scholarly journals Analysis of free and protein-bound nitrotyrosine in human plasma by a gas chromatography/mass spectrometry method that avoids nitration artifacts

2000 ◽  
Vol 345 (3) ◽  
pp. 453-458 ◽  
Author(s):  
Matthew T. FROST ◽  
Barry HALLIWELL ◽  
Kevin P. MOORE
Keyword(s):  
Reactive Nitrogen Species ◽  
Plasma Proteins ◽  
Biological Fluids ◽  
Plasma Concentrations ◽  
Normal Subjects ◽  
Gas Chromatography Mass Spectrometry ◽  
Highly Sensitive ◽  
Acidic Conditions ◽  
First Time

Measurement of nitrotyrosine in biological fluids and tissues is increasingly being used to monitor the production of reactive nitrogen species in vivo. The detection of nitrotyrosine in vivo has been reported with the use of a variety of methods including immunoassay, HPLC and GLC/MS. The validity of HPLC and immunoassays have been questioned with regard to their selectivity and sensitivity limits. In principle, the measurement of nitrotyrosine by GLC/MS permits a highly specific, highly sensitive and fully quantitative assay. The nitration of tyrosine under acidic conditions in the presence of nitrite is well documented. Derivatization for the full quantification of nitrotyrosine by using GLC/MS can lead to the artifactual nitration of tyrosine if performed under acidic conditions in the presence of nitrite. We describe a novel alkaline method for the hydrolysis and derivatization of nitrotyrosine and tyrosine, and demonstrate its applicability to the measurement of plasma concentrations of both free and protein-bound nitrotyrosine and tyrosine. A detection limit of 1 pg for nitrotyrosine and 100 pg for tyrosine has been achieved. Our method allows, for the first time, the analysis of free and protein-bound nitrotyrosine and tyrosine in biological samples. The plasma concentrations (means±S.E.M.) of free tyrosine and nitrotyrosine in eight normal subjects were 12±0.6 μg/ml and 14±0.7 ng/ml respectively. Plasma proteins contained tyrosine and nitrotyrosine at 60.7±1.7 μg/mg and 2.7±0.4 ng/mg respectively.

5α-Androstane-3α, 17β-diol and 5α-androstane-3α, 17β-diolglucuronide in plasma of normal children, adults and patients with idiopathic hirsutism: a mass spectrometric study

1996 ◽  
Vol 134 (1) ◽  
pp. 87-92 ◽  
Author(s):  
Stefan A Wudy ◽  
Ulrich A Wachter ◽  
Janos Homoki ◽  
Walter M Teller
Keyword(s):  
Reductase Activity ◽  
Plasma Concentrations ◽  
Mass Spectrometric Study ◽  
Normal Subjects ◽  
Mass Spectrometric ◽  
Gas Chromatography Mass Spectrometry ◽  
Spectrometric Study ◽  
Normal Children ◽  
Idiopathic Hirsutism ◽  
High Resolution Gas Chromatography

Wudy SA. Wachter UA, Homoki J, Teller WM. 5α-androstane-3α, 17β-diol and 5α-androstane-3α, 17β-diol-glucuronide in plasma of normal children, adults and patients with idiopathic hirsutism: a mass spectrometric study. Eur J Endrocrinol 1996;134:87–92. ISSN 0804–4643 We investigated the developmental patterns of 5α-androstane-3α, 17β-diol (AD) and 5α-androstane-3α, 17β-diol-glucuronide (ADG) in plasma of normal children and adults of both sexes and in patients with idiopathic hirsutism using a physicochemical method: high-resolution gas chromatography/ mass spectrometry (HRGC/MS). In children below the age of 11 years, AD and ADG increased with age showing no differences between sexes (mean ±sd nmol/l): normal subjects 3–6 years: AD in females 0.08± 0.03. in males 0.07 ±0.03; ADG in females 0.15 ±0.05, in males 0.14± 0.04: normal subjects 7–10 years: AD in females 0.17 ± 0.03, in males 0.17 ± 0.07; ADG in females 0.59 ± 0.12, in males 0.47 ± 0.14. Thereafter, AD and ADG showed a greater increase in males (normal subjects 11–15 years: AD in females 0.24 ± 0.06, in males 0.41 ±0.14: ADG in females 1.47± 0.36. in males 3.36 ± 1.22). In adults, plasma levels did not overlap between females and males (AD in females 0.24 ±0.07. in males 0.99± 0.31; ADG in females 2.32 ± 0.68, in males 13.01 ± 3.05). 5α-Androstane-3β, 17β-diol-glucuronide discriminated better between sexes than AD. In idiopathic hirsutism, mean plasma concentrations of AD and ADG were higher than those of healthy females (ages 11–15 years: AD 0.31 ± 0.10. ADG 3.48 ± 2.00; ages > 16 years: AD 0.44 ± 0.27, ADG 6.46 ± 3.11). but 54% of patients had normal plasma concentrations of AD and 29% had normal ADG values. Thus, ADG reflected androgenicity better than AD. However, both metabolites were imperfect markers of androgenicity in idiopathic hirsutism. Therefore, our findings do not support the concept of increased 5α-reductase activity in all patients with idiopathic hirsutism. Stefan A Wudy, Steroid Laboratory, First Department of Pediatrics, University of Ulm. D-89070 Ulm/Donau, Germany

scholarly journals In vivo measurement of synthesis rate of multiple plasma proteins in humans

2006 ◽  
Vol 291 (1) ◽  
pp. E190-E197 ◽  
Author(s):  
Abdul Jaleel ◽  
Vandana Nehra ◽  
Xuan-Mai T. Persson ◽  
Yves Boirie ◽  
Maureen Bigelow ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Synthesis ◽  
Stable Isotope ◽  
Plasma Proteins ◽  
Large Scale ◽  
Protein Quantification ◽  
Gas Chromatography Mass Spectrometry ◽  
Synthesis Rate ◽  
Wide Range

Advances in quantitative proteomics have facilitated the measurement of large-scale protein quantification, which represents net changes in protein synthesis and breakdown. However, measuring the rate of protein synthesis is the only way to determine the translational rate of gene transcripts. Here, we report a technique to measure the rate of incorporation of amino acids from ingested protein labeled with stable isotope into individual plasma proteins. This approach involves three steps: 1) production of stable isotope-labeled milk whey protein, oral administration of this intrinsically labeled protein, and subsequent collection of blood samples; 2) fractionation of the plasma and separation of the individual plasma proteins by a combination of anion exchange high-pressure liquid chromatography and gel electrophoresis; and 3) identification of individual plasma proteins by tandem mass spectrometry and measurement of stable isotopic enrichment of these proteins by gas chromatography-mass spectrometry. This method allowed the measurement of the fractional synthesis rate (FSR) of 29 different plasma proteins by using the same precursor pool. We noted a 30-fold difference in FSR of different plasma proteins with a wide range of physiological functions. This approach offers a tremendous opportunity to study the regulation of plasma proteins in humans in many physiological and pathological states.

scholarly journals In vivo and in vitro Volatile Constituents of the Flowers of Xylopia aromatica by HS‑SPME/GC-MS

2021 ◽  
Author(s):  
João Junqueira ◽  
Michelle do Nascimento ◽  
Lucas da Costa ◽  
Lincoln Romualdo ◽  
Francisco de Aquino ◽  
...  
Keyword(s):  
Solid Phase ◽  
Floral Scent ◽  
Principal Component ◽  
Gas Chromatography Mass Spectrometry ◽  
Large White ◽  
Volatile Constituents ◽  
Wide Range ◽  
First Time

Xylopia aromatica (Lam.) Mart. (Annonaceae) is a typical species from the Brazilian cerrado that presents medicinal properties. The plant is distinguished by its large white flowers which produce a pleasant fragrance. X. aromatica is characterized by a wide range of medicinal application. These characteristics have motivated us to investigate the flowers volatile organic compounds (VOCs) via in vivo and in vitro protocols by a headspace solid-phase microextraction (HS‑SPME) technique combined with gas chromatography-mass spectrometry (HS-SPME/GC‑MS). Four different fibers, extraction times and temperatures were the parameters changed to lead to the maximum profiling of the volatile constituents. Data were analyzed using principal component analysis (PCA). A total of 77 VOCs were extracted from the floral scent, with 52 and 68 extracted from in vivo and in vitro sampling, respectively, of which 48 were reported for the first time in the literature as volatile constituents from X. aromatica flowers. The extraction and identification of VOCs were successfully performed through HS-SPME/GC-MS. The PCA data allowed the identification of parameters that led to the maximum number of VOCs, which were polyacrylate (PA) and carboxen/polydimethylsiloxane (CAR/PDMS) fibers, 60 min extraction time and temperature of 29.0 °C. Among the volatile constituents identified, sesquiterpenes predominated, comprising about 61.04%.

Plasma pancreatic trypsinogens in chronic renal failure and after nephrectomy

1982 ◽  
Vol 242 (2) ◽  
pp. G177-G182
Author(s):  
M. C. Geokas ◽  
R. Reidelberger ◽  
M. O'Rourke ◽  
E. Passaro ◽  
C. Largman
Keyword(s):  
Renal Failure ◽  
Chronic Renal Failure ◽  
Plasma Levels ◽  
Intravenous Infusion ◽  
Plasma Proteins ◽  
Plasma Concentrations ◽  
Normal Subjects ◽  
Cationic Trypsinogen ◽  
Anephric Patients ◽  
Highly Correlated

The kidney has previously been shown to be a major site for the plasma clearance of pancreatic trypsinogens in the rat. This study investigated plasma concentrations of anionic and cationic trypsinogen in chronic renal failure and anephric patients. Plasma concentrations were significantly elevated in both groups of patients. Hemodialysis did not change their plasma levels. The plasma levels of anionic and cationic trypsinogens were highly correlated in patients and normal subjects; however, the relative concentrations of anionic trypsinogen were significantly higher in renal failure patients. This suggests that in patients with renal failure the secondary clearance mechanisms for these plasma proteins more efficiently clear cationic molecules. In normal dogs, intravenous infusion of synthetic octapeptide of cholecystokinin (CCK-8) resulted in small transitory increases in plasma trypsinogen levels. After nephrectomy, basal levels of anionic and cationic trypsinogen were elevated, and intravenous infusion of CCK-8 resulted in prolonged, high levels of plasma trypsinogens.

scholarly journals Platelet IgG, IgA, IgM, and albumin: correlation of platelet and plasma concentrations in normal subjects and in patients with ITP or dysproteinemia

Blood ◽  
1988 ◽  
Vol 72 (1) ◽  
pp. 362-365 ◽  
Author(s):  
JN George ◽  
S Saucerman
Keyword(s):  
Plasma Proteins ◽  
Plasma Concentrations ◽  
Normal Subjects ◽  
Enzyme Linked Immunosorbent Assay ◽  
Thrombocytopenic Purpura ◽  
Platelet Surface ◽  
Wide Range ◽  
Elisa Technique ◽  
Platelet Content ◽  
Monoclonal Antibody Binding

Abstract IgG, IgA, IgM, and albumin are primarily known as plasma proteins. Their presence in platelets is poorly understood. The total platelet content of IgG, IgA, and albumin, measured in solubilized platelets by an enzyme-linked immunosorbent-assay (ELISA) technique, was greater than 90% secreted after stimulation by thrombin, consistent with an alpha-granule location. The platelet concentrations of these proteins correlated with their plasma concentrations in normal subjects and over a wide range of abnormalities in patients with IgG or IgA myeloma or liver cirrhosis. IgM was not detectable in normal platelets but was measurable and related to the plasma IgM concentration in patients with macroglobulinemia. In patients with idiopathic thrombocytopenic purpura (ITP), the platelet concentrations of IgG, IgA, and albumin were all twofold to threefold higher than normal despite normal plasma concentrations. Platelet surface IgG, measured by 125I-monoclonal antibody binding, constituted less than 1% of the total platelet IgG, and it appeared to be a pool distinct from the alpha-granule IgG since its concentration in normal subjects and patients did not correlate with either plasma or total platelet IgG concentrations. These observations are consistent with hypotheses that megakaryocytes incorporate plasma proteins into developing alpha-granules by pinocytosis and that the increased ratio of platelet to plasma of IgG, IgA, and albumin in ITP may reflect a younger average age of these platelets.

In vivo evidence of cortisol secretion by aldosterone-producing adenomas

1984 ◽  
Vol 106 (4) ◽  
pp. 516-520 ◽  
Author(s):  
Kaoru Nomura ◽  
Mitsuhide Naruse ◽  
Kiyoko Naruse ◽  
Hiroshi Demura ◽  
Kazuo Shizume
Keyword(s):  
Primary Aldosteronism ◽  
Plasma Cortisol ◽  
Plasma Concentrations ◽  
Normal Subjects ◽  
Plasma Aldosterone ◽  
Adrenal Vein ◽  
Cortisol Secretion ◽  
Adrenal Vein Sampling ◽  
The Relationship

Abstract. This study was done to confirm that aldosterone-producing adenomas secrete cortisol in vivo. Plasma cortisol and aldosterone concentrations were measured in samples obtained by selective adrenal-vein sampling in 8 patients with primary aldosteronism due to unilateral adenoma. All cases revealed higher adrenal-vein plasma cortisol concentrations on the adenoma side than the opposite, irrespective of adenoma location. These concentrations correlated significantly with plasma aldosterone concentrations (r = 0.972, P < 0.001) in effluents from the adenoma side, but not from the opposite. Plasma concentrations also correlated significantly with estimated adenoma volume (r = 0.918, P < 0.05). These findings strongly suggest that aldosterone-producing adenomas secrete cortisol in vivo. In a second study, we used metyrapone to test 6 patients with adenomas. Their responsiveness to cortisol and corticotrophin was found to be the same as that in normal subjects, suggesting that adenoma-secreted cortisol did not disturb the relationship between corticotrophin and cortisol. We thus concluded that cortisol is secreted concomitantly with aldosterone from aldosterone-producing adenomas under corticotrophin influence.

scholarly journals Angiotensin (1–12) in Humans With Normal Blood Pressure and Primary Hypertension

Hypertension ◽  
2021 ◽  
Vol 77 (3) ◽  
pp. 882-890
Author(s):  
Carlos M. Ferrario ◽  
Seethalakshmi R. Iyer ◽  
John C. Burnett ◽  
Sarfaraz Ahmad ◽  
Kendra N. Wright ◽  
...  
Keyword(s):  
Plasma Concentrations ◽  
Plasma Renin ◽  
Normal Subjects ◽  
Potential Contribution ◽  
Ang Ii ◽  
Women Patients ◽  
Normal Blood Pressure ◽  
Angiotensin Ii Receptor ◽  
Normotensive Subjects ◽  
First Time

The importance of canonical versus noncanonical mechanisms for the generation of angiotensins remains a major challenge that, in part, is heavily swayed by the relative efficacy of therapies designed to inhibit renin, ACE (angiotensin-converting enzyme), or the Ang II (Angiotensin II) receptor. Ang (1–12) (angiotensin [1-12]) is an Ang II forming substrate serving as a source for Ang II–mediated tissue actions. This study identifies for the first time the presence of Ang (1–12) in the blood of 52 normal (22 women) and 19 (13 women) patients with hypertension not receiving antihypertensive medication at the time of the study. Normal subjects of comparable ages and body habitus had similar circulating plasma Ang (1–12) concentrations (women: 2.02±0.62 [SD] ng/mL; men 2.05±0.55 [SD] ng/mL, P >0.05). The higher values of plasma Ang (1–12) concentrations in hypertensive men (2.51±0.49 ng/mL, n=6) and women (2.33±0.63 [SD] ng/mL, n=13) were statistically significant ( P <0.02) and correlated with elevated plasma renin activity, systolic and pulse pressure, and plasma concentrations of NT-proBNP (N-terminal prohormone BNP). The increased plasma Ang (1–12) in patients with hypertension was not mirrored by similar changes in plasma angiotensinogen and Ang II concentrations. The first identification of an age-independent presence of Ang (1–12) in the blood of normotensive subjects and patients with hypertension, irrespective of sex, implicates this non-renin dependent substrate as a source for Ang II production in the blood and its potential contribution to the hypertensive process.

Abstract P183: Chemokine-like Receptor 1 Mediates the Vasoconstrictor Actions of Chemerin in vitro and in vivo

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Amanda Kennedy ◽  
Peiran Yang ◽  
Cai Read ◽  
Rhoda Kuc ◽  
Janet Maguire ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Smooth Muscle ◽  
Therapeutic Potential ◽  
Plasma Concentrations ◽  
Resistance Arteries ◽  
Resistance Vessels ◽  
Increased Risk ◽  
First Time

Hypertensive patients have significantly higher plasma concentrations of the adipokine chemerin compared with healthy controls, and levels of chemerin positively correlate with systolic and diastolic blood pressure. Chemerin activates chemokine-like receptor 1 (CMKLR1 or ChemR23) but it also activates the ‘orphan’ G protein-coupled receptor 1 (GPR1) which has been linked with hypertension. It is therefore crucial to determine whether one or both of these receptors mediate the constrictor actions of chemerin in the vasculature in order to identify a potential new therapeutic target for the treatment of hypertension. Using immunohistochemistry and molecular biology, we localized chemerin to the endothelium, smooth muscle and adventitia, and CMKLR1 and GPR1 to the smooth muscle in human conduit and resistance vessels. Chemerin activated β-arrestin via heterologously expressed receptors GPR1 (pD 2 =9.30±0.05) and CMKLR1 (pD 2 =9.23±0.03) with comparable potency. CCX832, a small molecule antagonist, was fully characterized as highly selective for CMKLR1, with no effect on GPR1 in binding or cell-based functional assays. The C-terminal fragment of chemerin, C9 (chemerin149-157) contracted human saphenous vein (pD 2 =7.30±0.31) and resistance arteries (pD 2 =6.23±0.16), and caused a significant increase in blood pressure in rats in vivo (0.2 μmol, 9.1±1.0 mmHg). These actions were blocked by CCX832, confirming for the first time that a single chemerin receptor, CMKLR1, mediates the constrictor response in humans and in vivo. Our data suggest that chemerin activation of CMKLR1 may contribute to elevated blood pressure; this in combination with the known roles of chemerin in metabolic syndrome and diabetes, could lead to increased risk of cardiovascular disease. This study provides proof of principle that the therapeutic potential of selective CMKLR1 antagonists should be explored.

scholarly journals Electrochemical Evaluation of the Compact and Nanotubular Oxide Layer Destruction under Ex Vivo Ti6Al4V ELI Transpedicular Screw Implantation

Materials ◽  
2020 ◽  
Vol 13 (1) ◽  
pp. 176 ◽  
Author(s):  
Katarzyna Arkusz ◽  
Marta Nycz ◽  
Ewa Paradowska
Keyword(s):  
Oxide Layer ◽  
Mechanical Stability ◽  
Ex Vivo ◽  
Electrochemical Methods ◽  
Orthopedic Implant ◽  
Transpedicular Screw ◽  
Highly Sensitive ◽  
First Time ◽  
Electrochemical Evaluation

Nano-engineered implants are a promising orthopedic implant modification enhancing bioactivity and integration. Despite the lack of destruction of an oxide layer confirmed in ex vivo and in vivo implantation, the testing of a microrupture of an anodic layer initiating immune-inflammatory reaction is still underexplored. The aim of this work was to form the compact and nanotubular oxide layer on the Ti6Al4V ELI transpedicular screws and electrochemical detection of layer microrupture after implantation ex vivo by the Magerl technique using scanning electron microscopy and highly sensitive electrochemical methods. For the first time, the obtained results showed the ability to form the homogenous nanotubular layer on an Ti6Al4V ELI screw, both in α and β-phases, with favorable morphology, i.e., 35 ÷ 50 ± 5 nm diameter, 1500 ± 100 nm height. In contrast to previous studies, microrupture and degradation of both form layers were observed using ultrasensitive electrochemical methods. Mechanical stability and corrosion protection of nanotubular layer were significantly better when compared to compact oxide layer and bare Ti6Al4V ELI.

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