9 1992 by The Humana Press, Inc. All rights of any nature, whatsoever, reserved. 0163-4984/92/3301-3--4)135$02.00
Metabolic Differences and Similarities of Selenium in Blood and Brain of the Rat Following the Administration of Different Selenium Compounds WANG ZI-JIAN,* ZHOU JIE, AND PENG AN Research Centre For Eco-Environmental Sciences, Academia Sinica, PO Box 2871, Beijing, PR China 100085 Received May 31, 1991; Accepted August 2, 1991
ABSTRACT A common intermediate, i.e., selenite, was found in the serum of the rat; the maximum levels occurred 3 h after administration independent of chemical forms. This indicates that both the reduction of selenate to selenite, and oxidation of seleno-dl-methionine to selenite existed in the metabolic pathways of the rat. We found that water-soluble selenium compounds led to a similar maximum content in blood and serum, but seleno-dl-methionine had a higher affinity for the brain and, by gel filtration chromatography, for the higher mol-wt (25-100 K Da) fractions of serum protein, when compared with inorganic forms. Index Entries: Selenium compounds; selenite; rat; metabolism.
INTRODUCTION Many authors have reported on the similarities and differences of different chemical forms in metabolic pathways of selenium, such as absorption (1), retention (2), and excretion (3). Previous research has dealt also with the metabolic transformation of different selenium corn*Author to whom all correspondence and reprint requests should be addressed. Biological Trace Element Research
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pounds, which emphasized the central role of selenite, but also indicated the characteristic pathways of each chemical form (4). Comparison of metabolic differences was mostly carried out by 75Se radioisotope, which was limited by available labeled chemicals. In the biological selenium cycle, the reduction from selenite to selenide and the methylation process are well documented (5). The reduction of selenate to selenite in biological system is less well investigated. Similarly, it is known that certain bacteria can oxidize elemental selenium to selenite and to traces of selenate (5), but the biological oxidation of selenide, as in seleno-methionine, to selenite, has not been studied in detail. Moreover, although metabolic similarities between selenium and sulfur have been reported by many authors, analogies between sulfite and selenite production have not been considered. The purpose of this article is to show that there exists a common pathway of different selenium compounds in the rat, although their incorporation into biological components differs significantly.
/vLA.TERIALSAND METHODS Chemicals and Instruments Seleno-dl-methionine (Se-met) was obtained from Sigma Company, Germany. Seleno-yeast (Se-yeast) was obtained from Huazhong University of Science and Technology. Selenite (Se[IV]) and selenate (Se[VI]) were obtained from Beijing Chemical Company. Other chemicals used were analytical grade. Selenium Determination Trace amounts of selenium were determined by an HPLC-FID technique developed in this laboratory. The total amount was determined after being wet digested in an HC103-HNO 3 mixture and derivatized with 2,3-diaminonaphthalene (DAN). Determination of selenite was performed through direct derivatization in acidic solution, based on the selective reaction between selenite and 2,3-diaminonaphthalene. The derivative product was extracted into cyclohexane. The selenium containing fluorescent product piazselenol (NSD) was separated by normalphase liquid chromatography, and was characterized by retention time and cochromatography with a synthesized standard sample of NSD. A detailed experimental procedure for speciation study was described in a previous paper (6).
Animals and Experimental Procedures Wistar rats (170 _+ 30 g) were fed a normal diet throughout the experiment (0.112 mg Se/kg), and maintained at 20-22~ and 50--70% of humidity. Selenite, selenate, seleno-dI-methionine, and seleno-yeast Biological Trace Element Research
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were prepared in a solution of 25 mg Se/L (or suspension in case of seleno-yeast). At zero time, animals were grouped into four and were orally administered 0.5 mg Se/kg body wt of each chemical. The blood of three rats in each group was drawn 1, 3, 6, 12, and 24 h away from orbit, and the rats were then sacrificed to obtain the whole brain. In three rats, only distilled water was administered to serve as a control group. From these animals, blood was drawn after 12 h, and the brains were obtained as mentioned above. For selenium determination, 0.2 mL of whole blood and 0.5-1.0 g of brain were used. The rest of the blood was centrifugated, and two portions of 0.2 mL each of serum were used immediately for total selenium and selenite determination. The rest of the serum from each group at the indicated time was mixed and stored at 0~ before sampling into a gel filtration column.
Gel Fgtration Chromatography Sephadex G-200 (Pharmacia Ltd., Switzerland) column (55 x 1.5 cm) was equilibrated with a 0.05M Na2HPO4-0.1M NaC1-0.01% NaN3 mixture (pH 6.8). The column was calibrated by mol-wt kit, and 0.5 mL of serum was injected into the column through a sample applicator SA-5 (Pharmacia Fine Chemical Co.). The eluates from the column were monitored by a Waters model 481 LC spectrophotometer for protein at 280 nm, and the fractioned eluates were collected for selenium determinations.
RESULTS AND DISCUSSIONS Time-Course The time-course of selenium content in serum (Table 1) and blood (Table 2) of the rat following an oral administration of different selenium compounds shows a rise from 1--6 h and a decline from 6-24 h. This significant increase and rapid equilibration between serum and blood cells in the experimental period with respect to the 12-h control group indicate a specific process of absorption. Selenium reached maximum levels in the time period of 3--6 h, depending on chemical forms. No significant difference (P > 0.05) in the maximum levels could be found among selenite, selenate, and seleno-methionine, suggesting that these three compounds are equally bioavailable for absorption. The results agree with previous studies of the absorption of selenite and selenomethionine; both chemicals were found to be equally well absorbed, i.e., 92 and 91%, respectively (2). However, the maximum level was reached earlier in the case of seleno-dl-methionine than with the other selenium compounds studied, suggesting a different absorption mechanism (7). The maximum level in the case of seleno-yeast was lower than with the Biological Trace Element Research
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138
Table 1 Time-Course of Selenium Content in Serum of the Rat Following an Oral Administration of Different Selenium Compounds
Selenium content, mg Se/L Time, h 1 3 6 12 24
Se (IV) 0.36 0.72 0.94 0.53 0.40
_+ 0.05 + 0.03 _+ 0.10 -4- 0.02 + 0.10
Se-met 0.56 0.95 0.90 0.49 0.45
_ 0.07 + 0.03 -+ 0.10 _+ 0.05 + 0.02
Se (VI) 0.40 0.85 1.08 0.53 0.46
+ -+ + + -+
0.08 0.06 0.21 0.07 0.03
Se-yeast 0.38 0.45 0.58 0.33 0.40
+ 0.06 _+ 0.07 + 0.01 + 0.07 _+ 0.04
Control 0.26 + 0.06 -
Table 2 Time-Course of Selenium Content in Blood of the Rat Following an Oral Administration of Different Selenium Compounds Selenium content, mg Se/L Time, h 1 3 6 12 24
Se (IV) 0.33 0.55 0.52 0.46 0.45
+ 0.05 + 0.02 + 0.05 - 0.02 _+ 0.05
Se-met 0.48 0.64 0.59 0.48 0.40
_+ 0.01 _+ 0.11 + 0.08 + 0.04 _+ 0.01
Se (VI) 0.45 0.55 0.69 0.50 0.47
-+ -+ -+ + +
0.18 0.07 0.08 0.07 0.00
Se-yeast 0.38 0.36 0.44 0.35 0.32
-+ + _+ -+ +
0.05 0.04 0.01 0.03 0.01
Control 0.27 ___ 0.08 -
o t h e r chemical f o r m s s t u d i e d , p r e s u m a b l y b e c a u s e of its insolubility in water. The increase of the s e l e n i u m c o n t e n t in b l o o d a n d s e r u m of the rat after a d m i n i s t r a t i o n c o u l d be o w i n g either to d e l a y e d a b s o r p t i o n or the release of the e l e m e n t f r o m the liver back into p l a s m a (8). C o m p a r i n g Table 1 a n d Table 2, it m a y be s e e n that this increase of s e l e n i u m in b l o o d is m a i n l y the result of the increase of the e l e m e n t in the s e r u m , i n d e p e n d e n t of the chemical f o r m s a d m i n i s t e r e d .
Selenium Content in the Brain The brain is o n e of the priority p a t h w a y s of s e l e n i u m a b s o r p t i o n u n d e r c o n d i t i o n s of i n a d e q u a t e intake (9). S e l e n i u m c o n t e n t s in the brain after a d m i n i s t r a t i o n of different s e l e n i u m c o m p o u n d s are given in Table 3. It m a y be s e e n that seleno-dl-methionine has a higher affinity for the brain t h a n the inorganic f o r m s of selenium. The result s h o w s that differe n c e s in the m e c h a n i s m of a b s o r p t i o n exist b e t w e e n selenite, selenate, a n d s e l e n o - m e t h i o n i n e . S e l e n o - m e t h i o n i n e b e h a v e d like an active f o r m of s e l e n i u m for i n c o r p o r a t i o n into brain tissue and, therefore, c o u l d s e r v e as a b e t t e r chemical for S e - s u p p l e m e n t a t i o n t h a n the inorganic forms. O n the o t h e r h a n d , the latter could be m o r e r a p i d l y bioavailable t h a n selenomethionine.
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Table 3 Time-Course of Selenium Content in Brain of the Rat Following an Oral Administration of Different Selenium Compounds Time, h 1 3 6 12 24
Selenium content, mg Se/kg Se-met Se (VI) Se-yeast
Se (IV) 0.15 0.21 0.23 0.22 0.20
_+ 0.01 _+ 0.02 _+ 0.02 ___ 0.02 + 0.01
0.34 0.37 0.30 0.29 0.26
_+ 0.01 + 0.02 + 0.03 ___ 0.01 _+ 0.04
0.15 0.20 0.18 0.24 0.22
_+ 0.03 + 0.04 _+ 0.01 + 0.04 + 0.02
0.17 0.21 0.19 0.18 0.18
_+ 0.05 ___ 0.11 + 0.02 _+ 0.02 ___ 0.02
Control 0.15 _+ 0.02 -
Selenite lVletabolite in the Serum Selenite could be detected as a metabolite in the serum of the rat administered different selenium c o m p o u n d s (Table 4). The concentration of selenite determined includes free ions in the serum and labile selenium attached to proteins at pH 2. The maximum level of selenite in the serum was reached in 3 h and declined, suggesting that the oxidation reduction of selenium in the rat is a rapid process. The concentrations of selenite in the serum decreased in the order selenate > selenite seleno-d/-methionine > seleno-yeast. The results suggest that both reduction of selenate to selenite and oxidation of seleno-methionine and seleno-yeast to selenite are possible in the rat. Earlier work has s h o w n evidence for the formation of adenosine-5'-phosphoselenate, which suggests that sulfate and selenate utilize the same e n z y m e system (10) for activation. A p a t h w a y for the conversion of selenite and seleno-cysteine to selenide has been discussed (11), and it was also suggested that either h y d r o g e n selenide or a reduction product of selenodiglutathione could be a m o n g the selenium products released from the erythrocyte (12,13). Similar pathways have been suggested in sulfur and selenium metabolism (14). Therefore, a c o m m o n pathway similar to sulfur might exist, w h e r e both selenite and selenide could be the important intermediates for the synthesis of seleno-cysteine. Our results suggest that the selenite metabolite could be p r o d u c e d either through reduction of selenate or oxidative degradation of organo-selenium c o m p o u n d s from diet.
Gel F~ltratJon Figure 1 shows the mol-wt distribution of selenium-bound protein components in serum of the rat after administration of selenite, selenate, and seleno-methionine in 1, 3, 6, and 24 h, respectively. Recovery of selenium in these experiments is 70-140% (data not given). Four protein peaks could be differentiated in the chromatogram (A280, not illustrated) as generally reported by using the same technique. Increased selenium contents were found in all protein classes separated by gel-filtration in 3
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Zi-jian, die, a n d A n
Table 4 Selenite Metabolite in S e r u m of the Rat Following a n Oral A d m i n i s t r a t i o n of Different Selenium C o m p o u n d s Selenite content, ~g/L Time, h
Se (IV) 3.7 20.6 6.2 4.6 3.4
1 3
6 12 24
Se-met
_+ 1.9 _+ 7.5 +_ 1.7 + 1.2 __+ 2.1
Se (VI)
4.0 + 1.2 11.0 _+ 1.1 4.9 + 0.4 1.0 _ 0.2 N.D.*
11.2 28.4 14.5 12.0 10.3
_+ +_ _+ + _+
10.6 9.9 4.7 1.1 2.2
Se-yeast
Control
4.0 -+ 2.0 4.6 --_ 0.8 1.8 _+_ 0.4 0.5 ---0.4 N.D.
-
N.D. -
*N.D. < 0.2 ~g/L.
f, f,
f~
f+ Fractions
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~ c
~ e
M. Vl. C a l i h r a l i o n
jw.,, 1 A 6h
z z
z
~
3h
gELENATE
SELENITE
SELENO-METHIONINE
Fig. I. Chromatographic time-course of selenium in serum of the rat following an oral administration of selenite, selenate, and seleno-dl-methionine. Molecular-weight calibration used Pharmacia Fine Chemical Co. kit, Switzerland; a--blue dextran 2000 (mol wt 2,000,000); b~bovine serum albumin (tool wt 67,000); c--ovalbumin (mol wt 43,000); d--ribonudease (tool wt 13,700); e~sodium fluorescein (mol wt 376). Fractions were divided into four parts as in Fig. 2; F , - - m o l w t 100-2000 kDa; F2--mol w t 25-100 kDa; F3--mol w t 10-25 kDa; F 4 - - m o l w t < 10 kDa.
Biological Trace Element Research
Vol. 33, 1992
IVletabolic Differences of Se Compounds A
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Biological Trace Element Research
VoL 33, 1992
142
Zi-jian, die, and An
and 6 h (Fig. 2), similar to the time-course of selenium content in serum, but the incorporation of selenium into protein components was different among the chemical forms applied. This is clearly demonstrated by Fig. 2, where the total eluent was divided into four parts (F1 to F4), each corresponding to a protein class as indicated in the figure captions. Relative accumulation was expressed for each of these four parts as the ratio of selenium in the Se-supplemented group to that in the control group (12 h). Figure 2 shows that the relative selenium accumulation of different selenium compounds in high-mol-wt fractions (F1, mol-wt 100-2000 kDa) reached 100% in 24 h, indicating that F1 represents only a transient carrier. Seleno-methionine tends to be accumulated in F2 (mol wt 25-100 kDa), whereas selenite and selenate tend to be accumulated in F 3 (tool wt 10-25 kDa) in 24 h, suggesting different incorporations into serum protein. Within 24 h, the accumulation and evolution patterns of the different selenium species in F 2 and F 4 ( < 1 0 kDa) became quite similar, suggesting that F2 is possibly interrelated with F4 for the recycling of low-mol-wt selenium components between erythrocyte and serum (15). The calculated percentage of selenite in F4 is ~29% of the total, indicating that other species, such as selenide, seleno-diglutathione, and so forth, are present.
ACKNOWLEDGMENT The authors gratefully acknowledge Professor H. D. Shu and Dr. L. P. Liang from Beijing Medical University for animal care. Thanks are also owing to Professor G. N. Schrauzer for kindly correcting the manuscript. This work was supported by the Chinese National Foundation.
REFERENCES 1. C. D. Thomson, R. D. H. Stewart, and M. F. Robinson, Br. J. Nutr. 33, 45-54 (1975). 2. C. D. Thomson and R. D. H. Stewart, Br. J. Nutr. 30, 139-147 (1973). 3. M. Richold, M. F. Robinson, and R. D. H. Stewart, Br. J. Nutr. 38, 19-29 (1977). 4. R. F. Burk, Trace Elements in Human Health and Disease, II, Essential and Toxic Elements, A. S. Prasad, ed., Academic, New York, 1976, pp. 105-133. 5. WHO, Environmental Health Criteria 58, Selenium, published under the joint sponsorship of UNEP, ILO and WHO, Geneva, 1987, pp. 41-43. 6. Z. J. Wang and A. Peng, J. Environm. Sci. (China) 1, 116-121 (1989) (Eng.). 7. K. P. McConnell and G. J. Cho, Am. J. Physiol. 208, 1191-1195 (1965). 8. H. W. Symonds, B. F. Sansom, D. L. Mather, and M. J. Vagg, Br. J. Nutr. 45, 117-120 (1981). 9. D. Behne, H. Hilmert, S. Scheid, H. Gessner, and W. Elger, Biochim. Biophysi. Acta 966, 12-21 (1988).
Biological Trace Element Research
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/vletabolic Differences of Se Compounds
143
10. I. Rosenfield and O. A. Beath, eds. Selenium Geobatany, Biochemistry, Toxicity and Nutrition, Academic, New York, 1964, pp. 335--365. 11. R. F. Burk and P. E. Gregory, Arch. Biochem. Biophys. 213, 73-80 (1982). 12. M. A. Motsenbocker and A. L. Tappel, Biochim. Biophys. Acta, 704, 253 (1982). 13. M. A. Beilstein and P. H. Whanger, J. Nutr. 113, 2138 (1983). 14. O. A. Levander, Trace Elements in Human Health and Disease, II, Essential and Toxic Elements, A. S. Prasad, ed., Academic, New York, 1976, pp. 135-163. 15. W. B. Davidson and C. H. Mcmurray, J. Inorg. Biochem. 34, 1-9 (1988).
Biological Trace Element Research
Vol. 33, 1992
Metabolic Differences and Similarities of Selenium in Blood and Brain of the Rat Following the Administration of Different Selenium Compounds WANG ZI-JIAN,* ZHOU JIE, AND PENG AN Research Centre For Eco-Environmental Sciences, Academia Sinica, PO Box 2871, Beijing, PR China 100085 Received May 31, 1991; Accepted August 2, 1991
ABSTRACT A common intermediate, i.e., selenite, was found in the serum of the rat; the maximum levels occurred 3 h after administration independent of chemical forms. This indicates that both the reduction of selenate to selenite, and oxidation of seleno-dl-methionine to selenite existed in the metabolic pathways of the rat. We found that water-soluble selenium compounds led to a similar maximum content in blood and serum, but seleno-dl-methionine had a higher affinity for the brain and, by gel filtration chromatography, for the higher mol-wt (25-100 K Da) fractions of serum protein, when compared with inorganic forms. Index Entries: Selenium compounds; selenite; rat; metabolism.
INTRODUCTION Many authors have reported on the similarities and differences of different chemical forms in metabolic pathways of selenium, such as absorption (1), retention (2), and excretion (3). Previous research has dealt also with the metabolic transformation of different selenium corn*Author to whom all correspondence and reprint requests should be addressed. Biological Trace Element Research
] 35
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136
Zi-jian, Jie, and An
pounds, which emphasized the central role of selenite, but also indicated the characteristic pathways of each chemical form (4). Comparison of metabolic differences was mostly carried out by 75Se radioisotope, which was limited by available labeled chemicals. In the biological selenium cycle, the reduction from selenite to selenide and the methylation process are well documented (5). The reduction of selenate to selenite in biological system is less well investigated. Similarly, it is known that certain bacteria can oxidize elemental selenium to selenite and to traces of selenate (5), but the biological oxidation of selenide, as in seleno-methionine, to selenite, has not been studied in detail. Moreover, although metabolic similarities between selenium and sulfur have been reported by many authors, analogies between sulfite and selenite production have not been considered. The purpose of this article is to show that there exists a common pathway of different selenium compounds in the rat, although their incorporation into biological components differs significantly.
/vLA.TERIALSAND METHODS Chemicals and Instruments Seleno-dl-methionine (Se-met) was obtained from Sigma Company, Germany. Seleno-yeast (Se-yeast) was obtained from Huazhong University of Science and Technology. Selenite (Se[IV]) and selenate (Se[VI]) were obtained from Beijing Chemical Company. Other chemicals used were analytical grade. Selenium Determination Trace amounts of selenium were determined by an HPLC-FID technique developed in this laboratory. The total amount was determined after being wet digested in an HC103-HNO 3 mixture and derivatized with 2,3-diaminonaphthalene (DAN). Determination of selenite was performed through direct derivatization in acidic solution, based on the selective reaction between selenite and 2,3-diaminonaphthalene. The derivative product was extracted into cyclohexane. The selenium containing fluorescent product piazselenol (NSD) was separated by normalphase liquid chromatography, and was characterized by retention time and cochromatography with a synthesized standard sample of NSD. A detailed experimental procedure for speciation study was described in a previous paper (6).
Animals and Experimental Procedures Wistar rats (170 _+ 30 g) were fed a normal diet throughout the experiment (0.112 mg Se/kg), and maintained at 20-22~ and 50--70% of humidity. Selenite, selenate, seleno-dI-methionine, and seleno-yeast Biological Trace Element Research
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Metabolic Differences of Se Compounds
137
were prepared in a solution of 25 mg Se/L (or suspension in case of seleno-yeast). At zero time, animals were grouped into four and were orally administered 0.5 mg Se/kg body wt of each chemical. The blood of three rats in each group was drawn 1, 3, 6, 12, and 24 h away from orbit, and the rats were then sacrificed to obtain the whole brain. In three rats, only distilled water was administered to serve as a control group. From these animals, blood was drawn after 12 h, and the brains were obtained as mentioned above. For selenium determination, 0.2 mL of whole blood and 0.5-1.0 g of brain were used. The rest of the blood was centrifugated, and two portions of 0.2 mL each of serum were used immediately for total selenium and selenite determination. The rest of the serum from each group at the indicated time was mixed and stored at 0~ before sampling into a gel filtration column.
Gel Fgtration Chromatography Sephadex G-200 (Pharmacia Ltd., Switzerland) column (55 x 1.5 cm) was equilibrated with a 0.05M Na2HPO4-0.1M NaC1-0.01% NaN3 mixture (pH 6.8). The column was calibrated by mol-wt kit, and 0.5 mL of serum was injected into the column through a sample applicator SA-5 (Pharmacia Fine Chemical Co.). The eluates from the column were monitored by a Waters model 481 LC spectrophotometer for protein at 280 nm, and the fractioned eluates were collected for selenium determinations.
RESULTS AND DISCUSSIONS Time-Course The time-course of selenium content in serum (Table 1) and blood (Table 2) of the rat following an oral administration of different selenium compounds shows a rise from 1--6 h and a decline from 6-24 h. This significant increase and rapid equilibration between serum and blood cells in the experimental period with respect to the 12-h control group indicate a specific process of absorption. Selenium reached maximum levels in the time period of 3--6 h, depending on chemical forms. No significant difference (P > 0.05) in the maximum levels could be found among selenite, selenate, and seleno-methionine, suggesting that these three compounds are equally bioavailable for absorption. The results agree with previous studies of the absorption of selenite and selenomethionine; both chemicals were found to be equally well absorbed, i.e., 92 and 91%, respectively (2). However, the maximum level was reached earlier in the case of seleno-dl-methionine than with the other selenium compounds studied, suggesting a different absorption mechanism (7). The maximum level in the case of seleno-yeast was lower than with the Biological Trace Element Research
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Zi-jian, Jie, and An
138
Table 1 Time-Course of Selenium Content in Serum of the Rat Following an Oral Administration of Different Selenium Compounds
Selenium content, mg Se/L Time, h 1 3 6 12 24
Se (IV) 0.36 0.72 0.94 0.53 0.40
_+ 0.05 + 0.03 _+ 0.10 -4- 0.02 + 0.10
Se-met 0.56 0.95 0.90 0.49 0.45
_ 0.07 + 0.03 -+ 0.10 _+ 0.05 + 0.02
Se (VI) 0.40 0.85 1.08 0.53 0.46
+ -+ + + -+
0.08 0.06 0.21 0.07 0.03
Se-yeast 0.38 0.45 0.58 0.33 0.40
+ 0.06 _+ 0.07 + 0.01 + 0.07 _+ 0.04
Control 0.26 + 0.06 -
Table 2 Time-Course of Selenium Content in Blood of the Rat Following an Oral Administration of Different Selenium Compounds Selenium content, mg Se/L Time, h 1 3 6 12 24
Se (IV) 0.33 0.55 0.52 0.46 0.45
+ 0.05 + 0.02 + 0.05 - 0.02 _+ 0.05
Se-met 0.48 0.64 0.59 0.48 0.40
_+ 0.01 _+ 0.11 + 0.08 + 0.04 _+ 0.01
Se (VI) 0.45 0.55 0.69 0.50 0.47
-+ -+ -+ + +
0.18 0.07 0.08 0.07 0.00
Se-yeast 0.38 0.36 0.44 0.35 0.32
-+ + _+ -+ +
0.05 0.04 0.01 0.03 0.01
Control 0.27 ___ 0.08 -
o t h e r chemical f o r m s s t u d i e d , p r e s u m a b l y b e c a u s e of its insolubility in water. The increase of the s e l e n i u m c o n t e n t in b l o o d a n d s e r u m of the rat after a d m i n i s t r a t i o n c o u l d be o w i n g either to d e l a y e d a b s o r p t i o n or the release of the e l e m e n t f r o m the liver back into p l a s m a (8). C o m p a r i n g Table 1 a n d Table 2, it m a y be s e e n that this increase of s e l e n i u m in b l o o d is m a i n l y the result of the increase of the e l e m e n t in the s e r u m , i n d e p e n d e n t of the chemical f o r m s a d m i n i s t e r e d .
Selenium Content in the Brain The brain is o n e of the priority p a t h w a y s of s e l e n i u m a b s o r p t i o n u n d e r c o n d i t i o n s of i n a d e q u a t e intake (9). S e l e n i u m c o n t e n t s in the brain after a d m i n i s t r a t i o n of different s e l e n i u m c o m p o u n d s are given in Table 3. It m a y be s e e n that seleno-dl-methionine has a higher affinity for the brain t h a n the inorganic f o r m s of selenium. The result s h o w s that differe n c e s in the m e c h a n i s m of a b s o r p t i o n exist b e t w e e n selenite, selenate, a n d s e l e n o - m e t h i o n i n e . S e l e n o - m e t h i o n i n e b e h a v e d like an active f o r m of s e l e n i u m for i n c o r p o r a t i o n into brain tissue and, therefore, c o u l d s e r v e as a b e t t e r chemical for S e - s u p p l e m e n t a t i o n t h a n the inorganic forms. O n the o t h e r h a n d , the latter could be m o r e r a p i d l y bioavailable t h a n selenomethionine.
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Table 3 Time-Course of Selenium Content in Brain of the Rat Following an Oral Administration of Different Selenium Compounds Time, h 1 3 6 12 24
Selenium content, mg Se/kg Se-met Se (VI) Se-yeast
Se (IV) 0.15 0.21 0.23 0.22 0.20
_+ 0.01 _+ 0.02 _+ 0.02 ___ 0.02 + 0.01
0.34 0.37 0.30 0.29 0.26
_+ 0.01 + 0.02 + 0.03 ___ 0.01 _+ 0.04
0.15 0.20 0.18 0.24 0.22
_+ 0.03 + 0.04 _+ 0.01 + 0.04 + 0.02
0.17 0.21 0.19 0.18 0.18
_+ 0.05 ___ 0.11 + 0.02 _+ 0.02 ___ 0.02
Control 0.15 _+ 0.02 -
Selenite lVletabolite in the Serum Selenite could be detected as a metabolite in the serum of the rat administered different selenium c o m p o u n d s (Table 4). The concentration of selenite determined includes free ions in the serum and labile selenium attached to proteins at pH 2. The maximum level of selenite in the serum was reached in 3 h and declined, suggesting that the oxidation reduction of selenium in the rat is a rapid process. The concentrations of selenite in the serum decreased in the order selenate > selenite seleno-d/-methionine > seleno-yeast. The results suggest that both reduction of selenate to selenite and oxidation of seleno-methionine and seleno-yeast to selenite are possible in the rat. Earlier work has s h o w n evidence for the formation of adenosine-5'-phosphoselenate, which suggests that sulfate and selenate utilize the same e n z y m e system (10) for activation. A p a t h w a y for the conversion of selenite and seleno-cysteine to selenide has been discussed (11), and it was also suggested that either h y d r o g e n selenide or a reduction product of selenodiglutathione could be a m o n g the selenium products released from the erythrocyte (12,13). Similar pathways have been suggested in sulfur and selenium metabolism (14). Therefore, a c o m m o n pathway similar to sulfur might exist, w h e r e both selenite and selenide could be the important intermediates for the synthesis of seleno-cysteine. Our results suggest that the selenite metabolite could be p r o d u c e d either through reduction of selenate or oxidative degradation of organo-selenium c o m p o u n d s from diet.
Gel F~ltratJon Figure 1 shows the mol-wt distribution of selenium-bound protein components in serum of the rat after administration of selenite, selenate, and seleno-methionine in 1, 3, 6, and 24 h, respectively. Recovery of selenium in these experiments is 70-140% (data not given). Four protein peaks could be differentiated in the chromatogram (A280, not illustrated) as generally reported by using the same technique. Increased selenium contents were found in all protein classes separated by gel-filtration in 3
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Vol. 33, 1992
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Zi-jian, die, a n d A n
Table 4 Selenite Metabolite in S e r u m of the Rat Following a n Oral A d m i n i s t r a t i o n of Different Selenium C o m p o u n d s Selenite content, ~g/L Time, h
Se (IV) 3.7 20.6 6.2 4.6 3.4
1 3
6 12 24
Se-met
_+ 1.9 _+ 7.5 +_ 1.7 + 1.2 __+ 2.1
Se (VI)
4.0 + 1.2 11.0 _+ 1.1 4.9 + 0.4 1.0 _ 0.2 N.D.*
11.2 28.4 14.5 12.0 10.3
_+ +_ _+ + _+
10.6 9.9 4.7 1.1 2.2
Se-yeast
Control
4.0 -+ 2.0 4.6 --_ 0.8 1.8 _+_ 0.4 0.5 ---0.4 N.D.
-
N.D. -
*N.D. < 0.2 ~g/L.
f, f,
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gELENATE
SELENITE
SELENO-METHIONINE
Fig. I. Chromatographic time-course of selenium in serum of the rat following an oral administration of selenite, selenate, and seleno-dl-methionine. Molecular-weight calibration used Pharmacia Fine Chemical Co. kit, Switzerland; a--blue dextran 2000 (mol wt 2,000,000); b~bovine serum albumin (tool wt 67,000); c--ovalbumin (mol wt 43,000); d--ribonudease (tool wt 13,700); e~sodium fluorescein (mol wt 376). Fractions were divided into four parts as in Fig. 2; F , - - m o l w t 100-2000 kDa; F2--mol w t 25-100 kDa; F3--mol w t 10-25 kDa; F 4 - - m o l w t < 10 kDa.
Biological Trace Element Research
Vol. 33, 1992
IVletabolic Differences of Se Compounds A
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Biological Trace Element Research
VoL 33, 1992
142
Zi-jian, die, and An
and 6 h (Fig. 2), similar to the time-course of selenium content in serum, but the incorporation of selenium into protein components was different among the chemical forms applied. This is clearly demonstrated by Fig. 2, where the total eluent was divided into four parts (F1 to F4), each corresponding to a protein class as indicated in the figure captions. Relative accumulation was expressed for each of these four parts as the ratio of selenium in the Se-supplemented group to that in the control group (12 h). Figure 2 shows that the relative selenium accumulation of different selenium compounds in high-mol-wt fractions (F1, mol-wt 100-2000 kDa) reached 100% in 24 h, indicating that F1 represents only a transient carrier. Seleno-methionine tends to be accumulated in F2 (mol wt 25-100 kDa), whereas selenite and selenate tend to be accumulated in F 3 (tool wt 10-25 kDa) in 24 h, suggesting different incorporations into serum protein. Within 24 h, the accumulation and evolution patterns of the different selenium species in F 2 and F 4 ( < 1 0 kDa) became quite similar, suggesting that F2 is possibly interrelated with F4 for the recycling of low-mol-wt selenium components between erythrocyte and serum (15). The calculated percentage of selenite in F4 is ~29% of the total, indicating that other species, such as selenide, seleno-diglutathione, and so forth, are present.
ACKNOWLEDGMENT The authors gratefully acknowledge Professor H. D. Shu and Dr. L. P. Liang from Beijing Medical University for animal care. Thanks are also owing to Professor G. N. Schrauzer for kindly correcting the manuscript. This work was supported by the Chinese National Foundation.
REFERENCES 1. C. D. Thomson, R. D. H. Stewart, and M. F. Robinson, Br. J. Nutr. 33, 45-54 (1975). 2. C. D. Thomson and R. D. H. Stewart, Br. J. Nutr. 30, 139-147 (1973). 3. M. Richold, M. F. Robinson, and R. D. H. Stewart, Br. J. Nutr. 38, 19-29 (1977). 4. R. F. Burk, Trace Elements in Human Health and Disease, II, Essential and Toxic Elements, A. S. Prasad, ed., Academic, New York, 1976, pp. 105-133. 5. WHO, Environmental Health Criteria 58, Selenium, published under the joint sponsorship of UNEP, ILO and WHO, Geneva, 1987, pp. 41-43. 6. Z. J. Wang and A. Peng, J. Environm. Sci. (China) 1, 116-121 (1989) (Eng.). 7. K. P. McConnell and G. J. Cho, Am. J. Physiol. 208, 1191-1195 (1965). 8. H. W. Symonds, B. F. Sansom, D. L. Mather, and M. J. Vagg, Br. J. Nutr. 45, 117-120 (1981). 9. D. Behne, H. Hilmert, S. Scheid, H. Gessner, and W. Elger, Biochim. Biophysi. Acta 966, 12-21 (1988).
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Vol. 33, 1992
/vletabolic Differences of Se Compounds
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10. I. Rosenfield and O. A. Beath, eds. Selenium Geobatany, Biochemistry, Toxicity and Nutrition, Academic, New York, 1964, pp. 335--365. 11. R. F. Burk and P. E. Gregory, Arch. Biochem. Biophys. 213, 73-80 (1982). 12. M. A. Motsenbocker and A. L. Tappel, Biochim. Biophys. Acta, 704, 253 (1982). 13. M. A. Beilstein and P. H. Whanger, J. Nutr. 113, 2138 (1983). 14. O. A. Levander, Trace Elements in Human Health and Disease, II, Essential and Toxic Elements, A. S. Prasad, ed., Academic, New York, 1976, pp. 135-163. 15. W. B. Davidson and C. H. Mcmurray, J. Inorg. Biochem. 34, 1-9 (1988).
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