Author(s):  
Francesco Caridi ◽  
Santina Marguccio ◽  
Alberto Belvedere ◽  
Maurizio D`Agostino ◽  
Giovanna Belmusto
Keyword(s):  
Vegetable Oils ◽  
Natural Radioactivity ◽  
Specific Activity ◽  
Mean Value ◽  
Vegetable Food ◽  
Fish Samples ◽  
Feeding Regimes ◽  
Italian Origin ◽  
Dose Levels

Background: In this article a comprehensive study was carried out for the determination of natural radioactivity in animal and vegetable food (meat, fish, milk and derivates, legumes, cereals and derivates, fruit, hortalizas, vegetables, vegetable oils) typical of different feeding regimes, for the age category higher than 17 years. Methods: A total of eighty-five samples of Italian origin, coming from large retailers during the years 2014, 2015 and 2016, were analyzed through HPGe gamma spectrometry. Results: The specific activity of 40K was investigated and its mean value was found to be: (106.3 ± 6.9) Bq/kg for bovine, swine and sheep meat; (116.5 ± 9.7) Bq/kg for fish; (52.9 ± 3.1) Bq/kg for milk and derivates; (271.9 ± 16.7) Bq/kg for legumes; (67.2 ± 4.7) Bq/kg for cereals and derivates; (52.7 ± 4.4) Bq/kg for fruit; (72.9 ± 5.6) Bq/kg for hortalizas; (83.9 ± 6.5) Bq/kg for vegetables; lower than the minimum detectable activity for vegetable oils. For animal food the highest mean 40K activity concentration was found in fish samples; for vegetable food the highest one was detected in legumes. Conclusion: The evaluation of dose levels due to the food ingestion typical of Mediterranean, Vegetarian and Vegan diets was performed. The annual effective dose was found to be 0.16 mSv/y, 0.41 mSv/y and 0.54 mSv/y, respectively.

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