BIOLOGICAL T R A C E ELEMENT RESEARCH 2, 31 39 (1980)
The Effect of Ethanol on Zinc and Copper Metabolism in Rats I V O R E. D R E O S T I , * R E G I N A L D A . BUCKLEY, AND IAN R . R E C O R D
CSIRO Division of Human Nutrition, Adelaide, South Australia 5000 Received September 12, 1979
Abstract Studies performed on adult female rats over a period of 10 weeks indicated that the consumption of alcohol (20% v/v) did not appear to disturb the zinc or copper balance, nor did it adversely affect tissue zinc or copper levels, even in zinc-restricted animals. On the contrary, higher plasma zinc levels were consistently observed in animals receiving alcohol together with the experimental diets. Index Entries: Ethanol, effects on Zn and Cu metabolism; zinc metabolism, effects of ethanol on; copper metabolism, effects of ethanol on; metabolism, of zinc and copper, effects of ethanol on.
Introduction Hyperexposure to alcohol in humans and animals severely disturbs many biochemical systems (1). Chronic alcoholism in humans is often, but not invariably, accompanied by hyperzincuria and low plasma zinc levels, which seem to be associated with cirrhosis of the liver and kidney damage, although the precise mechanism of zinc excretion under these conditions is not understood (2). Very little is known concerning the initial effect of ethanol consumption on zinc metabolism in unaddicted man or animals, although it has been reported that in rats, liver zinc levels fell by about 40% two weeks after receiving 20% ethanol in their drinking water (3). In addition plasma zinc declined consistently by approximately 3% per week from the start of the experiment. 9 1980 by The Humana Press Inc. All rights of any nature whatsoever reserved. 0163-4984/80/0300-0031502.00
31
32
DREOSTI, BUCKLEY, AND RECORD
Because of the importance of zinc for growth and fetal development (4) and because inadequate zinc nutriture is suggested to occur quite widely in humans (5), the possibility that alcohol might modify zinc metabolism is highly important, and it was therefore studied further. Investigations were made with rats to establish whether the consumption of ethanol altered in any way the balance of zinc or its distribution within the animal body. Attention was especially paid to the effect of alcohol on zinc levels in rats receiving a zinc-deficient diet, in order to ascertain whether alcohol might accelerate the rate of zinc depletion in these animals. Copper levels and the balance of copper were determined concurrently as the reciprocal relationship between copper and zinc in many tissues is well documented (2).
Methods Adult female rats of the Hooded Wistar strain weighing approximately 200 g were housed either individually in metabolism cages or in groups of four in plastic and stainless steel cages; animals received either the stock colony diet (W. Charlick Ltd., Adelaide, Australia), which contained on assay 109 _+ 2.1 ppm of zinc and 15.7 _+ 0.8 ppm of copper, or a zincdeficient or zinc-s upplemented diet. The zinc-deficient diet (6) was prepared from EDTA-extracted soyabean flour (7) and contained on assay less than 0.5 ppm zinc and 7.8 _+ 1.4 ppm copper.The zinc-supplemented rats received the same diet containing an additional 100 ppm of zinc as zinc sulfate. In some cases water was replaced as the drinking fluid by an aqueous solution of 20% (v/v) ethanol. Zinc and copper balance studies were performed on a 7-day basis and included measurements of total zinc and copper intake, and total urinary and fecal zinc and copper excretion. Tissues for zinc and copper analyses were removed from other groups of animals at appropriate time intervals. Zinc and copper were estimated using a Varian Techtron AA6 atomic absorption spectrophotometer (Varian Techtron Pty. Ltd., Melbourne, Australia).
Results and Discussion When animals were changed from receiving the zinc-supplemented diet and water for two weeks (control values) to the zinc-deficient diet, food intake declined progressively to a level approximately 68% of the control consumption (Table 1). Fecal output fell in parallel with the reduced food intake and urine volume was unchanged. With animals that received 20% ethanol concurrently with the introduction of the zinc deficient diet (Table 1), food intake fell immediately by about
33
ETHANOL IN ZINC AND COPPER METABOLISM
TABLE 1 Food Intake and Excretory Output in Rats Receiving a Zinc-Deficient Diet and Either (1) Water or (2) 20% Ethanol for Drinking a
Treatment
Food intake, Rat weight, g g/rat/wk
Excretory output, g/rat/wk Feces
Urine
(1) Water + Z n cont~'ol, 2 wk - Z n , 1 wk - Z n , 2 wk - Z n , 3 + 4 wk - Z n , 5 + 6 wk - Z n , 7 + 8 wk - Z n , 9 + 10 wk
239 + 6 225+6 190 + 9 194 • 5 200 • 5 192 + 7 188 + 7
96 _+ 3 98_+7 96 + 2 80 _+ 2 84 • 3 67 + 3 66 + 2
6.9 + 0.9 8.0+0.8 5.2 + l.l 4.3 _+_0.2 4.8_+ 0.4 5.1 + 0.4 3.1 + 0.3
67 _+ 9 63_+9 85 + 4 88 • 12 91 + 8 77 + 14 78 + 8
(2) 20% Ethanol +Zn + H20 control, 2 wks - Z n , 1 wk - Z n , 2 wk - Z n , 3 + 4 wk -Zn, 5+6wk - Z n , 7 + 8 wk - Z n , 9 + 10 wk
198 + 8 184_+ 11 179 • 12 166 • 12 146• 136 • 16 129 • 15
84 + 3 45+2 51 + 1 39 + 3 35+4 37 • 2 29 + 1
5.3 + 1.1 2.1 +0.2 2.7 + 0.4 2.5 + 0.2 1.6+0.1 1.5 + 0.5 1.9 + 0.2
73 + 15 13.5+2.4 11.5 + 2.5 9.9 + 1.3 7.5+1.8 6.2 • 1.3 7.9 + 0.6
aValues represent the means + S E M of 4 rats in each group.
40%, and thereafter declined progressively to a level around 35% of the control intake. Fecal output responded in a similar manner and urinary excretion was greatly reduced in volume (90%), which largely reflected the severely diminished ( > 60%) fluid intake of the animals consuming alcohol. In animals receiving the zinc-deficient diet and water fecal zinc excretion fell precipitately, so that after only two weeks of zinc restriction total fecal zinc loss amounted to less than 100/ag zinc/rat/week (Table 2). Urinary zinc levels delcined to a lesser extent over the same period and finally represented a zinc output a little less than one half of the control value. Animals immediately entered a negative zinc balance on receiving the zinc-deficient diet which persisted until the end of the experiment. When 20% ethanol was introduced into the dietary regimen together with the zinc deficiency fecal zinc output decreased to a level of about 26 # g z i n c / r a t / w e e k after 10 weeks of treatment (Table 2). Urinary zinc excretion too fell over this period, although again it was prop ortionately less affected than fecal zinc. These animals also abruptly entered into a negative zinc balance, which continued throughout the 10 weeks of study.
34
DREOSTI, BUCKLEY, AND RECORD
TABLE 2 Zinc Balance in Rats Receiving Receiving a Zinc-Deficient Diet and Either (1) Water or (2) 20% Ethanol for Drinkinga
Treatment
Zinc intake, #g/rat/wk
(1) Water +Zn control, 2 wk - Z n , 1 wk - Z n , 2 wk -Zn, 3+4wk
9648 _+ 302
The Effect of Ethanol on Zinc and Copper Metabolism in Rats I V O R E. D R E O S T I , * R E G I N A L D A . BUCKLEY, AND IAN R . R E C O R D
CSIRO Division of Human Nutrition, Adelaide, South Australia 5000 Received September 12, 1979
Abstract Studies performed on adult female rats over a period of 10 weeks indicated that the consumption of alcohol (20% v/v) did not appear to disturb the zinc or copper balance, nor did it adversely affect tissue zinc or copper levels, even in zinc-restricted animals. On the contrary, higher plasma zinc levels were consistently observed in animals receiving alcohol together with the experimental diets. Index Entries: Ethanol, effects on Zn and Cu metabolism; zinc metabolism, effects of ethanol on; copper metabolism, effects of ethanol on; metabolism, of zinc and copper, effects of ethanol on.
Introduction Hyperexposure to alcohol in humans and animals severely disturbs many biochemical systems (1). Chronic alcoholism in humans is often, but not invariably, accompanied by hyperzincuria and low plasma zinc levels, which seem to be associated with cirrhosis of the liver and kidney damage, although the precise mechanism of zinc excretion under these conditions is not understood (2). Very little is known concerning the initial effect of ethanol consumption on zinc metabolism in unaddicted man or animals, although it has been reported that in rats, liver zinc levels fell by about 40% two weeks after receiving 20% ethanol in their drinking water (3). In addition plasma zinc declined consistently by approximately 3% per week from the start of the experiment. 9 1980 by The Humana Press Inc. All rights of any nature whatsoever reserved. 0163-4984/80/0300-0031502.00
31
32
DREOSTI, BUCKLEY, AND RECORD
Because of the importance of zinc for growth and fetal development (4) and because inadequate zinc nutriture is suggested to occur quite widely in humans (5), the possibility that alcohol might modify zinc metabolism is highly important, and it was therefore studied further. Investigations were made with rats to establish whether the consumption of ethanol altered in any way the balance of zinc or its distribution within the animal body. Attention was especially paid to the effect of alcohol on zinc levels in rats receiving a zinc-deficient diet, in order to ascertain whether alcohol might accelerate the rate of zinc depletion in these animals. Copper levels and the balance of copper were determined concurrently as the reciprocal relationship between copper and zinc in many tissues is well documented (2).
Methods Adult female rats of the Hooded Wistar strain weighing approximately 200 g were housed either individually in metabolism cages or in groups of four in plastic and stainless steel cages; animals received either the stock colony diet (W. Charlick Ltd., Adelaide, Australia), which contained on assay 109 _+ 2.1 ppm of zinc and 15.7 _+ 0.8 ppm of copper, or a zincdeficient or zinc-s upplemented diet. The zinc-deficient diet (6) was prepared from EDTA-extracted soyabean flour (7) and contained on assay less than 0.5 ppm zinc and 7.8 _+ 1.4 ppm copper.The zinc-supplemented rats received the same diet containing an additional 100 ppm of zinc as zinc sulfate. In some cases water was replaced as the drinking fluid by an aqueous solution of 20% (v/v) ethanol. Zinc and copper balance studies were performed on a 7-day basis and included measurements of total zinc and copper intake, and total urinary and fecal zinc and copper excretion. Tissues for zinc and copper analyses were removed from other groups of animals at appropriate time intervals. Zinc and copper were estimated using a Varian Techtron AA6 atomic absorption spectrophotometer (Varian Techtron Pty. Ltd., Melbourne, Australia).
Results and Discussion When animals were changed from receiving the zinc-supplemented diet and water for two weeks (control values) to the zinc-deficient diet, food intake declined progressively to a level approximately 68% of the control consumption (Table 1). Fecal output fell in parallel with the reduced food intake and urine volume was unchanged. With animals that received 20% ethanol concurrently with the introduction of the zinc deficient diet (Table 1), food intake fell immediately by about
33
ETHANOL IN ZINC AND COPPER METABOLISM
TABLE 1 Food Intake and Excretory Output in Rats Receiving a Zinc-Deficient Diet and Either (1) Water or (2) 20% Ethanol for Drinking a
Treatment
Food intake, Rat weight, g g/rat/wk
Excretory output, g/rat/wk Feces
Urine
(1) Water + Z n cont~'ol, 2 wk - Z n , 1 wk - Z n , 2 wk - Z n , 3 + 4 wk - Z n , 5 + 6 wk - Z n , 7 + 8 wk - Z n , 9 + 10 wk
239 + 6 225+6 190 + 9 194 • 5 200 • 5 192 + 7 188 + 7
96 _+ 3 98_+7 96 + 2 80 _+ 2 84 • 3 67 + 3 66 + 2
6.9 + 0.9 8.0+0.8 5.2 + l.l 4.3 _+_0.2 4.8_+ 0.4 5.1 + 0.4 3.1 + 0.3
67 _+ 9 63_+9 85 + 4 88 • 12 91 + 8 77 + 14 78 + 8
(2) 20% Ethanol +Zn + H20 control, 2 wks - Z n , 1 wk - Z n , 2 wk - Z n , 3 + 4 wk -Zn, 5+6wk - Z n , 7 + 8 wk - Z n , 9 + 10 wk
198 + 8 184_+ 11 179 • 12 166 • 12 146• 136 • 16 129 • 15
84 + 3 45+2 51 + 1 39 + 3 35+4 37 • 2 29 + 1
5.3 + 1.1 2.1 +0.2 2.7 + 0.4 2.5 + 0.2 1.6+0.1 1.5 + 0.5 1.9 + 0.2
73 + 15 13.5+2.4 11.5 + 2.5 9.9 + 1.3 7.5+1.8 6.2 • 1.3 7.9 + 0.6
aValues represent the means + S E M of 4 rats in each group.
40%, and thereafter declined progressively to a level around 35% of the control intake. Fecal output responded in a similar manner and urinary excretion was greatly reduced in volume (90%), which largely reflected the severely diminished ( > 60%) fluid intake of the animals consuming alcohol. In animals receiving the zinc-deficient diet and water fecal zinc excretion fell precipitately, so that after only two weeks of zinc restriction total fecal zinc loss amounted to less than 100/ag zinc/rat/week (Table 2). Urinary zinc levels delcined to a lesser extent over the same period and finally represented a zinc output a little less than one half of the control value. Animals immediately entered a negative zinc balance on receiving the zinc-deficient diet which persisted until the end of the experiment. When 20% ethanol was introduced into the dietary regimen together with the zinc deficiency fecal zinc output decreased to a level of about 26 # g z i n c / r a t / w e e k after 10 weeks of treatment (Table 2). Urinary zinc excretion too fell over this period, although again it was prop ortionately less affected than fecal zinc. These animals also abruptly entered into a negative zinc balance, which continued throughout the 10 weeks of study.
34
DREOSTI, BUCKLEY, AND RECORD
TABLE 2 Zinc Balance in Rats Receiving Receiving a Zinc-Deficient Diet and Either (1) Water or (2) 20% Ethanol for Drinkinga
Treatment
Zinc intake, #g/rat/wk
(1) Water +Zn control, 2 wk - Z n , 1 wk - Z n , 2 wk -Zn, 3+4wk
9648 _+ 302
