scholarly journals Rat Serum Carboxylesterase Partly Hydrolyses Gamma-Butyrobetaine Esters

2009 ◽  
Vol 60 (2) ◽  
pp. 147-156 ◽  
Author(s):  
Lida Bagdonienė ◽  
Danutė Labeikytė ◽  
Ivars Kalviņš ◽  
Veronika Borutinskaitė ◽  
Aleksandrs Prokofjevs ◽  
...  
Keyword(s):  
Blood Serum ◽  
Esterase Activity ◽  
Colorimetric Determination ◽  
Reaction Products ◽  
Rat Serum ◽  
Rat Blood ◽  
Optical Absorbance ◽  
Hydrolysis Of ◽  
Related Compounds

Rat Serum Carboxylesterase Partly Hydrolyses Gamma-Butyrobetaine EstersAlthough described some time ago, gamma-butyrobetaine esters and related compounds have not gained much attention from researchers, and their physiological function remains obscure. Formerly we detected GBB-esterase enzymatic activity in rat blood serum using phenylated gamma-butyrobetaine as an artificial substrate of the enzyme and HPLC. The aim of the present work was to develop an assay that would enable spectrophotometric or colorimetric determination of the reaction products of GBB-esterase activity and to reveal individual proteins performing GBB-esterase activity in rat blood serum. For this purpose gamma-butyrobetaine 1-naphthyl ester was synthesised. Hydrolysis of this ester releases 1-naphthol, which increases the optical absorbance at 322 nm. We have shown that the enzymatic hydrolysis of GBB 1-naphthyl ester to 1-naphthol in rat blood serum is due to GBB-esterase activity. An attempt was done to purify the enzyme from rat blood serum. By combining DEAE Sepharose at pH 4.2 and affinity chromatography with procainamide we achieved a 68-fold enrichment of GBB-esterase activity in our preparations. Separation of fraction proteins in 2D protein electrophoresis with following mass-spectrometry indicated that GBB esterase activity in rat blood serum is performed in part by carboxylesterase.

scholarly journals Development of Simultaneous Determination Method of a Pregnan Steroid with Gestagenic Activity – Gestobutanoil and Two its Metabolites in Rat Serum

2021 ◽  
Vol 10 (2) ◽  
pp. 112-118
Author(s):  
E. S. Stepanova ◽  
L. M. Makarenkova ◽  
S. V. Goryainov ◽  
T. A. Fedotcheva ◽  
N. L. Shimanovsky
Keyword(s):  
Blood Serum ◽  
Simultaneous Determination ◽  
Megestrol Acetate ◽  
Reaction Products ◽  
Active Metabolites ◽  
Rat Serum ◽  
Rat Blood ◽  
Esi Ms ◽  
Pharmacologically Active

Introduction. Gestobutanoil is a synthetic pregnane steroid with gestagenic activity. Gestobutanoil has two pharmacologically active metabolites (AMOL and megestrol acetate). This implies the need for a detailed study of the kinetics of metabolites. It is rational to combine the study of the pharmacokinetics of gestobutanoil and its metabolites (AMOL and megestrol acetate). The simultaneous determination of several analytes in the rats’ serum can be carried out using chromatography-mass-spectrometry.Aim. Development of an analytical method for the simultaneous determination of gestobutanoil and two its metabolites in a biomatrix (rat serum).Materials and methods. The following methods were used to determine gestobutanoyl and two its metabolites in a biological matrix: GC-MS, HPLCESI-MS, HPLC-ESI-MS with derivatization, HPLC-APCI-MS.Results and discussion. When working with GC-MS, the chromatographic peaks of gestobutanoyl, AMOL, and megestrol acetate were strongly blurred and superimposed on each other, which is apparently due to the thermolability of the substances. The GC-MS method was abandoned in favor of HPLC. Analytes were separated by HPLC gradient elution on a C18 column. ESI ionization did not give typical protonated ions of gestobutanoyl and AMOL, and the intense signals of their cationized ions and fragment ions, which were observed in the spectra of AMOL and gestobutanoyl, could not ensure the reproducibility of the spectra, since the conditions of their formation are not suitable for routine analysis. Derivatization of analytes to form oximes and substituted hydrazones did not give the expected reaction products for HPLC-ESI-MS. APCI made it possible to remove intense cationized ions from the spectra of gestobutanoyl and AMOL and to increase the reliability of the method. The HPLCAPCI-MS technique was reproduced on model rat blood serum.Conclusion. An HPLC-MS method was developed for the simultaneous determination of gestobutanoyl, megestrol acetate, and AMOL. The technique was tested on a model rat blood serum containing all three analytes.

scholarly journals Protocol for purification of a rat blood serum protein fraction enriched in gamma-butyrobetaine esterase activity

2009 ◽  
Vol 63 (3) ◽  
pp. 93-99
Author(s):  
Lida Bagdonienė ◽  
Danutė Labeikytė ◽  
Ivars Kalviņš ◽  
Benediktas Juodka ◽  
Nikolajs Sjakste
Keyword(s):  
Enzymatic Activity ◽  
Blood Serum ◽  
Serum Protein ◽  
Esterase Activity ◽  
Protein Fraction ◽  
Physiological Function ◽  
Rat Blood ◽  
Serum Protein Fraction ◽  
Blood Serum Protein ◽  
Related Compounds

Protocol for purification of a rat blood serum protein fraction enriched in gamma-butyrobetaine esterase activity Although described some time ago, gamma-butyrobetaine esters and related compounds have not gained much attention from researchers, and their physiological function remains obscure. Formerly we detected GBB-esterase enzymatic activity in rat blood serum. The aim of the present work was to develop a protocol that would enable purification of the protein fraction enriched in GBB esterase activity from rat blood serum. Chromatography on DEAE Sepharose at pH 4.2 enabled to purify a protein fraction enriched in enzymatic activity, but represented by numerous polypeptides. Following separation of this fraction by means of chromatography on DEAE Sepharose at pH 6.5 or heparin Sepharose chromatography at pH 7.0 did not lead to significant decrease of polypeptide number. When the above fraction was further fractionated by means of DEAE Sepharose chromatography at pH 7.4 or Bio Gel P150 chromatography the enzymatic activity was lost. Combination of DEAE Sepharose at pH 4.2 and affinity chromatography with procainamide appears to be the most suitable approach.

Polarographic study of hydrolysis of [8-lysine]vasopressin and its derivatives with blood serum of pregnant women

1980 ◽  
Vol 45 (4) ◽  
pp. 1099-1108 ◽  
Author(s):  
Mikuláš Chavko ◽  
Michal Bartík ◽  
Evžen Kasafírek
Keyword(s):  
Blood Serum ◽  
Pregnant Women ◽  
Degree Of Hydrolysis ◽  
Polarographic Study ◽  
Ph Optimum ◽  
Lysine Vasopressin ◽  
Rate Of Hydrolysis ◽  
Hydrolysis Of ◽  
Vasopressin Analogues

A polarographic study of the hydrolysis of [8-lysine]vasopressin and some hormonogens of the vasopressin series with the blood serum of women in the last week of pregnancy was studied. The dependence of hydrolysis on pH (pH optimum: 7.4-7.50, substrate concentration (Km 1.2 . 10-5M), pH stability and thermal stability were determined. The rate of hydrolysis of individual vasopressin analogues decreases in the order: [8-lysine]vasopressin > Nα-glycyl-prolyl[8-lysine]-vasopressin > Nα-leucyl-[8-lysine]vasopressin > Nα-alanyl-[8-lysine]vasopressin > Nα-phenyl alanyl-[8-lysine]vasopressin > Nα-diglycyl-[8-lysine]vasopressin > Nα-prolyl-[8-lysine]vasopressin > Nα-triglycyl-[8-lysine]vasopressin > Nα-sarcosyl-glycyl-[8-lysine]vasopressin. The degree of hydrolysis gradually increases to a multiple with the length of the pregnancy in consequence of the presence of oxytocine. However, vasopressin is also hydrolysed to a small extent with the enzymes from the blood sera of non-pregnant women. Under similar analytical conditions oxytocin was not hydrolysed with the sera of non-pregnant women and therefore oxytocin is a more suitable substrate than vasopressin for polarographic determination of serum oxytocinase.

scholarly journals A simple method for the determination of the cholesterol esterase activity.

2013 ◽  
Vol 60 (3) ◽  
Author(s):  
Agnieszka Ewa Stępień ◽  
Mykhailo Gonchar
Keyword(s):  
Human Serum ◽  
Esterase Activity ◽  
Cholesterol Oxidase ◽  
Cholesterol Esterase ◽  
Cholesterol Esters ◽  
Simple Method ◽  
Bacterial Enzyme ◽  
Low Costs ◽  
Hydrolysis Of

The proposed method determines the activity of cholesterol esterase (CEH) and takes advantage of its ability to catalyze the hydrolysis of cholesterol esters naturally present in human serum. The assay is based on Allain's method of spectrophotometric determination of cholesterol by means of cholesterol oxidase, peroxidase, but using 3,5-dichloro-dihydroxybenzenesulfonic acid (DHBS) as phenolic chromogen and human serum as a source of substrate for the CEH as a novelty. Furthermore, it is characterized by low costs and high precision. It can be employed to control the activity of CE preparations used for the preparation of enzymatic kits for the determination of cholesterol or for screening of potential bacterial enzyme producers.

Enzymic colorimetric determination of phosphatidylglycerol in amniotic fluid.

1984 ◽  
Vol 30 (4) ◽  
pp. 534-537 ◽  
Author(s):  
J D Artiss ◽  
M W McGowan ◽  
D R Strandbergh ◽  
E Epstein ◽  
B Zak
Keyword(s):  
Amniotic Fluid ◽  
Fetal Lung ◽  
Aqueous Sample ◽  
Colorimetric Determination ◽  
Glycerol Phosphate ◽  
Ionic Detergent ◽  
Catalyzed Reactions ◽  
Enzyme Catalyzed Reactions ◽  
Hydrolysis Of

Abstract We describe a procedure for the enzymic, colorimetric determination of phosphatidylglycerol in amniotic fluid. After extraction into chloroform:methanol (2:1 by vol) and evaporation, the phospholipid-containing residue is redissolved in a non-ionic detergent, which thus provides an aqueous sample. The subsequent enzymic reaction sequence involves phospholipase-catalyzed hydrolysis of glycerol from its phospholipid. Subsequent enzyme-catalyzed reactions phosphorylate this glycerol and oxidize the resulting glycerol phosphate to produce hydrogen peroxide, which is reacted to produce an intense red chromogen in the peroxidase-catalyzed coupling of 4-aminoantipyrine and 2-hydroxy-3,5-dichlorobenzenesulfonate. When used in conjunction with previously reported enzymic techniques for determination of lecithin and sphingomyelin, this procedure may provide an accurate and precise "lung profile" for assessment of fetal lung maturity.

Determination of Sevin Insecticide Residues in Fruits and Vegetables

1964 ◽  
Vol 47 (2) ◽  
pp. 283-286
Author(s):  
D P Johnson
Keyword(s):  
Fruits And Vegetables ◽  
Collaborative Study ◽  
Colorimetric Determination ◽  
Average Recovery ◽  
Insecticide Residues ◽  
Chromogenic Agent ◽  
Hydrolysis Of ◽  
Concentration Levels ◽  
Good Agreement

Abstract Collaborative study of a method for Sevin insecticide residues was repeated with apples and lettuce as test crops. The method is based on alkaline hydrolysis of Sevin and colorimetric determination of the resulting 1-naphthol with p-nitrobenzenediazonium fluoborate as chromogenic agent. Data from 5 collaborators were in good agreement with an average recovery of 87.8% at 2 concentration levels. It is recommended that the method be adopted as official, first action.

Colorimetric Determination of Certain Phenothiazine Drugs by Using Morpholine and Iodine-Potassium Iodide Reagents

1986 ◽  
Vol 69 (3) ◽  
pp. 513-518 ◽  
Author(s):  
Adel F Youssef ◽  
Salwa R El-Shabouri ◽  
Fardous A Mohamed ◽  
Abdel Maboud I Rageh
Keyword(s):  
Potassium Iodide ◽  
Quantitative Estimation ◽  
Colorimetric Method ◽  
Molar Absorptivity ◽  
Dosage Forms ◽  
Colorimetric Determination ◽  
Reaction Products ◽  
Phenothiazine Drugs ◽  
Lower Molar

Abstract A colorimetric method was developed for the quantitative estimation of 11 phenothiazine drugs. The method is based on the interaction of unsulfoxidized drug with morpholine and iodine-potassium iodide reagents. The interaction for all studied phenothiazine drugs yields a blue product with 2 absorption maxima: one in the range of 620-640 nm with lower molar absorptivity and the other in the range of 662-690 nm with higher molar absorptivity. The color was stable for at least 10 h. The reproducibility and recovery of the method were excellent. The method was applied successfully to the analysis of various commercially available phenothiazines in different dosage forms. The results were comparable to those obtained by official procedures. The suitability of the method for detection and estimation of promethazine excreted in urine has been suggested by preliminary experiments. Reaction products have been isolated and identified.

An Automated Alkaline Phosphatase Assay with Phenolphthalein Monophosphate as Substrate

1967 ◽  
Vol 13 (4) ◽  
pp. 281-289 ◽  
Author(s):  
Kirsten Hviid
Keyword(s):  
Alkaline Phosphatase ◽  
Normal Values ◽  
Serum Alkaline Phosphatase ◽  
Colorimetric Determination ◽  
Alkaline Phosphatase Assay ◽  
Phosphatase Assay ◽  
Hydrolysis Of

Abstract The manual procedure of Babson et al. (1) for the determination of serum alkaline phosphatase has been automated. The assay is based on the colorimetric determination of phenolphthalein formed on hydrolysis of phenolphthalein monophosphate. The procedure utilizes 0.16 ml. of serum without dialysis. Blanks are required only for turbid sera. Results are compared with those obtained by the manual procedure, and data relating to sample interaction, precision, blank values, and normal values are presented.

Colorimetric Determination of Certain Phenol Derivatives in Pharmaceutical Preparations

1981 ◽  
Vol 64 (6) ◽  
pp. 1442-1445
Author(s):  
Sayed M Hassan ◽  
Mohammed I Walash ◽  
Sanaa M El-Sayed ◽  
Abdel Malek Abou Ouf
Keyword(s):  
Pharmaceutical Preparations ◽  
Sulfanilic Acid ◽  
Diazonium Salts ◽  
Colorimetric Determination ◽  
Reaction Products ◽  
Optimum Conditions ◽  
Phenol Derivatives ◽  
The Mean ◽  
Colorimetric Methods

Abstract Simple colorimetric methods are reported for determining both acetaminophen and oxyphenbutazone. These methods are based on coupling between the phenolic compound and the diazonium salts of both sulfanilic acid and p-nitroaniline; the optimum conditions for the reactions were carefully studied. For acetaminophen, the reaction products with diazosulfanilic acid and diazo-p-nitroaniline show maximum absorbance at 480 and 425 nm, respectively. The mean percentage recoveries for authentic samples were 99.5 ± 1.1 and 100.6 ± 0.66, respectively (P = 0.05). For oxyphenbutazone, the obtained colors showed maxima at 385 nm with diazosulfanilic acid and 490 nm with diazo-p-nitroaniline reactions. The mean percentage recoveries for authentic samples were 99.8 ± 0.27 and 100.1 ± 0.57, respectively (P = 0.05). The proposed methods were successfully applied to the analysis of commercial preparations; results were statistically compared with those of other methods.

THE PATTERN OF URINARY EXCRETION OF INDIVIDUAL 17-KETOSTEROIDS IN RHESUS MONKEYS

1971 ◽  
Vol 67 (4) ◽  
pp. 711-720 ◽  
Author(s):  
S. B. Pal
Keyword(s):  
Rhesus Monkeys ◽  
Enzyme Hydrolysis ◽  
Colorimetric Determination ◽  
Day Range ◽  
Extraction Separation ◽  
Normal Mean ◽  
The Individual ◽  
Hydrolysis Of ◽  
Steroid Conjugates

ABSTRACT A systematic procedure for the extraction, separation, purification and colorimetric determination of urinary individual 17-ketosteroids in normal rhesus monkeys is described. Dehydroepiandrosterone was extracted after hot hydrolysis of the urine at neutral pH and the remaining steroid conjugates were extracted at pH 2 by the addition of ammonium sulphate. The liberated steroids were extracted after enzyme hydrolysis and the ketonic steroids were separated from the non-ketones by the Girard T reaction. The 11-deoxy steroids were separated from the 11-oxygenated steroids on a Celite column. They were further separated by paper chromatography and after elution, the individual 17-ketosteroids were estimated by the Zimmermann reaction. Normal mean excretion values in 30 female monkeys for total neutral 17-ketosteroids were 1.8 mg/24 h (range 1.5–2.6); total 17-hydroxycorticosteroids, 3.1 mg/24 h (range 2.2–4.8); androsterone, 38.6 μg/kg/24 h (range 9.3–60.2); aetiocholanolone, 19.2 μg/kg/24 h (range 6.8–30.7); dehydroepiandrosterone, 30.8 μg/kg/24 h (range 13.4–50.6); 11-ketoandrosterone, 7.5 μg/kg/24 h (range 5.1–14.1); 11-ketoaetiocholanolone, 10.2 μg/kg/24 h (range 6.8–18.3); 11β-hydroxyandrosterone, 15.4 μg/kg/24 h (range 8.8–26.7); 11β-hydroxyaetiocholanolone, 19.6 μg/kg/24 h (range 12.2–33.1). For total neutral 17-ketosteroids in 20 males, they were 2.5 mg/day (range 2.3–4.1); total 17-hydroxycorticosteroids, 3.7 mg/day (range 2.8–5.3); androsterone, 51.5 μg/kg/24 h (range 12.3–76.7); aetiocholanolone, 22.6 μg/kg/24 h (range 8.7–42.3); dehydroepiandrosterone, 41.3 μg/kg/24 h (range 16.9–59.7); 11-ketoandrosterone, 9.6 μg/kg/24 h (range 6.4–16.3); 11-ketoaetiocholanolone, 12.3 μg/kg/24 h (range 8.2–20.5); 11β-hydroxyandrosterone, 17.5 μg/kg/24 h (range 11.7–29.2); 11β-hydroxyaetiocholanolone, 21.7 μg/kg/24 h (range 14.5–36.4).

Sign in / Sign up

Export Citation Format

Share Document