1. Introduction
Mongolia is a landlocked country located between Russian and China, known for its rich diversity of natural resources, including gold, silver, copper, coal, fluoride, iron, petroleum oil, and uranium [
1]. Traditionally, the main economic activity in Mongolia is based on agriculture, livestock, and the mineral industry. Mongolia’s economy has been expanding through exploitation of vast mineral resources during the past 20 years. The mining sector has supported economic growth and development in Mongolia, as mining recently contributed approximately 20% of gross domestic product (GDP) and about 80% of total exports in 2017 [
2,
3].
However, metal contamination derived from mining and industrial activity has been observed by many groups and has been reported worldwide [
4,
5,
6,
7]. For example, the area near mining site in Zambia recorded high concentration of metals including lead and cadmium, and geographic analysis revealed that the mining activity was possible source of pollution [
4]. Heavy metals represent one of the main environmental pollutants that cause adverse effects to human and animal health. These effects, such as neurotoxicity, hepatoxicity, carcinogenicity, teratogenicity, and nephrotoxicity, have also been reported [
8,
9]. The International Agency for Research on Cancer (IARC) has classified heavy metals such as arsenic, cadmium and nickel as Group1 (carcinogenic), and lead and cobalt as Group2B (possibly carcinogenic). While metals affect human health, many reports have also indicated heavy metal toxicity in animals. For instance, cadmium is highly toxic, affecting nearly all animal systems. Arsenic shows various toxicities with gastrointestinal and nervous symptoms in cattle and induced weight loss, reduction of milk yield, anemia, and liver and kidney damage by chronic exposure [
10]. Another consideration is that heavy metals cannot be degraded through physical processes. Therefore, they persist in the animal body and environment for a long period [
11].
Environmental contamination and exposure of humans to metals are practical concerns in Mongolia. Contamination with metals usually results from many different sources, such as industrial waste, chemicals used in agriculture, construction, vehicular exhaust, coal and fossil fuel combustion, and mining [
8,
12,
13]. In Ulaanbaatar, Mongolia, survey sampling in 2007 showed high arsenic contamination in soil. The possible source of this arsenic could be coal combustion and vehicles in the city [
12]. Another study of samples collected in 2019 also showed high arsenic contamination in soil in Ulaanbaatar, and arsenic and chromium have been evaluated as metals with high ecological risk [
14].
In recent years, environmental problems have gained attention in Mongolia. Economic losses because of metals through toxic effects on livestock and humans have also been found in other countries [
15,
16]. Concerning human health, various pathways through food, air, and water were considered as intake routes for metals, and animal products are one possible source of metals to humans [
8,
17]. Traditionally, Mongolians eat a lot of meats, and sheep meat is one of the main protein sources. The major livestock are sheep and cows for meat and milk in Mongolia. Mongolians eat not only muscle, but also liver, kidney, heart, etc. This unique food culture may increase the risk of metal contamination of animal products. As toxicity of metals in humans has been a concern in several countries, monitoring metals in grazing livestock in Mongolia is necessary in order to protect both animal health and human food safety. However, this kind of scientific research has not been pursued in detail in Mongolia at present. Therefore, monitoring environmental pollution that can cause toxicity to animals is desirable in Mongolia. Thus, this research targeted various livestock and soils in Mongolia, analyzing metal levels in collected samples.
2. Materials and Methods
2.1. Study Area
This study was conducted on animal farms in two areas: Dornogovi (Ulaanbadrakh: UB, Zuun-bayan: ZB and Airag: AE soum) and Tuv-Aimag (Zaamar soum). Dornogovi (44°53′N 110°09′E) is one of the 21 aimags (provinces) of Mongolia, located about 456 km southeast of the capital city, Ulaanbaatar. Dornogovi is known for its harsh weather conditions; it is located in the Gobi Desert, where frequent sand- and snowstorms occur. Tuv-aimag is another province of Mongolia, located about 45 km from the capital city. Many animal products, including milk and meat, are produced in both areas and distributed in Ulaanbaatar. Another notable product in Tuv-Aimag is gold from the mining located on the Tuul River. This area is still used for mining, and new mines are continuously being developed. Dornogovi also has mining areas, for fluorspar, uranium, etc. The soil and tissue samples were collected near mining areas. Details are shown in
Figure 1.
2.2. Sample Collection
All samplings were done in June, 2016, in Mongolia. In total, 53 tissue samples (liver and kidney) were collected from various deceased animals (horse, goat, sheep, and cow) in Dornogovi province for metal analysis. In addition, 55 soil samples were collected from the same area. Approximately 50 g of soil from a depth of 0–5 cm was collected for each sample and kept in a plastic tube. At least three composite soil samples were collected from each sampling point. Approximately 150 blood samples were randomly collected from various animals (horse, goat, sheep, cow, and camel) in Dornogovi and Tuv-Aimag. Collected tissue and blood samples were stored in cooler boxes with dry ice during the sampling on-site, and all samples were transported and kept at 4 °C for soil and at −20 °C for blood and organs at the School of Veterinary Medicine, Mongolian University of Life Sciences, before being transported and analyzed for metal concentration at the School of Veterinary Medicine, Hokkaido University, Japan around 20 days after sample collection.
2.3. Sample Preparation and Metal Extraction
All laboratory materials and instruments used in metal extraction were washed in 2% nitric acid (HNO
3) and oven-dried (50 °C). Metals were extracted from the samples (tissue, soil, and blood) by acid digestion with the method used in previous reports for tissue samples [
18] and environment samples [
19], with minor modifications.
Tissue samples (liver and kidney) were weighed (1 g) and dried for 48 h in an oven at 60 °C. Then, the dried tissue samples were digested using 5 mL 30% nitric acid (HNO3) and 1 mL 30% hydrogen peroxide (H2O2) (analytical grade, Kanto Chemical Corp. Japan) in a speed wave MWS-2 microwave oven system. Digestion of blood samples (1 mL) followed the same process as the tissue samples. The microwave system was programmed to ramp in five min to 160 °C (held for five min), followed by a ramp time of three min increased to 200 °C (held for 20 min), and then hold for 10 min at 75 °C. After digestion, samples were cooled and transferred to a 15 mL polypropylene tube followed by dilution to 10 mL with Milli Q water.
In the case of soil samples, extraction was performed using about 0.5 g of sample and 5 mL 60% HNO3, and 1 mL 30% H2O2. The microwave was programmed at an initial temperature of 150 °C (held for 5 min), then with a ramp time of 2 min increased to 175 °C (held for 10 min) and with a ramp time of three min increased to 190 °C (held for 30 min). After digestion, samples were filtered through ADVANTEC 5C filter paper into 15 mL polypropylene tubes and made up to 10 mL using Milli Q water. Analytical blanks were run for all samples.
Tissues, blood, and soil were assessed for concentrations of ten metals (Cr, Mn, Co, Ni, Cu, Zn, As, Se, Cd, and Pb) using an inductively coupled plasma –mass spectrometer (ICP-MS; 7700 series, Agilent Technologies, Tokyo, Japan) using 9Be, 115In, and 205Tl as internal standards.
2.4. Quality Control and Quality Assurance
All chemicals and standard stock solutions were analytical reagent grade (Wako Pure Chemicals, Osaka, Japan). For quality control, blanks were analyzed after every 10 samples, and a calibration curve for each element was constructed with an R2 value greater than 0.995. Analytical quality control was performed using certified reference materials: DOLT-4 (dogfish liver, National Research Council of Canada), DORM (fish protein, National Research Council of Canada, Ottawa, Canada), SRM-1944 (New/York/New Jersey Waterway Sediment, National Institute of Standards and Technology, New York, USA), and BCR-320 (channel sediment, Community Bureau of Reference of European Commission, Brussels, Belgium). Replicate analysis of these reference materials showed good recoveries, ranging from 90–110%.
2.5. Statistical Analysis
JMP 16 (SAS Institute, NC, USA) software was used for all statistical analyses, with a significance threshold of p < 0.05. The regional difference in soils was tested using Steel-Dwass tests. Outliers were detected assessing quantile range outliers (tail quantile 0.05, Q 10). Four metal concentrations in animal blood (from two individual outlier animals) were extracted. These two animals were also defined as the first and second biggest neighborhood distance calculated from K nearest neighbor outliers (K: 2–8, threshold was not decided). Thus, these two animals were excluded from the sample dataset. There was no outlier among soil samples, which were handled in the same way as animal samples.
3. Results
The metal concentration in the soil is shown in
Table 1 and
Supplementary Table S1. The Tuv-Aimag Zaamar region had generally higher concentrations of most metals; copper and chromium concentrations were significantly higher than in other areas (
p < 0.05) (
Supplementary Table S1).
Table 1 shows the median concentration of metals in the highest-concentration area, the highest concentration in this study, and the regulations (such as the maximum allowable level in Mongolia, maximum permissible level summarized in [
20] and Ecological Soil Screening Levels (Eco-SSLs) [
21,
22,
23,
24,
25,
26,
27,
28]). Eco-SSLs were provided from the U.S. Environmental Protection Agency (EPA). The Eco-SSLs suggest screening the level of contaminants in soil to indicate ecological risks. Contaminants exceeding their permissible concentration may require further evaluation for protection against ecological risk. The Eco-SSLs mention four different levels of concentration (plants, soil invertebrates, birds, and mammals); this study used its levels for mammals, as shown in
Table 1. Chiroma T. M et al. (2014) summarized the maximum allowable limit using World Health Organization (WHO), Food and Agricultural Organization (FAO), and Ewers, U. (1991), Standard Guidelines in Europe. Some of the highest concentrations of target metals exceeded these maximum levels. In particular, arsenic and selenium concentrations were high, and some soil samples have exceeded these levels (
Figure 2). For example, more than 50% of samples in Dornogovi AE exceeded the maximum permissible level of selenium (10 mg/kg) in Mongolia.
Blood metal concentration is described in
Table 2. Strong species differences were not observed for many metals in the same area. However, the camel accumulated a slightly higher concentration of zinc than the other animals in all areas. The median concentration of zinc in Dornogovi UB was 50.4 mg/kg (dry weight) in camels’ blood, while goats and sheep had blood levels of only 29.0 and 20.6 mg/kg, respectively. In the case of copper, the concentration in small ruminants, such as goat and sheep, was a bit higher than in other animals. The order of copper median concentration in Dornogovi ZB was goat > sheep > horse > camel > cow (
Table 2). Comparing the regional difference showed that the Tuv-Aimag Zaamar region had a higher concentration of arsenic than other areas, as the soil concentration had already suggested. However, in the case of zinc, selenium, and lead, the median concentration in soil in Dornogovi UB was the lowest, but comparatively high concentrations of metals were detected in blood in that area, as the median zinc and lead concentration (29.0 and 0.1 mg/kg dry weight, respectively) was the highest in goat among all areas.
To give a better understanding of the absolute concentrations of metals in livestock,
Supplementary Table S2 summarizes the normal range of metals in animal organs, referring to published values [
29]. The concentration in liver and kidney in sheep, goat, and cattle is shown in
Table 3. The sample number of organs was not enough to determine regional difference. Thus,
Table 3 shows mixed results from all areas (Dornogovi UB, ZB, AE). We also extracted the results of copper, zinc, and selenium in liver and kidney, as shown in
Figure 3, with toxic and deficient levels described in a previous report [
30]. The relatively high concentration of copper was also seen in sheep liver (median concentrations were 235.1, 92.4, and 148.3 mg/kg dry weight in sheep, goat, and cow, respectively). Arsenic concentration in organs was not as high compared with other high toxic metals, such as cadmium and lead (
Table 3).
We additionally collected only two horse kidney samples, but concentrations of cadmium (122.65 and 352.79 mg/kg) in kidney were quite high in both individuals, compared with other animals, as median concentration values in sheep, goat, and cattle were 0.28, 1.02, and 3.78 mg/kg respectively (data not shown).
To summarize our results, each metal concentration was normalized using its sum and standard deviation of all the samples. Then, the mean of each normalized concentration, as shown in
Supplementary Figure S1, was used to evaluate the accumulation pattern of each metal.
5. Conclusions
The international scientific report targeting metals in Mongolia was limited to the veterinary field. This paper revealed that arsenic and selenium were highly accumulated in Mongolian soil. They should be considered carefully as a risk, but the current concentration results in animal organs were not enough to conclude that animals suffer severe adverse effects because of exposure to these metals. This research also indicates that screening of all of soils, blood, and organs can be effective to monitor metals and to understand the inclusive polluted levels in Mongolia, as arsenic highly accumulated in soil but not in animals. For a better understanding of sources of heavy metal contamination, the monitoring of heavy metals in wild plant and livestock drinking water is desirable. We also focused on animal organs (liver and kidney), which are eaten in the countryside in Mongolia. Metals can be transferred to humans via animal products. Our results contribute to the protection of not only animal health, but also human food safety. This research will be beneficial to protect animal health, inform future studies targeting humans, and help reveal the source of metal contamination.