Distribution of Selenium and Mercury in Heilongjiang Province and Its Effect on Body of Beef Cattle

Guo Zhao-zhou, Cui Hong-xia, Dong Na, Wu Hong-zhi, Song Xin-fa, and Xu Li

College of Animal Sciences and Technology, Northeast Agricultural University, Harbin 150030, China

Abstract: The aim of this study was to study the distribution of Selenium (Se) and Mercury (Hg) in feed and drinking water in Heilongjiang Province and the effect on body of beef cattle. The experiment selected four areas of Qiqihar, Harbin, Jixi and Hegang Cities in Heilongjiang Province. In each area, a pasture was selected. Five beef cattle from each ranch were collected. At the same time, blood, liver, muscle, kidney, urine and faece of the cattle were collected, as well as drinking water, feed and formula feed from the ranch. The contents of Se and Hg in the collected samples were detected by inductively coupled plasma mass spectrometry. The results showed that the contents of Se in liver, kidney, urine and blood of beef cattle were increased with the increase of Se contents in drinking water, while the contents of Se in muscle tissue and manure were decreased. When the contents of Se in feed increased, the contents of them in muscle, manure and liver of beef cattle were increased first and then decreased, while the contents of Se in kidney,urine and blood were decreased first and then increased. When Hg contents in drinking water were increased, the contents of them in beef cattle tissue, blood and manure were increased first and then decreased, and the contents of Se in urine were increased first and them decreased. And when Hg contents in feed increased, the contents of Se and Me were also increased. In conclusion, Se and Hg contents in these areas were within the safety standards, meanwhile, the contents of Se and Hg in body tissues of beef cattle were affected to varying degrees by the contents of selenium and mercury in feed.

Key words: Selenium, Mercury, north temperate zone, beef cattle tissue, feed, drinking water

Introduction

In the process of growth and development of animals,a variety of trace elements and some low dose of heavy metals are needed to maintain their health and production performance. Different proportions and quantities of various kinds of elements can provide indications of the nutritional status of the animals. In the physiological process of beef cattle growth and development, reproduction and health, the deficiency and excess of certain trace mineral elements can cause the growth of beef cattle to be blocked, so that the economic benefit of the farm is damaged.

Selenium (Se) in human selenoproteins has at least 25 selenocysteine forms, it is found that rare earth amino acids involved in redox reactions, such as glutathione peroxidase and thiooxazole, when presented at the active site of the enzyme (Legrain et al., 2014; Pedrero and Madrid, 2009). It is an essential micro-mineral for animals and can reduce the incidence of metritis and ovarian cysts as well as the interval from calving to conception in lactating dairy cows (Harrison and Conrad, 1984; Werechiga et al.,1994). It also manages immune functions (Salman et al., 2009), and plays a major role in the formation and the activity of helper T and natural killer (NK)cells (Petrie et al., 1989). Se is present in all the cells and tissues. In animal visceral tissues, such as liver,kidney, pancreas and other organs. There is a higher Se contents, while in bone, muscle and blood, Se contents are low. (Lu, 2016). However, when Se is deficient, it will slow down the rate of growth (Gleed et al., 1983), reduce fertility (McClure et al., 1986)and the immune responses of animals (Erskine et al.,1989; Boyne and Arthur JR, 1979). Two thirds of counties in China are with low or no Selenium, but in Yunnan Province, Hunan Province and other provinces, they are considered as potentially available Selenium-rich areas. The areas where in China with high Selenium contents are mostly in the southeast and northwest areas, and the areas in the central area are relatively low (Wu and Chou, 1996). And for Mercury(Hg), it is a toxic metal associated with various health problems, including cardiovascular disease,developmental neurotoxicity, thyroid dysfunction and liver and kidney dysfunction (Clarkson et al.,2007; Houston, 2011; Karagas et al., 2012). When Mercury enters human and animal bodies, different kinds of Mercury accumulate in different parts of the bodies. For example, metallic Mercury accumulates mainly in kidney and brain. Inorganic Mercury accumulates mainly in kidney, while organic Mercury accumulates mainly in blood and the central nervous system. Another study shows that, Methylmercury(MeHg) and Se, Hg are closely linked, the two elements can be through MeHg or direct interaction,to maintain the balance of the body (Berglund et al.,2005, Carrier et al., 2001; MacDonald et al., 2015;Turner and Swick, 1983). In China, Mercury is more concentrated in coal. Among them, Mercury content is relatively low in the north and a part of the central region, including Heilongjiang Province,Sichuan Province and Xinjiang Province, etc (Yang et al., 2015). However, the world-class Mercury deposits are distributed in southern China, such as Guizhou, Hunan, etc. The distribution of bedded Mercury and Mercury content in the adjacent areas are much higher than the average surface abundance(Hua, 1985).

Most parts of China belong to the northern temperate zone, which are suitable for ruminant breeding. Tibet,Xinjiang, Inner Mongolia, and northeast area due to their geographical locations, which are suitable for breeding large ruminants. In this experiment, the influence of Selenium and Mercury on beef cattle,the contents of drinking water and feed of Selenium and Mercury on four different beef cattle farms in the temperate zone were studied. These four regions were respresentation in Heilongjiang Province, accounting for the total amount of Heilongjiang Province, as much as 40% of the total population. And they were located in the four locations in Heilongjiang Province. Five beef cattle were selected from each beef cattle farm.By studying the relationship between Selenium and Mercury in feed and drinking water, and Selenium and Mercury in the body tissues of beef cattle, this study provided some references for the rational application of trace elements of Selenium and Mercury in the beef temperate zone in the north temperate zone.

Materials and Methods

Sample collection

Drinking water was collected from four beef cattle farms in Heilongjiang Province, of which there were three samples of tap water, and the surface water of one sample. Feed samples were collected at the same farms in Heilongjiang Province, a total of 29 samples, including four samples of corn straw, four samples of vinasse, two samples of distillers dried grains solubles (DDGS), three samples of corn, three samples of wheat bran, two samples of soybean meal, three samples of bean cake, four samples of concentrate supplement material, four samples of the total mixed rations (TMR). The all-day urine was collected by urine bag for three days, then 5%of the total amount of urine was passed through four layers of gauze filter, after that, 0.5 L urine sample was stored at –20℃ refrigerator. 5% of the total amount of feces was accurately weighted from the all-day feces which was collected and mixed every day. Then, 10% of sulfuric acid solution was sprayed on the surface of the feces samples. After that, the sample was placed in the refrigerator to keep cold until analyzed. The action of collecting the feces sample would be continued for three days. When the sampling was finished, the 3-day feces was mixed and accurately weighted 2 kg, and then it was placed in the oven at a constant temperature of 65℃ to be dried, sifted through a 40 mesh sieve to prepare a dried sample, then it was stored into a clean valve bag, and stored at a shade place until analyzed. Ten mL blood was taken from the caudal vein, then the blood sample was applied of EDTA as anticoagulant, upside down and mixed, saved at –20℃. When all the cattles were slaughtered, the samples of muscl, liver and kidney were collected back for 1 kg of each of them, then they were also stored into a clean valve bag and cryopreserved under –20℃.

Instruments and reagents

Instrument: ICP-MS 7500 inductively coupled plasma mass spectrometer (Agilent companies in the United States), CEM MARS microwave digestion instrument(CEM companies in the United States), preparation of instrument, electronic balance.

Reagent: nitric acid, hydrogen peroxide, pure water, Se element standard liquid, Hg element standard liquid.

Determination method

For sample pretreatments, the samples of feed, tissues and feces were accurately weighted 0.4 g respectively in the digestion tank, 8 mL 65% nitric acid was added, after that, the soaked samples were slightly shaken, covered the safely valve, heated at 1 200 W,the temperature was uniformed to 120℃ within 5 min,held for 3 min; then the temperature was uniformed to 160℃ within 5 min, held for 3 min, and then a temperature of 10 ℃ · min-1was taken to rise to 180℃, held for 15 min. When the digestion tank was cooled to room temperature, the digestion solution was moved into a 50 mL volumetric flask, scaled with water volume, then mixed uniformity and standby. The samples of water, urine and blood were accurately volumed 0.25 mL respectively in the digestion tank,8 mL 65% nitric acid was added, according to the process of digestion, then the temperature was uniformed with a temperature of 5℃ · min-1to 150℃ within 5 min,2 500 kPa pressure; in the 5th min to 14th min, the temperature of digestion tank was heated to 160℃,the pressure and heating rate were the same as the first stage; in the 14th min to 29th min, a temperature of 5 ℃ · min-1was taken to heat to 180℃, 3 000 kPa pressure; in the 29th min to 39th min, a temperature of 2 ℃ · min-1was taken to heat to 200℃, 2 000 kPa pressure. After that, the digestion solution was moved into a 50 mL volumetric flask, scaled with water volume, then mixed uniformity and standby.

All the samples were analyzed by inductively coupled plasma mass spectrometry. Specific parameters were chosen as the followings: RF power was 1 150 W, plasma flow rate was 20 L · min-1, auxiliary air flow was 0.5 L · min-1, atomization air pressure was 0.24 MPa, flushing time was 30 s, washing pump speed was 130 r · min-1, analysis pump speed was 130 r · min-1, and the detection wavelength was 220 nm.

Data collation and statistics were analyzed by using SPSS 20.0, and represented by "mean±standard deviation". The single factor analysis of variance(ANOVA) and least significant difference (LSD) were chosen to multiple comparisons of different beef cattle breeding places between Selenium content in samples collected.

Results

Contents of Selenium and Mercury in drinking water, soil and feed

The contents of different pasture Selenium in feed,water and soil are presented in Table 1. The content of Se in corn stalk was between 0.02 and 0.72 mg · kg-1;Selenium content in DDGS was between 0.03 and 0.34 mg · kg-1; pasture one was slightly lower than the corresponding value 0.4 mg · kg-1of NRC beef cattle feed ingredients. While pasture four was higher than the corresponding value. As for wheat bran, all the pastures were lower than NRC beef cattle feed ingredients value 0.83 mg · kg-1. The contents of Se in soybean meal at all pastures were higher than NRC beef cattle feed ingredients value 0.22 mg · kg-1. At all the pastures of Se content in drinking water were between 0.04 and 2.1 μg · L-1. Compering with the feed composition of NRC beef cattle, the content of Se in samples such as distillers' grains was different.Concentrated supplement material was the cattle pastures basing on requirements to purchase a feed of semi-finished products, so its Se content was relatively high. Mixed with roughage, it could achieve the recommended amount of beef cattle feeding standard 0.1-2 mg · kg-1, the content of Se was between 0.11 and 0.41 mg · kg-1in TMR was also a direct proof of this point.

The contents of different pastures Mercury in feed, water and soil are presented in Table 2. The content of Hg in corn stalk was between 0.003 and 0.016 mg · kg-1; Mercury content in DDGS was between 0.001 and 0.002 mg · kg-1; as for wheat bran,all the pastures were lower than the GB13078-2001's value 0.1 mg · kg-1. The contents of Hg in soybean cake at all the pastures were lower than the GB13078-2001's value 0.1 mg · kg-1. At all the pastures of Hg content in drinking water were between 0.004 and 0.01 μg · L-1, and there was no more than 0.01 mg · kg-1of HJ568-2010. And in soil, the contents of Mercury in all the pastures were not exceed the GB15628-1995's secondary standards. The content of Hg of concentrate supplement material was between 0.002 and 0.02 mg · kg-1, among them, the content of Hg of concentrate supplement material in pasture two was higher than that of the GB13078-2001's value. Mixed with roughage, it did not exceed the limit imposed by the GB13078-2001, and the mixed feed could be safely fed to beef cattle. TMR content of Hg was between 0.003 and 0.004 mg · kg-1.

Content of Selenium in manure and urine of beef cattle

The content of different pastures Selenium in cattle excreta is presented in Fig. 1. The content of Se in each pasture cattle urine was between 0.001 and 0.067 mg · L-1,the numerical range was relatively large, among them,pasture one was the highest, and the difference was very significant (p＜0.01); the content of Se in each pasture cattle feces was between 0.109 and 0.588 mg · kg-1,pasture one was the highest, pastures one and three were higher than pasture four, and the difference was significant (p＜0.01). Selenium contents in beef cattle feces were far higher than those of urine.

Table 2 Content of Mercury in feed, water and soil of different pastures (mg·kg-1)

Fig. 1 Selenium content in fecaluria of beef cattle in different pastures (n=5 for each pasture, mean±SD)*P＜0.05, **P＜0.01.

Content of Mercury in manure and urine of beef cattle

The contents of different pastures of Mercury in cattle excreta are presented in Fig. 2. The results showed than that the content of Mercury in urine was higher than that in feces at four pastures.

The content of Hg in each pasture cattle urine was between 0.042 and 0.4 µg · L-1, pasture one was the highest, and the difference was very significant(p＜0.01); the content of Hg in each pasture cattle feces was between 0.011 and 0.024 mg · kg-1, the content of Hg in pasture three was the highest and the difference was significant (p＜0.01).

Fig. 2 Mercury content in fecaluria of beef cattle in different pastures (n=5 for each pasture, mean±SD)*P＜0.05, **P＜0.01.

Content of Selenium in blood and tissues of beef cattle

The contents of different pastures Selenium in beef cattle organization are presented in Fig. 3. The content of Se in each pasture cattle blood and kidney was between 0.051 and 0.206 mg · L-1, 0.198 and 1.332 mg · kg-1,respectively. Among them, pasture one was the highest, and the difference was very significant (p＜0.01);Selenium content in liver and muscle was between 0.126 and 0.624 mg · kg-1, 0.080 and 0.285 mg · kg-1,respectively. For liver and muscle, the content of Se according to the order from high to low was pasture two＞pasture one＞pasture three＞pasture four,pasture four＞pasture three＞pasture two＞pasture one,respectively. And for blood and kidney, the content of Se according to the order from high to low was pasture one＞pasture three＞pasture two＞pasture four,pasture one＞pasture three＞pasture two＞pasture four,respectively.

Content of Mercury in blood and tissues of beef cattle

The content of different pastures Mercury in beef cattle organization is presented in Fig 4. From the image, Mercury content in the blood, liver, kidney and muscle of each pasture beef cattle was between 0.08 and 0.75 μg · L-1; 0.6 and 1.6 μg · kg-1; 0.48 and 1.8 μg · kg-1;0.13 and 1.1 μg · kg-1, respectively. The range of values was relatively large, among them, pasture three was the highest and pasture one was the lowest, which was significantly different from other pastures (p＜0.01).

Fig. 3 Selenium content in tissues of beef cattle in different pastures (n=5 for each pasture, mean±SD)*P＜0.05, **P＜0.01.

Fig. 4 Mercury content in tissues of beef cattle in different pastures (n=5 for each pasture, mean±SD)*P＜0.05, **P＜0.01.

Correlation analysis of Selenium and Mercury content between dung, tissue and feed, drinking water

The correlation analysis of Selenium content in rearing environment and excreta and tissues is presented in Table 3. The content of Se in cattle feces and urine and tissue showed a positive correlation between feed and drinking water content of Selenium, among them, liver,kidney and blood Selenium content reached significant level (p＜0.05) with increasing Se content. Blood and kidney content Selenium had a significant positive correlation with drinking water. The correlation was also appropriate for liver, blood and urine to soil.

Table 3 Correlation analysis of Selenium content in excreta and tissues and rearing environment

The correlation analysis of Mercury content in rearing environment and excreta and tissues is presented in Table 4. There was a positive correlation between Mercury content in feces and tissues and Mercury content in feces, and Mercury content in feed was significant (p＜0.05). There was a significant positive correlation between Mercury content in muscle and liver and Mercury content in drinking water (p＜0.01). The Mercury content in urine was negatively correlated with Mercury content in feed and drinking water, and did not reach significant level.There was a negative correlation between Mercury content in kidney and feces and Mercury content in soil. Mercury content in urine and other tissues was positively correlated with Mercury content in soil,and the correlation between Mercury content in liver and Mercury content in soil was extremely significant(p＜0.01).

Average content of Selenium and Mercury in blood and tissues of beef cattle

The average content of Selenium and Mercury in different tissues of beef cattle is shown in Fig. 5. The order of Selenium in tissues was: kidney＞liver＞blood＞muscle, and the content of Selenium in kidney was significantly higher than that in other tissues(p＜0.01). The order of Mercury in tissues was:kidney＞liver＞ muscle＞blood, the content of Mercury in kidney was also higher than that in other tissues(p＜0.01).

Table 4 Correlation analysis of Mercury content in excreta and tissues and rearing environment

Fig. 5 Average content of Selenium and Mercury in different tissues (n=5 for each pasture, mean±SD)*P＜0.05, **P＜0.01.

Discussion

Distribution of Selenium and Mercury elements in soil, drinking water and feed

Se content of average topsoil was estimated to be 0.33 mg · kg-1on a world-wide basis (Kabata, 2011).Whilst the typical range was 0.01-2 mg · kg-1(Oldfield,2002), in this study, Se content of soil in four pastures,they were all near the average content. However,the contents of different pastures were different.Although the four pastures were in the same province,the differences in different regions were still very significant. Mercury was considered to be one of the most toxic contaminants, because it was harmful to the environment and public health (Rodriguez et al., 2017). So the control of Mercury content in soil was particularly important, according to the GB15628-1995, Hg contents of soil in four pastures were not more than national limit, did not affect the cultivation of crops, plants and animal body less affected by Mercury in soil. But it did not mean that there was no impact, although Mercury content did not exceed the limit, it was close to the upper limit of the limit. Therefore, more attention should be paid to the safety of the soil.

Drinking water with high concentrations of Selenium could increase Selenium intake (Barron et al., 2012).In recent years, various studies showed that Selenium concentrations in drinking water sample ranged from 0.07 to 1.12 µg · L-1and arsenic concentrations ranged from 0.3 to 10.7 µg · L-1(Guo and Wang, 2012).While the public water system in developed countries was generally within 10 μg · L-1of reference value(Nordberg et al., 2007). In this study, Se contents of drinking water in four pastures, were within 0.04-2.1 μg · L-1, and there was still some distance from the standards of developed countries. Pasture one was the highest content of Se in drinking water, but Se content of soil was not the highest one. There was no significant correlation between Selenium content in soil and drinking water in the same pasture. In water, Hg would affect the reproductive development of aquatic animals (Mohmood et al., 2016). For this purpose, the upper limit of Mercury content in drinking water was specified. The upper limit of EU Environmental Protection Agency was 1 μg · L-1, the upper limit of the United States was 2 μg · L-1, and the upper limit of China was 0.01 μg · L-1. In this study,Mercury content of drinking water in four pastures was within the standard and did not affect the physical health of beef cattle. Mercury content in pasture one and pasture four was zero, further indicating that there was less contamination of Mercury in the area without worrying about excessive Mercury levels in water.

Aforetime studies showed that supplementing inorganic Se in feed increased Se concentration in edible tissues of calves (Pavlata et al., 2001), lambs (Molnár et al., 1998), pigs and poultry (Leng et al., 2003).As for cattle, the recommended amount of mineral element Selenium in the beef cattle breeding standard(NRC, 1996) was 0.1 mg · kg-1and the maximum tolerance was 2 mg · kg-1. In this study, the content of Selenium in some feedstuffs was lower than that of the standard feed, and some others was higher than it, but the total mixed diets (TMR) obtained by mixing were 0.11-0.41 mg · kg-1, had reached more than the recommended amount of feeding standard of beef cattle, although the content of trace element manganese partial feed in the raw material shortage,the concentrate supplement added, could be fully complementary, in order to meet the needs of trace element manganese in beef cattle. According to the study of concentrate supplement and results of TMR and cost considerations, it was recommended that appropriate to a certain extent reduced the amount of Selenium added to the concentrate mixture. When Hg moved on the food chain, it entered into the environment through natural and anthropogenic sources and bioaccumulation, posing a special risk to animals with higher nutritional levels (Chumchal et al., 2011;Bond et al., 2015). The control of Mercury in feed was more important. In this study, Mercury content in feed was within the national standard, and the upper limit of Mercury content in the national standard was 0.1 mg · kg-1. It could be shown that feed ingredients and mixes in each pasture were not contaminated by Mercury metal and it could be fed directly to beef cattle.

Effects of Selenium and Mercury on beef cattle

In this study, the content of Selenium in urine of beef cattle was between 0.001 and 0.067 mg · L-1, the content of manganese in feces was between 0.109 and 0.588 g · kg-1, and the content of manganese in feces was nearly a hundred times that of urine. Facts clearly showed that a significant portion of endogenous Selenium was lost from the gastrointestinal tract. In each pasture, the Selenium content in liver and kidney of beef cattle was relatively high, but the content was low in muscle and blood. It could be seen that Selenium was mainly distributed in liver and kidney of beef cattle, and the content of muscle and blood was relatively low. In the report of Jotty et al (2009) they also pointed out the phenomenon. Urine was a good example of the determination of elements and inorganic Mercury. The number of more than 100 μg · L-1,resulting in neurological symptoms, and the concentration of more than 800 μg · L-1was often associated with death (Mehrdad et al., 2014). The content of Hg of urine sample in pasture one was the highest one in four pasture as 0.3 μg · L-1, and which was far enough to 100 μg · L-1, to a certain extent, the beef body was not subject to Mercury pollution. The amount of Mercury in feces of each pasture was lower than that in urine, further proving the health of the beef body.When the inorganic Mercury compound was absorbed into blood, the highest concentration (about 85%-90%)was found in kidney. Inorganic Mercury salts were absorbed and accumulated in the proximal tubules of kidneys (Park and Zheng, 2012). In this study, the content of Hg was mainly accumulated in liver and kidney, also confirmed this point.

The contents of Selenium and Mercury in beef cattle tissue and blood in different areas were large, but the contents of them in liver and kidney, or muscle or blood, increased with the increase of Selenium and Mercury contents in feed and drinking water, and maintained a positive correlation. However, when the Selenium content in soil increased, there was no obvious rule to follow.

The content of Selenium in drinking water was between 0.04 and 2.10 mg · L-1and TMR was between 0.11 and 0.41 mg · kg-1. Selenium in drinking water and feed had different effects on the distribution of Selenium in beef cattle. The Selenium in drinking water had more influence on the content of Selenium in kidney and blood of beef cattle. There was a significant positive correlation between Selenium content in kidney and blood and that in drinking water.This was consistent with the results of Loeschner et al (2014). However, there were relatively few reports on the impact of Selenium drinking water on body tissues. As for Mercury, drinking water and soil was mainly affected the liver and muscles (Tetsuya et al., 2003).

As for the correlation between feed and tissues,there was a significant positive correlation between the content of Selenium in liver, kidney and blood and that in feed. This was consistent with the results of the study that the total Se concentration of the whole blood and tissues was greater in those animals offered 0, 0.15, or 0.35 mg of Se · kg-1of DM, when compared with those receiving a comparable dose of 0.15 mg of Se · kg-1of DM, indicating an improvement in Se availability and tissue Se retention (Juniper et al.,2008). Similar results were also obtained in male goats(Li et al., 2011), deer (Stoebe et al., 2015) and sheep(Čobanová et al., 2017).

For Mercury, there was a significant positive correlation between the contents of Mercury in blood,and muscle, and the content of Selenium in feed. In previous studies, the order of the least to be most contaminated Mercury in body tissues was: fat＜heart＜lung＜spleen＜muscle＜kidney＜liver＜blood (Rodrigues et al., 2014). The results of the beef cattle tissue measurements in the four pastures of the study also met this conclusion. There was no significant correlation between the contents of Selenium and Mercuryin feces and urine and those in feed and drinking water.

Conclusions

The present study indicated that all the pasture feed contents of Selenium were not exceed recommended quantity, in order to save resources, the amount of Se element in the supplementary material should be appropriately reduced. Different pastures had different contents of Selenium, for these four pastures,Se contents in beef cattle tissue from high to low were according to the order of kidney, liver, muscle and blood. In addition, the study of correlation was important for cattle and animal husbandry staff, its results showed that Selenium content in feed affected the deposition of Selenium in beef cattle. As for Mercury, the content in drinking water, soil and feed was lower than that of the standard, which showed that the beef cattle breeding environment of these pastures was relatively healthy and less polluted. Hg contents in beef cattle tissue from high to low were according to the order of kidney, liver, muscle and blood. It could be seen that it was more likely to accumulate in the kidneys and liver, whether it was Selenium or Mercury. At the same time, the kidney and liver of beef cattle were positively correlated with the contents of Selenium and Mercury in feed and soil. In the process of beef cattle breeding, the safety of feed and raw material soil should be paid more attention.