The aim of this study was to investigate the effects of high dietary fluoride (F) on serum biochemistry and oxidative damage in broiler chickens. 280 one-day-old healthy avian broiler chickens were randomly allotted into four equal groups and fed with a corn-soybean basal diet containing 22.6 mg·F/kg (control group) or same basal diets supplemented with 400, 800, and 1200 mg·F/kg (high F groups I, II, and III) in the form of sodium fluoride for 42 days. At 42 days of age, the serum F content was markedly higher in the three high F groups than that in the control group. From 28 to 42 days of age, the contents of serum total protein (TP) and albumin (ALB) were significantly lower in the three high F groups. From 14 to 42 days of age, the activities of alkaline phosphatase (ALP), alanine aminotransferase (ALT) and the creatinine (Crea) contents in the serum showed a marked increase in the three high F groups; aspartate aminotransferase (AST) activity and uric acid (Ua) content were significantly increased, and a significant increase in the content of malondialdehyde (MDA) along with marked decreases in the activities of total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px), catalase (CAT), the glutathione (GSH) content and the ability to inhibit hydroxyl radical were observed in the high F groups II and III. In conclusion, F has accumulated in the blood circulatory system and dietary F in the range of 800 - 1200 mg/kg could significantly induce abnormalities of bone, liver and kidney, inhibit the synthesis of protein, enhance lipid peroxidation and disturb the antioxidative system of broiler chickens.
Fluoride (F) is known to cause fluorosis in large doses and can even lead to death in excessive amounts [
In our recent studies, we found that high dietary F induced oxidative stress in the cecal tonsil and the intestinal mucosa of broiler chickens [
Thus, the objective of this study was to investigate the effects of high dietary F on serum biochemical parameters and oxidative stress indicators by detecting the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), total superoxide dismutase (T-SOD), CAT, GSH-Px, the ability to inhibit hydroxyl radical and the contents of TP, ALB, creatinine (Crea), uric acid (Ua), GSH and MDA.
280 one-day-old healthy avian broiler chickens (purchased from Wenjiang commercial poultry hatchery in Sichuan Province) were randomly divided into four equal groups with 70 broiler chickens in each and fed a corn- soybean diet as follows: a control group with 22.6 mg·F/kg, and three high F groups I, II, and III with 400, 800, and 1200 mg·F/kg diet, respectively. They were housed in corresponding cages with electrically heated units and had unrestricted access to water and the above-mentioned diets for 42 days.
All the dietary nutrition requirements were adequate according to the US National Research Council (NRC) (1994) [
Animal care and protocol of the experiment were approved by Sichuan Agricultural University Animal Care and Use Committee.
At 14, 28, and 42 days during the experiment, five broiler chickens in each group were phlebotomized from jugular vein to collect serum after a 12-h overnight fast. Non-anticoagulative blood samples were clotted for 15 min at room temperature and then centrifuged at 3000 rpm for 15 min. The serum samples were collected in eppendorf tubes and stored at −20˚C until analysis.
Before determination, the serum samples were thawed. The content of serum F ions was determined with fluoride ion-selective electrode INESA PF-1-01 (China).
The activities of T-SOD, CAT and GSH-Px and the contents of GSH, MDA, and the ability to inhibit hydroxy radicals in the serum were assayed by biochemical reagent kits (T-SOD, A001-1; CAT, A007-2; GSH-Px, A005; GSH, A006-1; MDA, A003-1; ability to inhibit hydroxyl radicals, A018). Meanwhile, the activities of AST, ALT and ALP, the contents of TP, ALB, Crea, Ua were measured using the corresponding reagent kits (AST, C0010-2; ALT, C009-2; ALP, A059-2; TP, A045-2; ALB, A028-1; Crea, C011-1; Ua, C012, respectively). All the aforementioned commercial reagent kits were purchased from Nanjing Jiancheng Bioengineering Institute (China). All the experimental operating procedures followed the manufacturer’s instructions. The absorbances of T-SOD, CAT, GSH-Px, GSH, MDA, ability to inhibit hydroxy radicals were measured at 550, 240, 412, 420, 532, 550 nm and the absorbances of AST, ALT, ALP, TP, ALB, Crea, Ua were detected at 510, 510, 520, 595, 628, 510 and 690 nm, respectively, with a microplate reader Bio-Rad 680 (USA).
All the grouped data were statistically evaluated with SPSS software 17.0 for Windows and expressed as means ± standard deviation (SD). Comparison of means between four groups were conducted using one-way analysis of variance (ANOVA) followed by least significant difference (LSD) post hoc test and P-values of less than 0.05 (p < 0.05) were considered to indicate statistical significance.
As shown in
From 14 to 42 days of age, the activities of T-SOD, CAT, GSH-Px, the content of GSH and the ability to inhibit hydroxyl radicals were significantly lower (p < 0.01) in high F groups II and III than those in the control group, whereas the MDA content was significantly increased (p < 0.01) in the abovementioned high F groups in comparison with the control group (
As shown in
In blood, about 75% of F remains free in plasma, therefore, determination of the content of serum F reflects the F content in blood to some extent. In our present study, the serum F content was markedly higher in the three high F groups than that in the control group at 42 days of age (p < 0.01). The increased content of serum F was reported by many anthors [
The generation and concentration of reactive oxygen species (ROS) in the body is controlled by the antioxidative system, which enables transformation of ROS into inactive and harmless compounds or molecules [
. Changes of the serum F content in broilers
Days | Groups | F (μg/mL) |
---|---|---|
42 days | Control group | 0.06 ± 0.01 |
High F group I | 0.22 ± 0.01** | |
High F group II | 0.75 ± 0.05** | |
High F group III | 1.79 ± 0.09** |
Data are presented with the means ± standard deviation (n = 5). **p < 0.01, compared with the control group.
. Changes in the serum oxidative stress parameters of broilers
Parameters | Groups | Days | ||
---|---|---|---|---|
14 days | 28 days | 42 days | ||
T-SOD (U/ml) | Control group | 127.74 ± 4.15 | 150.91 ± 2.10 | 162.34 ± 3.14 |
High F group I | 127.48 ± 2.52 | 150.65 ± 2.69 | 161.92 ± 2.37 | |
High F group II | 102.25 ± 1.39** | 122.65 ± 1.93** | 145.77 ± 4.72** | |
High F group III | 101.56 ± 1.12** | 119.20 ± 2.20** | 140.91 ± 1.79** | |
GSH-Px (U/ml) | Control group | 544.70 ± 6.94 | 497.70 ± 5.33 | 442.57 ± 14.17 |
High F group I | 527.63 ± 11.06* | 483.51 ± 14.39 | 421.37 ± 11.12* | |
High F group II | 485.10 ± 4.95** | 382.90 ± 16.02** | 326.24 ± 10.19** | |
High F group III | 446.92 ± 13.87** | 348.97 ± 14.16** | 302.12 ± 6.55** | |
CAT (U/ml) | Control group | 8.24 ± 0.58 | 12.99 ± 0.75 | 13.76 ± 0.55 |
High F group I | 7.80 ± 0.36 | 12.39 ± 0.76 | 13.12 ± 0.77 | |
High F group II | 6.64 ± 0.52** | 9.34 ± 0.33** | 11.65 ± 0.51** | |
High F group III | 6.45 ± 0.50** | 8.17 ± 0.38** | 11.19 ± 0.57** | |
GSH (mgGSH/L) | Control group | 13.88 ± 0.71 | 16.96 ± 1.06 | 17.41 ± 0.65 |
High F group I | 13.61 ± 0.86 | 16.05 ± 0.61 | 16.60 ± 1.04 | |
High F group II | 11.57 ± 0.83** | 12.92 ± 0.93** | 13.78 ± 0.99** | |
High F group III | 11.02 ± 0.83** | 10.61 ± 0.73** | 11.83 ± 0.45** | |
MDA (nmol/ml) | Control group | 3.44 ± 0.18 | 3.84 ± 0.76 | 5.96 ± 0.09 |
High F group I | 3.64 ± 0.05 | 4.66 ± 0.03 | 6.45 ± 0.33* | |
High F group II | 5.03 ± 0.96** | 5.58 ± 0.85** | 6.73 ± 0.17** | |
High F group III | 5.27 ± 0.33** | 5.89 ± 0.46** | 8.90 ± 0.40** | |
The ability to inhibit hydroxyl radicals (U/ml) | Control group | 1357.86 ± 2.01 | 1361.47 ± 1.59 | 1362.05 ± 1.55 |
High F group I | 1356.67 ± 3.12 | 1357.80 ± 2.64 | 1360.33 ± 1.38 | |
High F group II | 1348.80 ± 0.18** | 1345.37 ± 3.59** | 1355.48 ± 0.98** | |
High F group III | 1345.24 ± 1.56** | 1344.64 ± 1.30** | 1350.09 ± 3.30** |
Data are presented with the means ± standard deviation (n = 5). *p < 0.05, compared with the control group. **p < 0.01, compared with the control group.
GSH-Px and CAT, as preventive antioxidants, and T-SOD, a chain-breaking antioxidant, play an important role in protecting against the deleterious effects of lipid peroxidation [
. Changes in the serum biochemical indexes of broilers
Parameters | Groups | Days | ||
---|---|---|---|---|
14 days | 28 days | 42 days | ||
TP (g/L) | Control group | 27.47 ± 0.92 | 28.34 ± 1.13 | 30.85 ± 0.43 |
High F group I | 27.12 ± 1.00 | 26.77 ± 0.49* | 25.73 ± 1.02** | |
High F group II | 24.33 ± 1.07** | 23.57 ± 0.65** | 25.94 ± 0.98** | |
High F group III | 18.09 ± 0.60** | 18.72 ± 1.02** | 22.49 ± 0.56** | |
ALB (g/L) | Control group | 16.10 ± 0.77 | 19.90 ± 0.65 | 31.51 ± 1.13 |
High F group I | 17.60 ± 0.59** | 17.30 ± 0.40** | 28.93 ± 0.92** | |
High F group II | 19.80 ± 0.56** | 17.10 ± 0.37** | 27.27 ± 0.40** | |
High F group III | 18.70 ± 0.65** | 15.19 ± 0.37** | 23.70 ± 0.92** | |
ALP (U/100mL) | Control group | 597.71 ± 8.82 | 781.70 ± 16.58 | 1260.39 ± 19.40 |
High F group I | 998.03 ± 12.11** | 1884.29 ± 21.05** | 2228.92 ± 21.15** | |
High F group II | 1133.82 ± 23.53** | 2237.26 ± 14.23** | 2407.08 ± 17.71** | |
High F group III | 1483.33 ± 15.56** | 2566.02 ± 26.35** | 2491.80 ± 22.19** | |
Crea (μmol/L) | Control group | 204.28 ± 5.35 | 239.64 ± 2.48 | 269.11 ± 4.13 |
High F group I | 243.56 ± 4.36** | 246.19 ± 2.96** | 275.03 ± 5.27* | |
High F group II | 261.26 ± 4.59** | 251.42 ± 2.71** | 294.64 ± 1.39** | |
High F group III | 267.15 ± 3.34** | 280.89 ± 1.61** | 304.46 ± 3.08** | |
Ua (mg/L) | Control group | 33.80 ± 0.75 | 36.62 ± 0.94 | 38.73 ± 0.92 |
High F group I | 34.68 ± 0.97 | 39.79 ± 0.84** | 40.66 ± 0.67** | |
High F group II | 39.08 ± 1.30** | 42.72 ± 0.80** | 46.24 ± 0.92** | |
High F group III | 42.02 ± 0.84** | 47.72 ± 1.31** | 48.83 ± 0.97** | |
AST (U/L) | Control group | 37.86 ± 2.85 | 40.05 ± 1.64 | 43.51 ± 1.96 |
High F group I | 40.94 ± 1.59 | 46.56 ± 1.90** | 62.76 ± 1.29** | |
High F group II | 41.76 ± 1.37* | 48.79 ± 1.70** | 68.04 ± 1.91** | |
High F group III | 53.94 ± 2.49** | 68.13 ± 2.87** | 70.13 ± 2.09** | |
ALT (U/L) | Control group | 2.50 ± 0.16 | 2.72 ± 0.09 | 6.21 ± 0.21 |
High F group I | 3.53 ± 0.14** | 4.04 ± 0.11** | 9.05 ± 0.74** | |
High F group II | 7.41 ± 0.42** | 8.25 ± 0.66** | 14.87 ± 0.56** | |
High F group III | 10.30 ± 0.34** | 13.93 ± 0.99** | 16.70 ± 0.92** |
Data are presented with the means ± standard deviation (n = 5). *p < 0.05, compared with the control group. **p < 0.01, compared with the control group.
[
F toxicity in animals is multifarious [
As a very active site of metabolism, the liver is especially susceptible to F intoxication [
The major route for the removal of F from the body is by the kidney [
Based on the results observed in the present study and the aforementioned discussion, it is concluded that F has accumulated in the blood circulatory system and dietary F in the range of 800 - 1200 mg/kg could significantly induce abnormalities of bone, liver and kidney, inhibit the synthesis of protein, enhance lipid peroxidation and disturb the antioxidative system of broiler chickens. Our data provide some information for clinical diagnosis of fluorosis and for further studying the mechanism of excessive F accumulation on the damage of soft tissues in broilers.
Yubing Deng and Hengmin Cui designed the study; Yubing Deng analyzed the data; Yubing Deng wrote the paper. This research was supported by the program for Changjiang scholars and the university innovative research team (IRT 0848), and the Education Department of Sichuan Province (09ZZ017).