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- Kushnir, Mark M., et al.
(författare)
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Beta-methylamino-L-alanine analysis by liquid chromatography tandem mass spectrometry with iTRAQ as the derivative
- 2009
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Ingår i: European journal of mass spectrometry. - : SAGE Publications. - 1469-0667 .- 1751-6838. ; 15:3, s. 439-443
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Tidskriftsartikel (refereegranskat)abstract
- Amino acid BMMA is produced by cyanobacteria and has been linked to the development of neurodegenerative diseases. We developed a method for quantitative analysis of BMAA in biological samples and plant extracts. The method is utilizing iTRAQ and LC-MS/MS detection using multiple reaction monitoring mode. The method uses 50 pL of sample and has a Limit of quantitation of 300 ng mL(-1), within-run run imprecision below 1%. Using this method we analyzed human serum samples, human cerebrospinal fluid samples and extract of the cycad seed. No BMAA could be detected in the human samples. Content of BMAA in the seed was 50 mg kg(-1).
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- Kushnir, Mark M., et al.
(författare)
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High-Sensitivity Tandem Mass Spectrometry Assay for Serum Estrone and Estradiol
- 2008
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Ingår i: American Journal of Clinical Pathology. - 0002-9173 .- 1943-7722. ; 129:4, s. 530-539
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Tidskriftsartikel (refereegranskat)abstract
- High-sensitivity measurement of serum estrogens is important in adult and pediatric endocrinology and oncology. We developed a high-sensitivity liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for simultaneous measurement of estrone (E-1) and estradiol (E-2). Aliquots of 200 mu L of serum were spiked with internal standard, extracted, derivatized with dansyl chloride, and analyzed by LC-MS/MS using 2-dimensional chromatographic separation. Total imprecision for the method was less than 11%; the limit of quantitation was 1 pg/mL. Reference intervals were established with samples from more than 900 healthy postmenopausal women, men, girls, and boys. Concentrations of estrogens in children reached adult levels by Tanner stage 3. In men and postmenopausal women, the median concentrations of total estrogens (E-1 + E-2) were 39 and 22 pg/mL, and the median E-2/E-1 ratios were 0.98 and 0.55, respectively. The method requires a small sample volume and has adequate sensitivity and specificity for analyzing estrogens in samples from postmenopausal women, men, and children.
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- Kushnir, Mark M., 1957-
(författare)
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Mass Spectrometric Applications for Diagnosing Metabolic and Endocrine Diseases
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Disease-specific compounds (biomarkers) are analyzed in clinical laboratories to assist with diagnosing diseases. This thesis describes development and validation of liquid chromatography tandem mass spectrometry (LC-MS/MS) based tests for diagnosing a diverse group of endocrine and metabolic diseases. The analytical methods used on-line and off-line sample extraction and analytical derivatization as means of enhancing the analytical sensitivity, specificity and clinical utility. All developed methods were extensively validated and reference intervals for the biomarker concentrations were established in blood samples of healthy adults and children. Advantages of the LC-MS/MS as an analytical technique include possibility of simultaneous measurement of multiple analytes and ability of confirming their identity. In this thesis we proposed and evaluated approaches for the assessment of the specificity of analysis in the methods that use tandem mass spectrometry detection. To enhance throughput of the LC-MS/MS tests for the biomarkers that have endogenous or exogenous isomers an approach was developed for quantitation of isomers from unresolved chromatographic peaks. Using methods developed in this thesis we performed a study of the steroidogenesis in ovarian follicles of healthy women and women with polycystic ovary syndrome (PCOS). Obtained data on the steroid concentrations and associations between the steroid metabolites in the pathway would be helpful for better understanding of the ovarian pathophysiology. Potential biomarkers of PCOS were identified in the thesis; further studies will be necessary to confirm their clinical utility.
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- Kushnir, Mark M, et al.
(författare)
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Steroid profiles in ovarian follicular fluid from regularly menstruating women and women after ovarian stimulation
- 2009
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Ingår i: Clinical Chemistry. - : Oxford University Press (OUP). - 0009-9147 .- 1530-8561. ; 55:3, s. 519-526
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Information on the concentrations of steroids in ovarian follicular fluid (FF) from regularly menstruating (RM) women has been limited because of the absence of methods for the simultaneous quantification of multiple steroids in small volumes of FF. We studied steroid profiles in FF during the early follicular phase of the menstrual cycle and after ovarian stimulation for in vitro fertilization (IVF), and compared concentrations with published values obtained by immunoassay (IA). METHODS: We used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to measure 13 steroids in 40-microL aliquots of FF samples from 21 RM women and from 5 women after ovarian stimulation for IVF. Relationships between concentrations of steroids and their ratios (representations of the enzyme activities) were evaluated within and between subgroups. RESULTS: The concentrations of testosterone (Te), androstenedione (A4), and estradiol (E2) measured by LC-MS/MS were lower than those previously reported in studies with IAs. In RM women, androgens were the most abundant class of steroids, with A4 being the major constituent. The concentrations of 17-hydroxyprogesterone (17OHP), total androgens, and estrogens were 200- to 1000-fold greater in FF than in serum. Compared with RM women, FF samples from women undergoing ovarian stimulation had significantly higher concentrations of E2 (P = 0.021), pregnenolone (P = 0.0022), 17OHP (P = 0.0007), and cortisol (F) (P = 0.0016), and significantly higher ratios of F to cortisone (P = 0.0006), E2 to estrone (P = 0.0008), and E2 to Te (P = 0.0013). CONCLUSIONS: The data provide the first MS-based concentration values for 13 steroids in ovarian FF from RM women, from estrogen- and androgen-dominant follicles, and from women after ovarian stimulation for IVF.
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