Relative Bioavailability Of Cu Research Articles

A field study of vertical mobility and relative bioavailability of Cu and Ni in calcareous soil

ABSTRACT The vertical mobility of heavy metals in the soil profile may affect their bioavailability. In the present study, a field experiment was conducted for 11 years to investigate the vertical mobility of copper (Cu) and nickel (Ni) and the effects on their bioavailability in soil. It was found that the type and concentration of heavy metals are factors that affect mobility. After 11 years, the mobility and movement of Ni were greater than that of Cu from topsoil (0–20 cm) to subsoil (20–60 cm). The vertical mobility of Cu decreased with the increased addition of Cu, while Ni had the highest vertical mobility when its concentration was the highest in soil. The aging processes and vertical movement could lead to decreases in Cu availability (68–78%) and Ni availability (83–89%). For the decrease of Cu availability in soil, the main driving factor was the vertical movement at low added concentrations, but gradually became aging processes with the increase of added Cu concentrations, while Ni had the opposite tendency. Compared with Cu, aging and vertical mobility played a more prominent role in decreasing the bioavailability of Ni, the bioavailability of Cu changed less than that of Ni after 11 years. The decreased rate of Cu in wheat grains was about 0.26–1.05% while that of Ni was about 2.48–9.46% per year from 2009 to 2018. Therefore, the influence of the vertical downward movement of heavy metals should not be ignored in the assessment of the risk and bioavailability of heavy metals in soil.

Relative bioavailability of copper in tribasic copper chloride to copper in copper sulfate for laying hens based on egg yolk and feather copper concentrations

This experiment was conducted to determine the relative bioavailability (RBV) of Cu in tribasic copper chloride (TBCC) to Cu in copper sulfate (monohydrate form; CuSO4·H2 O) for layer diets based on egg yolk and feather Cu concentrations. A total of 252, 72-wk-old Hy-Line Brown laying hens were allotted to 1 of 7 treatments with 6 replicates consisting of 6 hens per replicate in a completely randomized design. Hens were fed corn-soybean meal-based basal diets supplemented with 0 (basal), 100, 200, or 300 mg/kg Cu from CuSO4 or TBCC for 4 wk. Results indicated that egg production, egg weight, and egg mass were not affected by dietary treatments. However, increasing inclusion levels of Cu in diets from CuSO4 decreased (P < 0.05) feed conversion ratio (FCR), whereas increasing inclusion levels of Cu in diets from TBCC did not affect FCR, indicating significant interaction (P < 0.05). Increasing inclusion levels of Cu from TBCC or CuSO4 increased (P < 0.05) Cu concentrations of egg yolk and feathers. Feather Cu concentrations were greater (P < 0.01) for hens fed diets containing CuSO4 than for hens fed diets containing TBCC. The values for the RBV of Cu in TBCC to Cu in CuSO4 based on log10 transformed egg yolk and feather Cu concentrations were 107.4% and 69.5%, respectively. These values for the RBV of Cu in TBCC did not differ from Cu in CuSO4 (100%). The RBV measured in egg yolk did not differ from the RBV measured in feather. In conclusion, the RBV of Cu in TBCC to Cu in CuSO4 can be determined using Cu concentrations of egg yolk and feathers although the values depend largely on target tissues of laying hens. For a practical application, however, the RBV value of Cu in TBCC to Cu in CuSO4 could be 88.5% when the RBV values determined using egg yolk and feather Cu concentrations were averaged.

Determination of relative bioavailability of copper in tribasic copper chloride to copper in copper sulfate for broiler chickens based on liver and feather copper concentrations

This experiment was conducted to determine the relative bioavailability (RBV) of copper (Cu) in tribasic copper chloride (TBCC) to Cu in copper sulfate (monohydrate form; CuSO4·H2O) as a reference Cu source for broiler diets based on liver and feather Cu concentrations. A total of 588 1-day-old broiler chicks were randomly allotted to 1 of 7 dietary treatments with 7 replicate cages of 12 birds each (6 males and 6 females). Birds were fed the corn–soybean meal-based basal diet (8.76mg/kg Cu), or basal diets supplemented with 0, 100, 200, or 300mg/kg Cu from either CuSO4 or TBCC for 21 d. Results indicated that birds fed diets containing 300mg/kg of Cu from CuSO4 had the least (P<0.05) average daily gain (ADG) and average daily feed intake (ADFI) among 7 dietary treatments. The ADG and ADFI were less (P<0.01) for birds fed diets containing CuSO4 than for those fed diets containing TBCC. Birds fed diets containing 300mg/kg of Cu had greater (P<0.01) liver and feather Cu concentrations than those fed diets containing 100 or 200mg/kg of Cu. The RBV of Cu in TBCC to Cu in CuSO4, when it was calculated based on log10 transformed liver Cu concentrations and added Cu intake, was 92.6±7.9%, whereas the RBV of Cu in TBCC to Cu in CuSO4 determined using feather Cu concentrations was 84.3±8.4%. However, these two values were not significantly different. In conclusion, the RBV of Cu in TBCC to Cu in CuSO4 varies with target tissues, but the values are not significantly different. Thus, the RBV of Cu in TBCC to Cu in CuSO4 can also be determined using feather Cu concentrations in broiler chickens. From a practical standpoint, the RBV is likely close to 88.5% when values determined from liver and feather Cu concentrations are averaged.

Plankton copper requirements and uptake in the subarctic Northeast Pacific Ocean

We undertook the first measurements of metabolic Cu requirements (net Cu:C assimilation ratios) and steady-state Cu uptake rates (ρCu ss) of natural plankton assemblages in the northeast subarctic Pacific using the short-lived radioisotope 67Cu. Size-fractionated net Cu:C assimilation ratios varied ∼3 fold (1.35–4.21 μmol Cu mol C −1) among the stations along Line P, from high Fe coastal waters to the Fe-limited open ocean. The variability in Cu:C was comparable to biogenic Fe:C ratios in this region. As previously observed for Fe uptake, the bacterial size class accounted for half of the total particulate ρCu ss. Interestingly, carbon biomass-normalized rates of Fe uptake from the siderophore desferrioxamine B (DFB) (ρFe DFB; a physiological proxy for Fe-limitation) by the >20 μm size class were positively correlated with the intracellular net Cu:C assimilation ratios in this size class, suggesting that intracellular Cu requirements for large phytoplankton respond to increased Fe-limitation. At Fe-limited Ocean Station Papa (OSP), we performed short-term Cu uptake (ρCu L) assays to determine the relative bioavailability of Cu bound to natural and synthetic ligands. Like the volumetric ρCu ss measured along Line P, the bacterial size class was responsible for at least 50% of the total ρCu L. Uptake rates of Cu from the various organic complexes suggest that Cu uptake was controlled by the oxidation state of the metal and by the metal:ligand concentration ratio, rather than the concentration of inorganic species of Cu in solution. Collectively, these data suggest that Cu likely plays an important role in the physiology of natural plankton communities beyond the toxicological effects studied previously.

Bioavailability of copper from copper glycinate in steers fed high dietary sulfur and molybdenum1,2

Sixty Angus (n = 29) and Angus-Sim-mental cross (n = 31) steers, averaging 9 mo of age and 277 kg of initial BW, were used in a 148-d study to determine the bioavailability of copper glycinate (CuGly) relative to feed-grade copper sulfate (CuSO(4)) when supplemented to diets high in S and Mo. Steers were blocked by weight within breed and randomly assigned to 1 of 5 treatments: 1) control (no supplemental Cu), 2) 5 mg of Cu/kg of DM from CuSO(4), 3) 10 mg of Cu/kg of DM from CuSO(4), 4) 5 mg of Cu/kg of DM from CuGly, and 5) 10 mg of Cu/kg of DM from CuGly. Steers were individually fed a corn silage-based diet (analyzed 8.2 mg of Cu/kg of DM), and supplemented with 2 mg of Mo/kg of diet DM and 0.15% S for 120 d (phase 1). Steers were then supplemented with 6 mg of Mo/kg of diet DM and 0.15% S for an additional 28 d (phase 2). Average daily gain and G:F were improved by Cu supplementation regardless of source (P = 0.01). Final ceruloplasmin, plasma Cu, and liver Cu values were greater (P < 0.05) in steers fed supplemental Cu compared with controls. Plasma Cu, liver Cu, and ceruloplasmin values were greater (P < 0.05) in steers supplemented with 10 mg of Cu/kg of DM vs. those supplemented with 5 mg of Cu/kg of DM. Based on multiple linear regression of final plasma Cu, liver Cu, and ceruloplasmin values on dietary Cu intake in phase 1 (2 mg of Mo/kg of DM), bioavailability of Cu from CuGly relative to CuSO(4) (100%) was 140 (P = 0.10), 131 (P = 0.12), and 140% (P = 0.01), respectively. Relative bio-availability of Cu from CuGly was greater than from CuSO(4) (P = 0.01; 144, 150, and 157%, based on plasma Cu, liver Cu, and ceruloplasmin, respectively) after supplementation of 6 mg of Mo/kg of DM for 28 d. Results of this study suggest that Cu from CuGly may be more available than CuSO(4) when supplemented to diets high in S and Mo.

Pinto beans are a source of highly bioavailable copper in rats.

The trace element copper (Cu) is a required nutrient in the diets of humans. It has been found in animal studies to be essential for efficient iron absorption and oxygen utilization and for aiding free-radical degradation. Dry beans (Phaseolis vulgaris) are potentially good sources of Cu; thus, the objective of this study was to determine the bioavailability of Cu from dry beans using the pinto bean as the source. Dry beans were obtained from a local market, cooked according to package directions, and dried. Weanling male Sprague-Dawley rats (8 groups of 8 rats each) were fed a Cu-deficient diet (AIN-93G) for 4 wk followed by 2 wk of Cu repletion with diets containing 0-6.5 mg Cu/kg diet added as CuSO(4) or with 0.6 and 1.5 mg Cu/kg incorporated into rat diets as pinto beans at 10 and 20%. Standard response curves were developed based on repletion-induced recovery of 10 indices of Cu status, including organ Cu concentrations and Cu-dependent enzyme activities, in response to increasing dietary Cu as CuSO(4). Recovery of these variables in rats fed the pinto bean diets was compared with the standard response curve at similar levels of dietary Cu. Based on the recovery of all 10 variables, the relative bioavailability of Cu from dry beans was at least 100% of that with the highly available CuSO(4). For 3 of the variables, liver and heart Cu concentrations and serum superoxide dismutase 3 activity, estimated bioavailability values of Cu from beans were 138, 140, and 134%, respectively, of those from CuSO(4). We conclude that the dry pinto bean is a good source of dietary Cu with respect to both concentration and bioavailability.

Bioavailability of copper from tribasic copper chloride and copper sulfate in growing cattle

Two experiments were conducted to determine the bioavailability of copper (Cu) from tribasic Cu chloride (Cu 2(OH) 3Cl) relative to Cu sulfate in growing steers. Experiment 1 compared tribasic Cu chloride to sulfate in terms of ability to maintain Cu status when supplemented to steers fed diets high in the Cu antagonists, molybdenum (Mo) and sulfur (S). Sixty Angus and Angus × Hereford steers (257 ± 2 kg body weight) were stratified by body weight and randomly assigned to treatments. Treatments consisted of 0, 5 or 10 mg supplemental Cu/kg diet DM from either Cu chloride or Cu sulfate. All diets were supplemented with 5 mg Mo/kg and 1.5 g S/kg. The control corn silage based diet analyzed 4.9 mg Cu/kg and 6.9 mg Mo/kg and was calculated to contain 3.0 g S/kg. Plasma and liver Cu concentrations and plasma ceruloplasmin activity decreased ( P < 0.01) in all treatment groups during the 98-day study. Copper supplemented steers had higher ( P < 0.01) plasma Cu, plasma ceruloplasmin, and liver Cu than controls at the end of the study. Steers supplemented with 10 mg Cu/kg had higher ( P < 0.01) plasma Cu, plasma ceruloplasmin, and liver Cu than those receiving 5 mg Cu/kg diet. Bioavailability of Cu from Cu chloride, relative to Cu sulfate, was estimated from plasma Cu and ceruloplasmin on day 84 and liver Cu on day 98 using multiple linear regression and a slope ratio technique. Compared with Cu sulfate (1.00), relative bioavailability of Cu from tribasic Cu chloride was 1.32 ( P < 0.08), 1.18 ( P < 0.38) and 1.96 ( P < 0.04) based on plasma Cu, plasma ceruloplasmin and liver Cu, respectively. In experiment 2, 43 Angus and Simmental steers (375 ± 7 kg BW) that had previously been depleted of Cu were used in a 21-day repletion study. Steers were randomly assigned within breed to treatment, and individually fed a corn silage based diet low in Mo (1.18 mg/kg). Treatments consisted of 0, 50 or 100 mg supplemental Cu/day from either Cu chloride or Cu sulfate. Plasma Cu, plasma ceruloplasmin and liver Cu increased ( P < 0.01) in Cu supplemented, but not in control, steers. Plasma and liver Cu concentrations increased ( P < 0.01) to a greater extent in steers receiving 100 mg Cu/day compared to those given 50 mg Cu/day. Tribasic Cu chloride and Cu sulfate were similar ( P > 0.10) in their ability to increase Cu status in Cu depleted steers fed a diet low in Mo. Tribasic Cu chloride is more bioavailable than CuSO 4 when added to diets high in the Cu antagonists Mo and S. When evaluated in Cu depleted steers fed diets low in Mo, the two Cu sources had a similar bioavailability.

Chemical characteristics and relative bioavailability of supplemental organic copper sources for poultry.

Five commercially available organic Cu products and reagent-grade CuSO4 x 5H2O (Cu Sulf) were evaluated by polarographic analysis and solubility in 0.1 M K2HPO4-KH2PO4 buffer (pH 5), 0.2 M HCl-KCl buffer (pH 2), or deionized water. Fractions from these solubility tests were evaluated by gel filtration chromatography for structural integrity. The organic sources were Cu lysine complex (Cu Lys), Cu amino acid chelate (Cu AA), Cu proteinate A (Cu ProA), Cu proteinate B (Cu ProB), and Cu proteinate C (Cu ProC). Separation of peaks in the chromatograms for the soluble Cu fraction from deionized water indicated that 77, 31, 69, 94, and 16% of the Cu remained chelated for the above sources, respectively. Two experiments were conducted to estimate the relative bioavailability of Cu from the organic Cu supplements for chicks when added at high dietary concentrations to practical corn-soybean meal diets. Liver Cu concentration increased (P < 0.0001) as dietary Cu increased in both experiments. When Cu Sulf was assigned a value of 100% as the standard, linear regression slope ratios of log10 liver Cu concentration regressed on added dietary Cu concentration gave estimated relative bioavailability values of 124 +/- 5.1, 122 +/- 5.3, and 111 +/- 6.0 for Cu Lys, Cu AA, and Cu ProC, respectively, in Exp. 1. The bioavailability estimates for Cu Lys and Cu AA were greater (P < 0.05) than that for Cu Sulf. Values in Exp. 2 were 111 +/- 7.6, 109 +/- 8.4, and 105 +/- 7.5 for Cu Lys, Cu ProA, and Cu ProB, respectively, and all sources were similar in value for chicks. Solubility of Cu in pH 2 buffer provided the best prediction of bioavailability (r2 = 0.924). Other indicators of chelation integrity and solubility had little value as predictors of bioavailability (r2 < or = 0.445).

Relative bioavailability of copper in a copper-lysine complex or copper sulfate for ruminants as affected by feeding regimen

An experiment was conducted with 40 crossbred wether lambs to compare the relative bioavailability of Cu from a Cu-lysine complex with that from Cu sulfate under two feeding regimens. Liver biopsy samples were collected by a laparotomy technique and lambs were given 12 days to recover and adapt to individual pens. Lambs were fed either a control basal diet (9.8 mg kg −1 Cu dry matter (DM) basis) or basal supplemented with 180 mg kg −1 Cu as either reagent grade CuSO 4 · 5H 2O or a feed-grade Cu-lysine complex. There were 12 lambs fed control diet and 14 lambs fed each Cu-supplemented diet. Half of the lambs given each diet were fed 1.0 kg of their respective diet once daily at 08:00 h. The remaining half were fed 1.0 kg in equal portions (166 g) six times during a 24 h period at 04:00 h, 08:00 h, 12:00 h, 16:00 h, 20:00 h and 24:00 h. Lambs were fed experimental diets for 10 days, then killed and livers removed for Cu analysis. Lambs fed once daily accumulated more ( P < 0.05) Cu in liver than those fed every 4 h. There was no Cu source × feeding regimen interaction ( P > 0.10). Based on multiple linear regression slope ratio of log 10 transformed final liver Cu concentration on total intake of Cu (mg day −1) with the initial liver biopsy Cu concentration as a covariate, bioavailability of Cu from Cu-lysine relative to Cu sulfate was 93.4% ( P > 0.05) for combined feeding regimens.

Bioavailability of feed-grade copper sources (oxide, sulfate, or lysine) in growing cattle

Plasma Cu concentrations and ceruloplasmin (a Cu metalloenzyme) activity were used to assess relative Cu bioavailability from different Cu sources for growing cattle. In Exp. 1, 18 calves (average BW 207 +/- 7.7 kg) that had been fed Cu-deficient diets since birth were randomly assigned to treatment. Treatments consisted of control (n = 3) or 30 mg/d of supplemental Cu (n = 5/treatment) from Cu oxide (CuO), Cu sulfate (CuSO4), or Cu lysine. Blood samples were obtained for plasma Cu and ceruloplasmin activity on d 0, 7, 14, and 21. Plasma Cu and ceruloplasmin activity were greater (P < .05) on d 7, 14, and 21 for calves supplemented with CuSO4 than for controls. Copper status of calves fed Cu lysine did not differ from that of calves fed CuSO4. Compared with d-0 values, plasma Cu concentrations by d 21 had increased by 95 and 98% in calves supplemented with CuSO4 and Cu lysine, respectively. Copper oxide supplementation did not increase Cu status above that observed in control calves. In Exp. 2, 36 calves (average BW 211 +/- 4.4 kg) were used to compare the relative bioavailability of Cu from CuO and CuSO4 when supplemented to corn silage-based diets high in the Cu antagonists iron (Fe) or molybdenum (Mo). Treatments consisted of no supplemental Cu or 8 mg of supplemental Cu/kg diet from either CuSO4 or CuO. Within each Cu treatment, diets were supplemented with 600 mg of Fe or 5 mg of Mo/kg diet. In calves fed 5 mg of Mo/kg, plasma Cu was lower (P < .05) in those fed no supplemental Cu or CuO compared with calves fed CuSO4 by d 91 and at subsequent sampling dates throughout the 154-d study. Plasma ceruloplasmin activity was decreased (P < .01) by Mo and was increased (P < .05) by CuSO4 compared with CuO. Based on plasma Cu and ceruloplasmin activity, CuSO4 and Cu lysine were similar in bioavailability, but CuO was essentially unavailable.

Relative bioavailability of copper in a copperlysine complex for chicks and lambs

Two experiments were conducted to study the relative bioavailability of Cu in a Cu-lysine complex with that of reagent grade CuSO 4·5H 2O for chicks and lambs. Chicks were fed a basal diet (11 mg kg −1 Cu, dry matter (DM) basis) or the basal diet supplemented with 150, 300 or 450 mg kg −1 Cu from either of the two Cu sources for 18 days. Birds were killed and liver Cu concentrations determined. Liver biopsy samples were collected from wether lambs who were then fed the basal diet (9.5 mg kg −1 Cu DM) or the basal diet supplemented with 60, 120 or 180 mg kg −1 Cu from either source for 10 days. Copper in the initial biopsy was used as a covariate in the multiple linear regression of log 10 final liver Cu concentration on added dietary Cu concentration. Slope ratio comparisons gave relative Cu bioavailability values of 99 and 68 for Cu-lysine in chicks and lambs, respectively, compared with 100 for CuSO 4·5H 2O for both species.

Nutritional Evaluation of a Copper-Methionine Complex for Chicks

Three chick experiments were conducted to evaluate a new Cu complex product, Cu-Met. Relative bioavailability of Cu was assessed with two bioassay systems, one employing bile Cu concentration as the criterion at Cu-deficient levels and another employing liver Cu concentration at Cu-excess levels. Relative bioavailability of Met was assessed separately. Common-intercept multiple linear regression (slope-ratio) of bile Cu concentration or liver Cu concentration regressed on supplemental Cu intake was used to assess Cu bioavailability relative to the standard, analytical-grade CuSO4·5H2 O, which was set at 100%. In addition, Cu-Lys and CuCl (compounds previously evaluated by bile Cu concentration) were evaluated by the liver Cu accumulation method. Relative bioavailability of Cu in Cu-Met was estimated as 96% by the bile Cu assay and 88% by the liver Cu assay, which were not different (P > .05) from 100%. The liver Cu assay estimated that relative Cu bioavailabilities in Cu-Lys and CuCl of 114 and 145% (P < .05), respectively. These values were very close to those previously estimated by the bile Cu assay. Relative Met bioavailability of Cu-Met was 100% of the standard, i.e., feed-grade DL-Met.

Estimates of Copper Bioavailability from Liver of Different Animal Species and from Feed Ingredients Derived from Plants and Animals

Bile Cu accumulation in Cu-depleted chicks fed Cu concentrations between .56 and 1.56 mg/kg was used to estimate Cu bioavailability in several feed ingredients from both plant and animal sources, including liver from different species. Liver from slaughtered animals is rich in minerals and vitamins and is a significant source of Cu in canned pet foods. Liver from different species, however, was found to vary widely in Cu bioavailability (relative to CuSO4·5H2 O, which was set at 100%). The bioavailability of Cu in freeze-dried (FD) chicken liver and poultry by-product meal was 116 and 97%, respectively, but that in FD pork liver was not different from zero. Relative bioavailability of Cu in FD beef, sheep, and turkey liver was 82, 113, and 83%, respectively. Copper in FD liver from the rat, a species that does not possess a gall bladder, was 21% bioavailable. Copper in the feed ingredients from plants: corn gluten meal, dehulled soybean meal, cottonseed meal, peanut hulls, and soy mill run was 48, 38, 41, 44, and 47% bioavailable, respectively. In addition, when the fibrous ingredients peanut hulls or soy mill run were added to the basal diet containing .5 mg Cu/kg from CuSO4·5H2 O, Cu bioavailability in CuSO4·5H2 O was reduced. The results of this study demonstrate a wide variation in Cu bioavailability among feed ingredients originating from plants and animals.

Bioavailability of Copper in Analytical-Grade and Feed-Grade Inorganic Copper Sources when Fed to Provide Copper at Levels Below the Chick’s Requirement

Young chicks were fed a casein-soy protein concentrate basal diet (.56 mg Cu/kg) containing graded levels of added Cu (0, .5, 1.0 mg/kg) from analytical-grade (AG) CuSO4·5H2 O, Cu2 O, CuO, CuCO3·Cu(OH)2, CuCl, and also from feed-grade (FG) CuSO4·5H2 O and FG-CuO. Weight gain, hematocrit, hemoglobin, plasma Cu, liver Cu, gall bladder (bile) Cu, and tendon lysine were assessed to determine whether dose-dependent Cu responses would occur. Gall bladder Cu concentration as well as total Cu content of the gall bladder were found to be linear functions of supplemental Cu intake. Common-intercept multiple linear regression (slope-ratio) of bile Cu concentration regressed on supplemental Cu intake indicated Cu bioavailability values (relative to the standard, AG-CuSO4·5H2 O, set at 100%) of 97.9% for AG-Cu2 O, 93.5% for FG-CuSO4·5H2 O, and 112.9% for AG-CuCO3·Cu(OH)2. These values were not different (P > .05) from 100%. Relative bioavailability of Cu in AG-CuCl was 142.5%, which was different (P < .05) from the standard. Both AG-CuO and FG-CuO gave Cu bioavailability estimates not different (P > .05) from zero. The results indicate that when Cu levels are fed below the chick’s requirement, bile Cu concentration is a sensitive indicator of net gut absorption of Cu.

Nutritional Evaluation of Copper-Lysine and Zinc-Lysine Complexes for Chicks

Three chick experiments were conducted to evaluate two mineral-Lys complexed products, Cu-Lys and Zn-Lys. Relative bioavailability of Lys, Cu, and Zn were assessed separately using diets specifically deficient in each of these nutrients and by employing criteria that respond in a straight-line fashion to supplementation with the nutrient in question. Common-intercept multiple linear regression (slope-ratio) of weight gain, bile Cu concentration, and total tibia Zn regressed on supplemental intake of Lys, Cu, and Zn, respectively, were used to assess bioavailability values relative to the standards (i.e., L-Lys·HCl, CuSO4·5H2 O, and ZnSO4·H2 O). Lysine bioavailability of Cu-Lys and Zn-Lys were estimated as 89 and 109%, respectively, which were not different (P > .05) from 100%. Relative bioavailability of Cu in Cu-Lys and Zn in Zn-Lys were estimated as 120 and 106%, respectively. These values were not different (P > .05) from the standards, i.e., CuSO4·5H2 O for Cu and ZnSO4·H2 O for Zn.