Development and Validation of an LC/MS/MS Procedure for the Quantification of Endogenous <i>myo</i>-Inositol Concentrations in Rat Brain Tissue Homogenates

Kindt, Erick; Shum, Y. Y.; Badura, Lori L.; Snyder, Peter J.; Brant, A. W.; Fountain, Scott; Szekely-Klepser, Gabriella

doi:10.1021/ac049746w

Cited by 39 publications

(31 citation statements)

References 28 publications

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“…Several examples of the use of surrogate matrices for quantitation of exogenous and endogenous molecules have been reported. [28][29][30][31][32][33] We chose rabbit serum because initial observations indicated no interferences with either human or mouse hepcidin detection. Due to the difficulty of synthesizing human and mouse hepcidin, we were not able to use a stablelabeled internal standard that would have been preferred for development of this mass spectrometry-based assay.…”

Section: Resultsmentioning

confidence: 99%

Quantitation of hepcidin from human and mouse serum using liquid chromatography tandem mass spectrometry

Murphy¹,

Witcher

Luan

et al. 2007

Blood

149

131

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The hepatic peptide hormone hepcidin is considered the central regulator of iron metabolism. Characterizing the circulating levels of this peptide is critical to understanding its role in the development of clinically relevant syndromes, such as anemia of inflammation/chronic disease, and may provide insight into potential clinical interventions. While quantitative methods have been published for the determination of urinary hepcidin and serum prohepcidin, no definitive methods have been published for the determination of hepcidin in serum. In this report, we describe a quantitative method for the determination of both human and mouse hepcidin in serum and plasma. The method employs protein precipitation and solid-phase extraction followed by liquid chromatographic separation and tandem mass spectrometry detection. The method has a quantitative range of 0.25 ng/mL to 500 ng/mL serum for mouse hepcidin and 1 ng/mL to 500 ng/mL serum for human hepcidin. The method uses small sample volumes (50 L for mice and 100 L for humans) and 96-well formats for rapid sample processing. The method was used to establish baseline serum and plasma concentrations of hepcidin in normal C57Bl/6 mice and healthy human volunteers. (Blood.

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Section: Resultsmentioning

confidence: 99%

Quantitation of hepcidin from human and mouse serum using liquid chromatography tandem mass spectrometry

Murphy¹,

Witcher

Luan

et al. 2007

Blood

149

131

View full text Add to dashboard Cite

show abstract

“…This method was established using Kindt et al (Kindt et al, 2004) as a reference for optimization of extraction and liquid chromatography-tandem mass spectrometry (LC-MS/MS) procedures.…”

Section: Myo-inositol Quantitative Assaymentioning

confidence: 99%

Tilapia (Oreochromis mossambicus) brain cells respond to hyperosmotic challenge by inducingmyo-inositol biosynthesis

Gardell

Yang

Sacchi

et al. 2013

Journal of Experimental Biology

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SUMMARYThis study aimed to determine the regulation of the de novo myo-inositol biosynthetic (MIB) pathway in Mozambique tilapia (Oreochromis mossambicus) brain following acute (25 ppt) and chronic (30, 60 and 90 ppt) salinity acclimations. The MIB pathway plays an important role in accumulating the compatible osmolyte, myo-inositol, in cells in response to hyperosmotic challenge and consists of two enzymes, myo-inositol phosphate synthase and inositol monophosphatase. In tilapia brain, MIB enzyme transcriptional regulation was found to robustly increase in a time (acute acclimation) or dose (chronic acclimation) dependent manner. Blood plasma osmolality and Na + and Cl -concentrations were also measured and significantly increased in response to both acute and chronic salinity challenges. Interestingly, highly significant positive correlations were found between MIB enzyme mRNA and blood plasma osmolality in both acute and chronic salinity acclimations. Additionally, a mass spectrometry assay was established and used to quantify total myo-inositol concentration in tilapia brain, which closely mirrored the hyperosmotic MIB pathway induction. Thus, myo-inositol is a major compatible osmolyte that is accumulated in brain cells when exposed to acute and chronic hyperosmotic challenge. These data show that the MIB pathway is highly induced in response to environmental salinity challenge in tilapia brain and that this induction is likely prompted by increases in blood plasma osmolality. Because the MIB pathway uses glucose-6-phosphate as a substrate and large amounts of myo-inositol are being synthesized, our data also illustrate that the MIB pathway likely contributes to the high energetic demand posed by salinity challenge.

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“…The sugar phosphates and C5-polyols were detected using an MS/MS method. 2,19 The source settings (cone voltage, extraction cone, gas flow, and temperature) were optimized for the IS peak; however, the collision energy was optimized for each detected peak (15 V for the C5-polyols, 12 V for the IS, 6PG and S7P), as mentioned above.…”

Section: Chromatographic and Mass-spectrometric Optimizationmentioning

confidence: 99%

Study of transaldolase deficiency in urine samples by capillary LC‐MS/MS

et al. 2006

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Transaldolase (TAL) is a key enzyme of the pentose phosphate pathway (PPP). TAL deficiency is a newly recognized cause of liver cirrhosis. We have developed an ion-pair LC separation combined with negative ion electrospray MS/MS detection method to assess PPP metabolites in urine samples from TAL-deficient mice. Sedoheptulose 7-phosphate (S7P), C5-polyols D-arabitol and D-ribitol, and 6-phosphogluconate (6PG) levels were markedly increased in urine of TALdeficient mice with respect to those of wild-type and heterozygote littermates. The detection limits of S7P, D-arabitol, and 6PG were 0.15 ± 0.015 pmol, 3.5 ± 0.41 pmol, and 0.61 ± 0.055 pmol, respectively. The limit of quantitation was 0.4 ± 0.024 nmol/ml for S7P, 1.6 ± 0.11 nmol/ml for 6PG and 10 ± 0.7 nmol/ml for D-arabitol. Additional metabolites,

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Development and Validation of an LC/MS/MS Procedure for the Quantification of Endogenous myo-Inositol Concentrations in Rat Brain Tissue Homogenates

Cited by 39 publications

References 28 publications

Quantitation of hepcidin from human and mouse serum using liquid chromatography tandem mass spectrometry

Quantitation of hepcidin from human and mouse serum using liquid chromatography tandem mass spectrometry

Tilapia (Oreochromis mossambicus) brain cells respond to hyperosmotic challenge by inducingmyo-inositol biosynthesis

Study of transaldolase deficiency in urine samples by capillary LC‐MS/MS

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