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PubMed1955

Effect of blood thrombokinase, as influenced by soy bean trypsin inhibitor, ultracentrifugation, and accessory factors.

1. Crystallized soy bean trypsin inhibitor, at a concentration of 100 microg./ml., suppressed the production of thrombin from a mixture of prothrombin and blood thrombokinase. The experiment was performed in the presence of 0.011 M oxalate, in order to minimize the possibility of participation by accessory factors which require ionic calcium. The results are in accord with the view that thrombokinase is a trypsin-like enzyme. 2. When a solution of blood thrombokinase was centrifuged at 85,000 g for 120 minutes, almost all the activity remained in the supernate. This supernate activated the supernate from a prothrombin solution which had been similarly centrifuged. The activation of prothrombin by thrombokinase can proceed in the absence of material completely sedimentable in 120 minutes at 85,000 g. 3. An "accelerator" reagent was prepared by treating bovine serum with barium carbonate, and then passing the serum through a column of diatomaceous earth. This "accelerator" was used together with prothrombin, blood thrombokinase, Howell's cephalin, and calcium chloride to compose a five-reagent thrombin-producing system. In this system, no thrombin was produced without thrombokinase. On the other hand, thrombin was produced from prothrombin and thrombokinase, even when all the other reagents were omitted. When calcium was omitted, thrombokinase was able to function; but cephalin and the "accelerator" reagent were ineffective. 4. Quantitative tests indicated that the "accelerator" reagent exerted an effect distinct from those of thrombokinase and cephalin. However, it is not certain whether the "accelerator" reagent functioned as an accessory factor, as a potential source of more thrombokinase, or both. In the experiments reported, thrombokinase was primary to, or necessary for, the effect of "accelerator." 5. The effectiveness of thrombokinase was multiplied a hundred times or more, when complemented by calcium, cephalin, and "accelerator" reagent. Ionic calcium was a necessary component of this complementing system. This may help to explain why removal of calcium ions keeps blood fluid, even though thrombokinase, by itself, is little influenced either by calcium ions or by oxalate.

Animals↗
PubMed1987

Monoclonal IgM with lupus anticoagulant activity in a case of Waldenström's macroglobulinaemia.

Strong lupus anticoagulant activity was detected in the plasma of a 73-yr-old woman with Waldenström's macroglobulinaemia. Purified patient IgM caused significant prolongation of the APTT in concentrations as low as 30 micrograms/ml, and its activity was inhibited by intact or (more effectively) by frozen-thawed blood platelets. Employing varying concentrations of cephalin in the APTT assay, it was demonstrated that the macroglobulin had a direct cephalin-neutralizing effect. When tested in an ELISA system using phospholipid-coated microtiter wells, the IgM was shown to possess antibody activity against cephalin. With purified phospholipids as antigens, the protein was found to display antibody activity against phosphatidyl serine and phosphatidyl ethanolamine, which are main constituents of cephalin. These observations were confirmed in absorption experiments.

Aged↗
PubMed1990

Quantitation of anticephalin antibodies in a computer-assisted enzyme-linked immunosorbent assay (ELISA): relation to lupus anticoagulant.

Lupus anticoagulants (LA) are IgG or IgM antibodies which prolong phospholipid-dependent coagulation tests. For the detection and quantitation of such antibodies, we have developed an ELISA with cephalin as the coating antigen. The sensitivity of this assay was compared to the activated partial thromboplastin time (APTT). LA was defined as greater than or equal to 5 sec prolongation of the APTT with standard cephalin dilution, or greater than or equal to 10 sec prolongation with a high cephalin dilution, on a 1:1 mixture of patient and control plasma. Plasma samples from 158 healthy individuals were tested for anticephalin antibodies. The 97.5 percentile was chosen as the upper reference limit and allocated a value of 1 ELISA unit. A "four-parameter logistic" model was used for transformation of the absorbances to ELISA units. Of 314 plasma samples referred for LA screening, positive results were found in 62 by both APTT and ELISA. Twenty-three samples were ELISA positive and APTT negative; this finding may be explained by greater sensitivity of the ELISA, which gave positive results in a four-fold greater dilution than the APTT. Prolongation of the APTT without antibody activity was found in 8 samples of which 2 had an inhibitor of factor VIII:C, the remaining 6 probably had true LA. In conclusion, our computer-assisted ELISA is a sensitive and reliable test method for quantitation of anticephalin antibodies. This assay has a high concordance with LA as detected with the APTT.

Adult↗
PubMed1977

[Chronology of the ultrastructural modifications during the growth of Gregarina blaberae].

In an ultrastructural study the development of the sporozoite as well as the growth and development of the trophozoite of Gregarina blaberae were followed in the course of experimental infections of larvae of the cokroach Blaberus craniifer. The spectacular growth involved the transformation within 18 days of the sporozoite, measuring 15 X 1 micronm, to a cephaline--trophozoite affixed to the intestinal epithelium--of 250 micronm length and 65 micronm diameter. The sporozoite's ultrastructure is not different from that of sporozoites of other Sporozoa studies to date--the conoid and dense bodies are present. The pellicle consists of 3 membranes, but there are some interruptions in the internal membrane complex. The first dictyosomes are formed from the nuclear envelope. The migration of the nucleus and of the dense bodies, followed by the regression of all the structures characteristic of the sporozoites, and the establishment of a cortical zone that comes to cover the epimerite, take place within 48 h after infection and mark the transformation of the sporozoite into the trophozoite. Development of the cephaline involves the formation of the epicytic folds, which occurs at the base of the deutomerite, starting on the 3rd day of development. A regular system of longitudinal or epicytic folds is formed over the entire surface of the gregarine. On the 4th and 5th days of development, a vacuolar system and a chondriome become differentiated in the epimerite, while a fibrillar septum separates the protomerite from the deutomerite. The next stage, starting on the 6th day, is characterized by distribution of polysaccharide reserves between these 2 segments. The model studied allows us to determine the role of the epimerite in the parasite's nutrition, as well as the development of the chondriome and of the cortical membranes in the course of the vegetative growth phase of the cephaline gregarine.

Amylopectin↗
PubMed1990

The significance of high molecular weight kininogen for contact activation of rat blood coagulation, in vitro.

The involvement of the high molecular weight rat kininogen in the activation of the rat contact system by kaolin-cephalin, kaolin, sulfatides and ellagic acid has been investigated, using a rat plasma congenitally devoid of this kininogen. Coagulation times induced by these activators were shorter in normal as well as in deficient rat plasma than in normal human plasma. Coagulation times were prolonged in deficient rat plasma, when the incubation times was three min or less. By kaolin or cephalin-kaolin, this prolongation disappeared when the incubation time reached ten min. The activation of plasma prekallikrein developed slowly in deficient plasma with all the triggers but reached control level after ten min of incubation. By kaolin-cephalin, the activation of Hageman factor was weak and slow in deficient plasma during the ten min of incubation. In rat, high molecular weight kininogen plays thus a role in the activation of the contact system by these triggers. But this role seems to be less important than in human plasma.

Animals↗
PubMed1975

[Relations between the physico-chemical properties, the chemical reactivity and the local-anesthetic activity/part 33: studies on the interactions of local-anesthetically active cinchocaine homologues with phospholipids (author's transl)].

The general part consists of a review on nerve stimulus mechanism as well as the nerve structure and the function of nerve membrane as the site of action of local anesthetics. Furthermore a possibility of interaction between local anesthetics and the membrane components especially phospholipids is discussed. These phospholipids show interesting properties with respect to ions as well as local-anesthetics, which allow us to suppose that they are important participants in nervous stimulus transmission. In the experimental part two methods are described. The first deals with the measurement of electrical resistance as function of time at cephalin and cholesterin impregnated filter membrane in solutions of cinchocain homologues. An increase in the resistance with the increase in concentration of test substances was observed. The same was the effect of increasing chain length in alkoxy group where after butoxy derivative, a deformation of membrane was observed. In the second method the drug binding capacity of cephalin dispersed in aqueous medium was measured. Here, too, the increase in the binding capacity with the increase in alkoxy chain was observed. The large difference in free binding energy between two subsequent homologues is explained as the effect of increase in van der Waals' forces and hydrophobic interactions on one hand, and a change in colloidal form of cephalin dispersion on the other hand.

Absorption↗
PubMed1979

[Effect of phospholipase D on micellar phospholipids and the role of calcium ions].

The rates of the reaction products formation under simultaneous phospholipase D effect on phosphatidyl ethanolamine and phosphatidyl choline were studied. The hydrolysis of cephalin, unlike the phospholipase D effect on lecithin, does not require Ca2+ ions. Ca2+ does not affect the enzymatic degradation of lecithin and inhibits the reaction with cephalin in "inorganized" phospholipid emulsions. The hydrolysis of micellar phospholipids by phospholipase D (in the presence of the anionic detergent sodium dodecyl sulfate) is accelerated by Ca2+ ions for both substrates. The apparent Km value is equal to 1.5 mM and does not depend on the phospholipid type. In contrast, the value of kcat for lecithin is twice as high as that for cephalin. It was demonstrated that the phase state of the phospholipids and the chemical nature of the alcohol residue in the phospholipid molecule are essential for the substrate specificity of phospholipase D.

Calcium↗
PubMed1966

New, partially hydrolyzable synthetic analogues of lecithin, phosphatidyl ethanolamine, and phosphatidic acid.

The synthesis of two new synthetic analogues of lecithin, two of phosphatidyl ethanolamine ("cephalin"), and one new phosphatidic acid analogue is described. They comprise one of each of the following types: the "isosteric" diether lecithin and cephalin analogues ROCH(2)CH(OR)- CH(2)CH(2)P(O) (O(-))OCH(2)CH(2)N(+)R'(3) (R = C(18)H(37); R' = H or CH(3)); and the "hydrocarbon" analogues of phosphatidic acid, lecithin, and cephalin, C(17)H(35)CH(2)CH(C(18)H(37))CH(2)P(O)(R) = (R'); [R = R' = OH; R = O(-), R' = OCH(2)CH(2)N(+)(CH(3))(3); and R = O(-), R' = OCH(2)CH(2)N(+)H(3)]. Infrared spectra and other properties of these compounds are described.

Chemical Phenomena↗
PubMed1984

The inhibition of the anticoagulant activity of heparin by platelets, brain phospholipids, and tissue factor.

Platelets and phospholipids have been shown to protect factor Xa from inhibition by the heparin--antithrombin III complex. The studies reported herein investigated the effects of gel filtered platelets, activated platelets, brain phospholipids (cephalin), and brain tissue factor on the inactivation of thrombin and factor Xa by the heparin--antithrombin III complex. In addition, the relative anticoagulant effects of heparin on the extrinsic and intrinsic coagulation pathways were investigated. Our results suggest that gel filtered platelets, activated platelets, cephalin and tissue factor protect thrombin, as well as factor Xa, from inactivation by the heparin--antithrombin III complex. Tissue factor had the greatest anti-heparin activity. Activated platelets, gel filtered platelets, cephalin and tissue factor did not alter the protease--antithrombin III reaction rates measured in the absence of heparin. These observations are consistent with the hypothesis that platelets, brain phospholipids, and tissue factor, in the presence of calcium, partition heparin from antithrombin III, and thus prevent full expression of the antithrombin III-dependent anticoagulant activity of heparin.

Antithrombin III↗
PubMed1983

A family with heterozygous factor X Friuli defect outside Friuli.

Three members of the same family were found to have a clotting defect consistent with the diagnosis of heterozygous factor X Friuli disorder. The main features of the defect were a mild prolongation of prothrombin time and partial thromboplastin time, but a normal Stypven-Cephalin clotting time. Factor X activity was 40-50% of normal using tissue thromboplastin, but was perfectly normal using Russell's viper venom and cephalin. Using chromogenic substrate S-2222 the level was 30% of normal. Immunologically, factor X was normal. Bleeding manifestations were mild if any. The hereditary pattern was autosomal. The family comes from an area far away from Friuli and represents the first example of factor X Friuli discovered outside the Friuli.

Adult↗
PubMed1976

[About a substance with antithrombinic activity, found in the latero-oesophageal glands of Meinertia oestroides (Risso, 1826) (Isopoda, Flabellifera, Cymothoidae; parasite of fishes) (author's transl)].

An anticlotting factor was found recently in the cephalothoracic latero-oesophageal glands (G.L.O.) of Cymothoid Isopods (blood-sucking fishparasites). This substance is efficient on the blood of hosts such as Boops boops (L.), and also on the human plasma. The way of action and the chemical kind of this factor were investigated: Plasmatic coagulation-tests (Howell, Quick, Cephalin-kaolin, Stypven, Stypven-cephalin, Thrombin and Thrombin coagulase times) were realized on citrated plasmas of Boops and man; enzymatic activities of latero-esophageal glands and hepatopancreas of Meinertia oestroides (Risso, 1826) were also studied. Our results show that the anticlotting action of G.L.O. extracts proceeds from an anti-thrombinic factor, certainly carbohydrate-like as heparin.

Animals↗
PubMed1988

Phase diagrams of pseudo-binary phospholipid systems. I. Influence of the chain length differences on the miscibility properties of cephaline/cephaline/water systems.

The miscibility properties of homologous cephalines (PEs) were studied by means of differential scanning calorimetry (DSC). The phase diagrams of 5 pseudo-binary cephaline/cephaline/water systems (50% water) are discussed. In the high temperature L alpha-phase, all the homologous cephalines of fatty acid chain length from C = 12 to C = 18 were completely miscible. On the other hand in the low temperature L beta-phase, a miscibility gap occurred in dependence on the differences of the acyl chain lengths. Further, a distinct succession of the phase diagram types was observed according to increasing chain length differences of the PEs: complete miscibility (systems di-(C12:O)-PE/di-(C14:O)-PE/H2O; di-(C14:O)-PE/di-C(16:O)-PE/H2O)----peritectic mixing behaviour (systems di-(C12:O)-PE/di-(C16:O)-PE/H2O; di-(C14:O)-PE/di-(C18:O)-PE/H2O)----eutectic mixing behaviour (system di-(C12:O)-PE/di-(C18:O)-PE/H2O). The change in the type of phase diagram from azeotropic to semi-azeotropic and from semi-azeotropic to eutectic is interpreted by means of the Landau theory.

Calorimetry, Differential Scanning↗
PubMed1988

Effects of varying dietary proteins on plasma lipids and rabbit platelet function.

The aim of this paper was to study the effect of dietary proteins on rabbit platelet aggregation induced by ADP, collagen and thrombin. In addition, blood coagulation tests (Stypven, Stypven cephalin and cephalin kaolin plasma clotting times) were performed on these rabbits fed semi-synthetic diets containing animal protein (casein) or vegetable proteins (walnut or soybean). As part of this study, total cholesterol, HDL cholesterol, phospholipid and triglyceride plasma levels were determined and platelet fatty acid composition was analysed. The results show that the addition of walnut meal in the diet decreases platelet aggregability and total plasma cholesterol. The relative effects of casein, soybean and walnut protein on platelet aggregability best correlated with the ratio of lysine to arginine in the protein and the ratio of HDL to LDL. An increase in the ratio C20:5 n-3/C20:4 n-6 in platelet fatty acids was also seen in walnut and soybean protein-fed animals, but the changes were greatest in soybean protein-fed animals while inhibition of platelet aggregation was greatest in walnut protein-fed animals.

Animals↗
PubMed1990

[Mechanisms and prediction of hemorrhagic complications during surgery of thoraco-abdominal aortic aneurysms].

This study, including 33 consecutive patients was designed to assess the haemostatic alterations occurring during repair of thoracoabdominal aneurysms. The surgical procedure consisted in Dacron graft replacement of the diseased aorta, using neither cardiopulmonary bypass, nor any shunting technique, nor any heparin. Blood samples were drawn before anaesthesia, before and 30 min after unclamping, and on the first postoperative day. The measured parameters were: haematocrit, platelet count, bleeding, activated cephalin, thrombin and prothrombin times, and concentrations of fibrinogen, factors V, VII, X and II, anti-thrombin III, proteins C and S, fibrin degradation products, D-dimers, alpha 2-antiplasmin, plasminogen, tissue plasminogen activator, plasminogen activator inhibitor, and serum protein. Eight patients developed severe multiple haemorrhages; 3 of them died during the procedure because of uncontrollable bleeding. Although the measured parameters were similar in the "bleeding" and control (n = 25) groups before surgery, there was, before unclamping in the first group, an important increase in activated cephalin and thrombin times, with a fall in concentrations of factor II and V, protein C, fibrinogen, and alpha 2-antiplasmin, and in platelet numbers. After unclamping, these changes worsened further, with an increase in prothrombin time and in fibrinogen levels (0,8 g.l-1), without any increase in fibrin degradation products. Abnormal bleeding started about 30 min after this in all the patients of the "bleeding" group. These changes, involving the fibrinolytic system as well as a fall in concentration of all the coagulation factors, can probably be partly explained by the clamping and unclamping of mesenteric vessels.(ABSTRACT TRUNCATED AT 250 WORDS)

Aortic Aneurysm↗
PubMed1975

Diol lipids of rat liver. Quantitation and structural characteristics of neutral lipids and phospholipids derived from ethanediol, propanediols, and butanediols.

Specific enzymatic and chemical degradation of neutral lipid and phospholipid fractions from rat liver revealed the presence of novel types of lipid metabolites bearing a short-chain diol backbone. Diol-derived lecithin and cephalin analogs were readily cleaved by phospholipase C (EC 3.1.4.3) from Bacillus cereus, although the cephalin analogs required "carrier" lecithin to sustain hydrolysis. The products of phosphilipase hydrolyses as well as the neutral lipid fractions were subjected to alkaline and acidic methanolysis, and constituent short-chain diols were analyzed as long-chain cyclic acetals. Gas chromatographymass spectrometry confirmed that 1,2-ethanediol, 1,2-propanediol, 1,3-propanediol, and 1,3 butanediol can form the polyol backbone of neutral lipids and phospholipids. [1,1,2,2-2H]Ethanediol monohexadecanoate, dihexadecanoate, hexadecanoylphosphorylcholine, hexadecanoylphosphorylethanolamine were synthesized chemically and served as internal standards to assure accurate quantitation of the low levels of diol lipids (350 mug/g ot total lipid) present in rat liver.

Animals↗
PubMed2001

Solid-phase extraction followed by liquid chromatography-mass spectrometry for trace determination of beta-lactam antibiotics in bovine milk.

A confirmatory assay able to unambiguously identify and quantify 10 approved-for-use beta-lactam antibiotics in milk below stipulated U.S. and EU tolerance levels is presented. beta-Lactams are extracted from 10 mL of intact milk by a Carbograph 4 cartridge. After solvent removal, residue reconstitution, and filtration, a completely transparent and uncolored extract is injected into a liquid chromatography -mass spectrometry (LC-MS) instrument equipped with an electrospray (ES) ion source and a single quadrupole. During the chromatographic run, the ES/MS system is operated first in the positive-ion mode (PI) and then in the negative-ion (NI) mode. This is done to circumvent matrix interferences resulting in remarkable signal weakening of the last-eluted analytes, when detecting them as [M+H]+ adduct ions. MS data acquisition is performed by a time-scheduled three-ion selected ion monitoring program. At the 5 ng/mL level, recoveries of the beta-lactams are between 70 (nafcillin) and 108% (cephalin), with relative standard deviations ranging between 5 (oxacillin) and 11% (amoxicillin and ceftiofur). The response of the ES/MS detector is linearly related to injected amounts up to 500 ng, irrespective of the chemical characteristics of the beta-lactams and the acquisition mode selected (PI or NI modes). Limits of quantification, based on a minimal value of the signal-to-noise ratio of 10, were estimated to be within 0.4 (cephalin) and 3 ng/mL (dicloxacillin). Analyses of milk samples taken after intramammary application of amoxicillin showed that 1.2 ng/mL of this penicillin was still present 6 days after treatment. At this concentration level, the identification power of the method is not weakened, as signals of the three product ions of amoxicillin are still well distinguishable from the background noise.

Animals↗
PubMed1990

Cytochrome c oxidase and cardiolipin alterations in response to skeletal muscle ischaemia and reperfusion.

The effect of 2 and 4 h of tourniquet ischaemia followed by 1 h of reperfusion on the major mitochondrial phospholipids and on the cytochrome c oxidase kinetic parameters has been investigated in rat skeletal muscle. There was no change either in the mitochondrial phospholipid content or in the Vmax and the Km of the enzyme after 2 h of ischaemia with and without subsequent reperfusion. Four hours of ischaemia had no effect on the lecithin and the cephalin content, while the cardiolipin content decreased as well as the Vmax of the enzyme (P less than 0.05). Tissue reperfusion caused a dramatic decrease in both cardiolipin (55% of the control, P less than 0.001) and Vmax (38% of the control, P less than 0.001). The corresponding reduction in lecithin and cephalin contents was 12% and 14% respectively (P less than 0.05). The Km remained unchanged at all conditions. These findings suggest that mitochondrial dysfunction in response to ischaemia and reperfusion could be a consequence of the reperfusion itself following severe ischaemia. The results are discussed in terms of cardiolipin peroxidation and cytochrome oxidase as a functional parameter.

Animals↗
PubMed1962

Toxicity of blood clotting factors.

Pure bovine thrombin was separated from autoprothrombin C, and the lethal intravenous dose for mice weighing 25 g was 0.4 ml of a solution containing 50 units of thrombin per milliliter. Autoprothrombin C was not toxic alone, but with crude cephalin it was fatal. The clotting time of human plasma was only slightly accelerated by autoprothrombin C alone; the clotting time was as short as 5 seconds with a combination of autoprothrombin C and cephalin.

Animals↗