A method for the isotopic determination of selenium in biological matrices is described. The method is based on hydride generation inductively coupled plasma mass spectrometry (ICP-MS). The development is specifically related to the requirements of stable isotope tracer studies in human subjects. The method is based on isotope dilution using 82Se as the in vitro spike and can quantify the 74Se and 77Se contents of samples. It involves wet oxidation (HNO3 - H2O2 or HNO3 - HClO4) of the 82Se-spiked matrix, reduction to selenite by boiling with HCl followed by measurement of the isotope ratios (82Se/77Se and 74Se/77Se) in the gas stream (H2Se) generated from on-line reduction of the sample selenite with NaBH4. Compared with the isotopic signal resulting from a selenite solution containing 5 ng ml-1 of Se, the total sample blank contributions at m/z = 74, 77 and 82 were less than 5% of the respective isotope signal. Worst-case absolute detection limits were 0.2-0.9 ng of Se, depending on the isotope used. Ion beam intensity ratios were measured with an over-all precision [relative standard deviation (RSD)] of 1% for both isotope pairs. Measured ratios (MRa/b) were stable during a given day's operation within the expected precision of the measurements but varied for different days. The magnitude of MRa/b was generally independent of the nature of the matrix. Highly linear relationships were found between ion beam intensity ratios (MRa/b) and the corresponding true isotope ratios for calibration solutions whose isotope ratios had been altered by as much as one order of magnitude.(ABSTRACT TRUNCATED AT 250 WORDS)
We examined the effect of chronic selenite supplementation on whole body and selected organ selenium (Se) accumulation, urine excretion of total Se and trimethylselenonium ion, and Se balance in adult male rats. Animals were housed in metabolic cages and given either deionized water or water containing 4 micrograms of Se/mL as selenite for 30 d. Absorption of selenite was nearly complete, with only approximately 10% of ingested Se appearing in feces. There was a rapid rise in urinary Se that reached a plateau within a few days and accounted for 54 +/- 2% of the intake. Excretion of trimethylselenonium ion (TMSe) in urine increased rapidly, representing 35-40% of urinary Se in the supplemented animals compared with only 2% for the control group. In one experiment, rats were killed at 30 d and total carcass Se was measured using isotope dilution analysis. Supplemented rats had only a modest increase in whole body Se (94 +/- 4 micrograms Se vs. 66 +/- 3 in controls). Calculation of Se balance in the supplemented rats showed that approximately 35% of ingested Se could not be accounted for by urine plus fecal losses combined with the portion retained in the carcass. The results from this study demonstrate that under the condition of supplementation at 4 micrograms of Se/mL of drinking water, pathways other than urinary and fecal excretion may account for a substantial portion of Se loss.
This study explored the quantitative relationship between the size of the selenite-exchangeable metabolic pool (WSe-EMP) and total body or liver Se in rats of varying age and past Se intake. We performed four experiments. In one, weanling rats were fed either a Se-deficient or Se-supplemented diet for 30 d, followed by measurement of WSe-EMP and total body Se. For the other experiments, rats were fed natural sources of Se without added selenite until adult age and then either subjected to acute Se restriction during the 7 d of measurements or maintained on a Se-sufficient diet. For the animals fed the selenite diet, the 7-d average ratio of WSe-EMP:total body Se (Se(end),0) was 0.370 +/- 0.009, which was not significantly different from the corresponding value (0.350 +/- 0.018, P greater than 0.05) for the Se-deficient group. When the group mean values of WSe-EMP were correlated with the corresponding mean values of Se(end),0 for all experiments, we obtained highly linear relations (r2 greater than 0.96). When WSe-EMP for each animal was correlated with the corresponding value of total body endogenous Se (Se(end)) or liver Se(end) (for t = 1 or 7 d), we found equally strong linear relations (r2 greater than or equal to 0.99). We concluded that WSe-EMP accurately reflected total body Se content or the Se content of such organs as liver, regardless of past Se intake, chemical form of Se or age and size of the animals.
The quantitative relationship between the size of the selenite-exchangeable metabolic pool (WSe-EMP) and whole body endogenous selenium (Seend) was investigated in adult male rats. Two experiments based on multiple labeling with stable isotopes were performed. One focused on short-term (7 d, Expt. 1) and the other on long-term (60 d, Expt. 2) relationships. Rats were fed a Torula yeast diet and water supplemented with [76Se]selenite at 0.1 micrograms Se/mL; the in vivo [74Se]selenite tracer was administered orally. Groups of three or four animals were killed at timed intervals and whole carcass or selected organs were analyzed for the stable isotopes 74Se, 77Se and 82Se with hydride generation/inductively coupled plasma mass spectrometry. The value of WSe-EMP was determined from plasma or urine isotope ratios. In Experiment 1, with plasma as the sampling compartment, WSe-EMP at 24 h was 36.5 +/- 1.2% of the baseline value of whole body endogenous selenium (Seend) and 36.3 +/- 1.8% at 7 d. When urine was the sampling compartment, the corresponding values were 3.9 +/- 0.3% and 43.1 +/- 2.8%, respectively. In Experiment 2, WSe-EMP (plasma) was 38.9 +/- 1.3% of Seend at 7 d, increasing to 45.5 +/- 1.6% at 60 d. The corresponding values for urine as the sampling compartment were 45.5 +/- 2.0% (7 d) and 61.5 +/- 1.7% (60 d), respectively.
Plasma Pi in early life is high and renal Pi excretion low relative t o that in adults. Renal tubular Pi transport kinetics and membrane LF was investigated in apical membrane vesicles (BBMV) in 6 fetal (F) lambs and Dregnant ewes (E). Results In t h e adult kidney, carrier mediated Pi co-transport is highly Na+ specific. Studies in our laboratory showed that Pi uptake (PiU) in renal brush border membrane vesicles (BBMV) from FL kidneys is also stimulated by Nat.In The effect of trans-stimulation was studied by preloading RBMV with 5mM Pi. Initial 12s PiU in preloaded BBMV was 1007t181 pMol/rng BRMV protein and was 4-fold higher than PiU in non-loaded BBMV (p<.001). The effect of extravesicular (0) pH was studied by measuring initial 15s PiU in BBMV with constant intravesicular pH 7.4 and varying O pH from 6.0, 7.4 and 8.0. PiU was 700264, 19832230 and 411510 pMol/mg RBMV protein for OpH 6.0, 7.4 and 8.0 respectively. PiU at pHo=pHi was higher than that for OpH 6.0 and OpH 8.0 (p<.01; p<.001).The results demonstrate Kt-Pi symport in FL kidney. The co-transport system may represent shared affinity by K+ with Na+ for the cation site on t h e carrier or a different carrier system. RENAL ALPHA ( a ) AND BETA ( 6 ) -, E f f e c t s of i n t r a r e n a l boluses of epinephrine (E) (0.00625 t o 0 . 2 ug/kg of B.W.) on r e n a l blood flow (RBF) were s t u d i e d i n c h r o n i c a l l y c a t h e t e r i z e d f e t a l (F) lambs (125-137 days; term 145 days) and a d u l t (A) sheep using a doppler flowmeter. E f f e c t s of E a l o n e and during a, a l , and 62 a d r e n e r g i c blockade were s t u d i e d . Blood pressure was unchanged during E i n f u s i o n . The E dose t h a t reduced RBF (XARBF) by 50% (ED50) was 0.0014 ug/kg/ml RBF i n F and 0.0001 ug/kg/ml RBF i n A; t h i s response was blocked by t h e a 1 a n t a g o n i s t prazosin. Following a blockade w i t h phentolamine, E produced v a s o d i l a t i o n (%A RBF-a) i n F and A which was blocked by t h e 6 2 a n t a g o n i s t butoxamine. When E was given d u r i n g a blockade a t a dose equal t o ED50 f o r v a s o c o n s t r i c t i o n , E produced a 33% r i s e i n RBF i n F and 10% i n A (p<0.05 ---23'4 P r e s e n t r e s u l t s demonstrate t h a t : 1 ) t h e E dose r e q u i r e d f o r v a s o c o n s t r i c t i o n , which i s a 1 mediated, i s 10 times g r e a t e r i n F than A, 2) t h e lower a receDtor resDonse i n F mav be secondarv PAUCITY OF MINIMAL CHANGE (MC) LESION IN YOUNG CHILDREN WITH EARLY FREQUENTLY RELAPSING STEROID SENSITIVE (FRSS) NEPHROTIC SYNDROME (NS). Kishore Phadke*, Anthony N i c a s t r i , Howard Trachtman*, Fred C a r r o l l * , C. Chen* and Amir T e j a n i , (Introduced by Laurence Finberg). Downstate Medical Center, Brooklyn, New York.The morphological l e s i o n i n young c h i l d r e n w i t h e a r l y FRSSNS i s presumed t o be MC; however, t h e r e a r e no p r o s p e c t i v e s t u d i e s t h a t v a l i d a t e t h i s a s s e r t i o n . From 1980-84, we performed a percutaneous r e n...
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