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A rapid and sensitive high-performance liquid chromatographic procedure is described for the simultaneous determination of piroxicam (CAS 36322-90-4) and tenoxicam (CAS 59804-37-4) in blood, plasma and buffer solutions used in the equilibrium dialysis studies. The method can be used to measure either piroxicam and tenoxicam in these three fluids using the other as internal standard. A Nucleosil C18 and a mobile phase consisting of an acetonitrile-distilled water-acetic acid (58 : 38 : 4) mixture were used. The flow rate was 1 ml/min and the effluent was monitored at 365 nm with 0.02 AUFS (absorbance units full scale). The sensitivities of this method were 0.2 microgram/ml levels of piroxicam and tenoxicam in the plasma, blood and buffer solutions samples.
Concentration of (total) globulin was relatively stable in blood plasma of mares, but rapidly decreased in colostrum to very low levels within 2 days after parturition. In foals, after intake of the first colostrum, globulin increased within 1 day in blood plasma, but remained at lower concentrations than those measured in mare plasma. Concentrations of immunoreactive insulin (iI) were high during the first 2 months of lactation in blood plasma of mares and then decreased, were high in first colostrum and then decreased drastically, and remained at low concentrations up to weaning in blood of foals. In mares, concentration of immunoreactive insulin-like growth factor I (iIGF-I) in plasma increased during late pregnancy, peaked 2 days after parturition, and then gradually declined until weaning. iIGF-I was highest in first colostrum and then dramatically decreased within the first 2 days of lactation. In foals, iIGF-I gradually increased over the first 2 months of life. IGF-I in the horse appears to be bound to proteins of similar molecular weight as in cattle. The study demonstrates different patterns of changes in plasma iIGF-I and iI concentrations in mares and their foals, whereas iI, iIGF-I and globulin changes in colostrum and milk occurred in parallel. Furthermore, plasma iI and iIGF-I behaved differently, while colostrum iI and iIGF-I behaved similarly, in mares compared with dairy cows.
There are conflicting results regarding the activity of glutathione (GSH) in patients with kidney disease. We examined whole blood-, plasma- and erythrocyte concentrations of GSH and cysteine in 12 patients with renal disease and in 14 patients on maintenance hemodialysis. Total plasma GSH concentrations were higher in patients with kidney disease than in patients maintained on hemodialysis (23.5 +/- 13.1 vs 9.1 +/- 7.5 mumol/l, p < 0.01) but erythrocyte concentrations of GSH were similar. Whole blood-, plasma and erythrocyte concentrations of GSH remained stable during hemodialysis. The total concentration of cysteine in plasma was significantly higher in patients on hemodialysis (556 +/- 258 mumol/l) as compared to patients with kidney disease (248 +/- 53 mumol/l, p < 0.001) but red blood cell concentrations were similar. Whole blood- and plasma concentrations of cysteine decreased significantly during hemodialysis. There were no significant correlations between serum creatinine and GSH or cysteine concentrations in patients on hemodialysis but when all patients were evaluated together, a significant correlation between serum creatinine and total plasma cysteine concentrations was observed (r = 0.52, p = 0.0017).
Murine blood plasma rapidly inactivates nitrosamides and nitrosocarbamates but not nitrosoureas. The mechanism of this inactivation in murine blood plasma has been investigated. The vast majority of activity (greater than 97%) was inhibited by serine hydroxylase inhibitors. Also, 92% of the activity was inhibited by bis(p-nitrophenyl)phosphate, a selective inhibitor of carboxylesterases. Decomposition products formed after blood plasma action on N-ethyl-N-nitrosoacetamide or N-methyl-N-nitrosoethylcarbamate were separated and identified by gas chromatography. The products formed were consistent with a hydrolytic cleavage of the amidic bond. These observations are consistent with the idea that the major active factor(s) in plasma is a carboxylesterase(s).
The phospholipid composition of blood plasma and erythrocytes from artery, hepatic venous, and sagittal sinus blood was investigated in dogs under pentobarbital anesthesia both before and after continuous infusion of norepinephrine. Norepinephrine infusion caused a very significant increase of phosphatidylglycerol (GPG) in the erythrocytes and plasma from arterial blood. Other phospholipids were not affected by the infusion. Significant release of GPG across the brain and splanchnic area was noted both in plasma and erythrocytes prior to, but not following, norepinephrine infusion. The importance of the differences between the respective phospholipid compositions of erythrocytes and plasma is emphasized. The biologic significance of the changes of the phospholipids, particularly of GPG during stress, is discussed.
Human blood plasma contains low-molecular substances that inhibit in a dose-dependent manner both high-affinity specific binding of imipramine and reverse serotonin uptake by platelets. Incubation of human blood plasma with alumina was made use of to extract and study these imipramine-like inhibitors. The extract obtained from human blood plasma inhibited imipramine binding and reverse uptake of serotonin with median inhibitory concentrations of 0.18 +/- 0.1 and 0.36 +/- 0.15 mg/ml, respectively. After gel chromatography on Biogel P-2 the elution profile of the extract showed 2 major peaks of reverse serotonin uptake and imipramine binding inhibition and 3 additional peaks of reverse serotonin uptake inhibition, which did not have any considerable effect on imipramine specific binding. It is assumed that endogenous inhibitors of imipramine binding and reverse serotonin uptake are involved in the development of affective disorders.
In the Polish Navy, deep-water dives, performed for the needs of the maritime industry, are conducted using our own national technology and trimix as a breathing medium. In this paper are presented the results obtained during a short-time deep-water diving test using the principles of US Navy technology, combined with the use of diving equipment type AF-2 and heliox-type breathing mixture in the open circuit. In the performed examinations changes in clinical parameters were assessed viz.: blood morphology, hematocrit level, concentration of C3c, C4, IgG, IgA, IgM, CRP, concentration of fibrinogen and factor XII level, obtained 30 minutes prior to commencement, immediately after completion, and 24 hours after termination of the exposure. The results thus generated were subjected to a preliminary analysis by the description of trends observed. It was revealed that the diving technology employed did not generate substantial changes in the examined parameters of blood in divers, and the increase of neutrophils, blood platelets and fibrinogen concentration in the blood plasma immediately after diving is of temporary character, being a typical reaction observed during diving.
Plasma levels of interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) were determined in healthy individuals and patients with aplastic anemia (ApAn). IFN-gamma was not detected in normal peripheral blood plasma (PBP) or bone marrow plasma (BMP) and was present in PBP from only 2 of 22 patients and in BMP from 1 of 14 patients and the levels were low (< 1.5 U/ml). Elevated levels of TNF-alpha were present in BMP and PBP from patients but not in control (healthy donor) PBP and BMP. Eleven of twenty-four patients had elevated levels of TNF-alpha in their PBP and 6 of 13 patients had detectable levels of TNF-alpha in their BMP. Only one of the 14 healthy control donors had detectable TNF-alpha and the level was very low (7 pg/ml), while 13 of the 27 ApAn patients had detectable TNF-alpha (P = .009, chi-square test). Not surprisingly, the centers of the distributions of TNF-alpha concentrations of the controls and ApAn patients differed significantly (P < .017 for control and patient PBP and P < .056 for control and patient BMP, Wilcoxon rank-sum test). Spontaneous production of IFN-gamma and TNF-alpha by cultured bone marrow mononuclear cells was observed in four of seven patients but not in the six healthy controls (P = 0.026). Spontaneous production of IFN-gamma and TNF-alpha by cultured peripheral blood mononuclear cells from patients and controls was however similar. Phytohemagglutinin (PHA)-induced production of IFN-gamma and TNF-alpha by cultured mononuclear cells did not differ significantly between ApAn patients and normal controls. The significance of overproduction of TNF-alpha in the pathophysiology of ApAn is discussed.
The blood plasma cholesterol (CS) level increased by 170% due CS of atherogenic lipoproteins (LP) in dogs that received atherogenic rations including lamb fat (LF) (group 1) during 2 months. In dogs of group 2 that received atherogenic rations including sunflower oil (SFO), the total content of CS in the blood plasma rose only by 50%, the portion of high density LP (HDLP) decreased from 73 to 25% in both groups of animals. LF and SFO produced no effect on the plasma CS level, when the dogs did not receive excessive CS in the ration (groups 3 and 4). SFO induced reduction in the rate of the lecitin-cholesterolacyltransferase (LCAT) reaction, both in normal and excessive CS consumption. Nevertheless, the ratio free CS/CS esters was lowered that denoted the appearance in the blood of CS esters, the nature of which was not associated with the LCAT reaction. In the animals of groups 1, 2 and 3 the share of oleic acid in phosphatidylcholine of HDLP increased (in group 3 this increase was most manifest), while the portion of stearic acid significantly diminished.
The feasibility of using plasma, blood and haemoglobin adducts for monitoring occupational exposure to the suspected human carcinogen 4,4'-methylenebis(2-chloroaniline) (MOCA) was investigated. A method utilising capillary gas chromatography-negative-ion chemical-ionisation mass spectrometry (GC-MS) for the determination of pentafluoropropionyl (PFP) derivatives of MOCA, released by alkaline hydrolysis from protein adducts and conjugates, was both sensitive and selective. When selected ion monitoring was used, sub-femtomole amounts of PFP-MOCA could be measured. The detection limit for haemoglobin adducts of MOCA was below 10 fmol/g Hb, well below the levels found for occupationally exposed individuals. Capillary GC with electron-capture detection also had the required sensitivity for the determination of MOCA in blood samples, however, the presence of interfering compounds in some samples limited its use. The levels of MOCA in the blood and urine of five individuals who were exposed to MOCA during the manufacture of polyurethane elastomers were determined by the GC-MS method. The MOCA concentrations for the various blood fractions and urine were within the following ranges: haemoglobin adducts, 0.73-43.3 pmol MOCA/g Hb; plasma alkaline hydrolysate, 0.05-22.0 nmol/l; whole blood, 0.13-17.4 nmol/l; urine, 4.5-2390 nmol/l. Because the products of MOCA in the blood reflect metabolic activation of MOCA and integrate exposure over a period of weeks, the use of blood samples for monitoring exposure to MOCA offers advantages over the currently used urinary MOCA measurements.
Oncogene mutations are frequently found in several tumour types and, among these, point mutations of the ras gene are particularly significant. A predominance of N-ras mutations has been found in the bone marrow DNA of patients with myelodysplastic syndrome (MDS) or acute myelogenous leukaemia (AML). On the other hand, increased levels of plasma DNA have previously been observed in patients suffering from various malignant diseases. In the present work we have investigated, by polymerase chain reaction (PCR), point mutations of the N-ras gene in the DNA of plasma, blood cells and bone marrow of 10 patients suffering from AML or MDS. The different ras mutations detected in five cases were always present in the plasma DNA while sometimes absent in the DNA of peripheral blood cells or bone marrow. This indicates that a bone marrow biopsy or aspiration does not necessarily contain all the malignant clones involved in the disease. Plasma could thus prove to be an easily accessible and useful material for detection and monitoring of myeloid disorders.
Fifty-two patients with chronic hepatitis C virus (HCV) infection were treated with standard doses of interferon alfa-2b. During treatment, HCV RNA detection was studied in samples of whole blood (WB), plasma (Pl), and peripheral blood mononuclear cells (PBMCs). Individuals were classified as sustained responders (SRs), complete responders with relapse (CRs), partial responders (PRs), or nonresponders (NRs) according to normalization of serum alanine transaminase (ALT) during treatment and follow-up. Before treatment, 100% of WB samples and more than 95% of Pl and PBMC samples were positive for HCV RNA. During treatment, there was progressive clearance of HCV RNA from Pl and PBMCs in SRs and CRs, but CRs had significantly more positive WB samples during and following treatment (P <.0001). At 6 months, only 10% of CR patients were positive by Pl assay, but 50% were positive by WB assay (P <.01). In the PR group, all WB samples remained positive throughout treatment, although 25% to 40% of PBMC and Pl samples became negative for HCV RNA during the first 2 months of therapy (WB > Pl or PBMC; P < .001). However, at later times during treatment most Pl and PBMC samples in the PR group were positive. Samples from the NR group showed no clearance of HCV RNA from WB, Pl, or PBMC fractions. These data document the increased sensitivity of WB assays for detecting HCV RNA in the peripheral blood of patients during interferon therapy. Furthermore, our findings suggest that WB analysis of HCV RNA may be a useful parameter to monitor in determining the end point of interferon therapy.
We examined the spontaneous and metal-ion-catalyzed oxidative modification plasma blood proteins in the group of healthy adults and elderly ones and patients with vascular dementia (mild and severe). We determined the spectrophotometric measurement of 2,4-dinitrophenylhydrazone derivatives formed by reactions with protein carbonyls. The level of metal-ion-catalyzed oxidation proteins in the aged patients both with and without dementia was high in comparison to the healthy adults. The patients with severe dementia showed lower amount of 2,4-dinitrophenylhydrazone deriviates. Low levels of metal-ion-catalysed protein oxidation strongly correlated with the degree of psychoorganic disturbances. The elderly persons with both and without dementia showed a high level of plasma nonenzymatic H2O2 scavenging in comparison with the healthy adult ones. We discovered an imbalance between enzymatic and nonenzymatic components of the antioxidant system. The latter indicates that the oxidative modification of brain tissue proteins probably plays an important role in aging and mental disorders.
A high-performance liquid chromatographic method was developed to measure the concentration of brodimoprim and its metabolite, hydroxybrodimoprim, in small volumes of blood, plasma and urine. The procedure involved a simple extraction step with chloroform, followed by chromatographic separation on a short reversed-phase column deactivated for the analysis of basic compounds. The column effluent was monitored by fluorescence (excitation wavelength 290 nm, emission wavelength 340 nm). The recoveries of both compounds were similar in all three biological fluids, and averaged 84 and 72%, respectively. The detection limit for both compounds reached 5 ng/ml. No endogenous compound interfered in the assay. The linearity of the method and its within- and between-day precision were analytically satisfactory.
BACKGROUND: The working group "Blood Plasma Constituents" of the DGTI has carried out a multicenter study with human immunoglobulins for intravenous administration (IVIG) and a NIBSC serum, with the designation 67/98C, since the European Pharmacopoeia IVIG monograph gives no information for the IgG subclass determination. Aim of the ring study was 1) the determination of the IgG subclass distribution with a standardized method and 2) the determination of the IgG subclass distribution in the NIBSC serum 67/98C and to test if the NIBSC serum coded 67/98C is suited as a reference serum for the IgG subclass determination. MATERIAL AND METHODS: Ten laboratories from Europe participated in the ring study. The IgG subclasses were determined by radial immunodiffusion (RID). RESULTS: The results showed good consistency between the participating laboratories and showed distinct differences in the IgG subclass composition of the tested IVIGs. The reproducibility of the method showed coefficients of variation of approximately 10%. CONCLUSION: The NIBSC serum 67/98C is suitable as reference serum for the IgG subclass determination in sera and IVIGs.