Methods were developed for determining the kininogen fractions, kininase and prekallikrein. The plasma prekallikrein was activated by 20 % (v/v) acetone for about 17 hours (20-24"). Urine kallikrein was prepared by dialysis of urine against running tap water for about 24 hours. Kininase activity was eliminated in plasma, plasma kallikrein and urine kallikrein by incubation at 37" with EDTA-2Na (1.0 X 10-2 M) for about 24 hours. Kinin assays were carried out on the isolated rat uterus. Released kinin was calculated as pg bradykininlml plasma. The total kininogen, whether determined by activation with acetone (16 % vlv for not less than 5 hours) and subsequent incubation with plasma kallikrein, by incubation with plasma kallikrein and then urine kallikrein, or by incubation with acetone (20 % vlv for 17 hours) and subsequent evaporation of the acetone, was found to be the same, 2.0 pglml plasma as an average value of 7 plasma batches corresponding to a total of 90 rats ( S . D. = 0.09). The average values of kinin released by incubation with plasma kallikrein and by urine kallikrein were 1.5 pg/ml and 1.4 pg/ml plasma respectively with S. D. values of 0.11 and 0.06 respectively. The procedures for kininase and for prekallikrein determinations corresponded closely to previously published methods for estimation of the same parameters in human plasma (RINVIK, DYRUD & BRISEID 1966; BRISEID, DYRUD & ARNTZEN 1968). TechniqueA . Materials and assays. Source of rats. Male Wistar albino rats (body weight 180-450 g) obtained from the National Institute of Public Health, Oslo, were used for the preparation of 7 batches of plasma (corresponding to batches 1-7, tables 2 and 3), and male rats (unknown strain, body weight 350-450 g) obtained from the Department of Pharmacology and Toxicology, The Veterinary College of Norway, Oslo, were used for the preparation of 2 batches of plasma (corresponding to batches 8-9, tables 2 and 3). Rat plasma.From male rats anaesthetized with ether, blood was collected from the inferior vena cava into siliconized syringes containing 1 ml of a 3.1 % sodium citrate dihydrate solution per 9 ml of blood. The pooled blood from 6 to 20 rats was used for the preparation of batches of rat plasma. The citrated blood was transferred to siliconized centrifuge tubes, and centrifuged at 1.3 X 103 g for 30 minutes at 10". The citrated plasma was stored at -20" as 1 ml samples or used at once for the preparation of kininase, kininogen, or kallikrein preparations.Rat plasma kininase. Citrated plasma was shaken with silica powder (50 mg/ml) for 30 minutes at room temperature (20-24"). The silica was removed by centrifugation and decantation, and the enzyme preparation was stored at -20" as 1 ml samples.Rat plasma kininogen. 1.00 ml citrated rat plasma was incubated for about 24 hours at 37" with 4.0 mg EDTA-2Na dissolved in 0.04 ml water. The kininogen preparation was stored at -20".Rat plasma kallikrein. For activation of prekallikrein 0.25 ml acetone per ml citrated plasma (20 % v/v of acetone) was added, and th...
Methods have been described to obtain a kininase free substrate from a turpentine-induced rat pleural exudate (24 h post-turpentine) and for the determination of kininase activity and total kininogen in such exudates. With the methods used, a mean of 7.4 min was necessary for 0.25 ml of exudate to inactivate 50% of a given amount (1 µg) of added brady-kinin. Total kininogen in exudate with leukocytes and without leukocytes were 1.97 µg/ml and 1.80 µg/ml, respectively, calculated as bradykinin. It was concluded that most of the kininogen in pleural exudate is present in the cell-free supernatant. Evidence was put forward that the rat uterus-stimulating agents that develop in the exudate upon activation with acetone are kinins not significantly contaminated with other smooth muscle-stimulating agents such as, for instance, histamine and serotonin. The conclusion was drawn that pleural exudate obtained 24 h post-turpentine in the rat is a transudate of rat plasma and that the kinin system could play a role, locally, in bringing about formation of the exudate.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.