Assessment of some heavy metals in waste water and milk of animals grazed around sugar cane plants in Sudan

This investigation was conducted to evaluate the heavy metal concentration of waste water from sugar cane plants and milk from animals grazed around the plants. A total of 106 samples of waste water and 94 samples of milk (cow, goat, sheep) were collected from Guneid, Sinnar, Assalaya, Kenana and New Halfa sugar cane plants. The waste water and milk samples were subjected to heavy metal analysis (Cu, Zn, Pb, Cd, Cr, Co) by flame atomic absorption spectrophotometer.

Results indicated that Zn and Cu were detected in the waste water of all sugar cane plants except Assalaya and Kenana respectively, Pb in the waste water of Kenana and New Halfa, Cd in the waste water of Guneid and Sinnar, Co in the waste water of Guneid and Cr in the waste water of all plants. Cu and Zn were detected in the milk of all plants, Pb in the milk of all plants except New Halfa, Cd in the milk of Guneid, Sinnar and Assalaya, Cr in the milk of Guneid, Co in the milk of Guneid and Sinnar. Heavy metals were detected in the milk of all species under study in varying concentrations except Cr which was not detected in sheep milk.

Key words: cadmium, chromium, cobalt, concentration, copper, cow, goat, lead, sheep, zinc

Introduction

Milk is nearly a complete food for its good source of protein, fat and minor minerals, being a complex bioactive substance that promotes growth and development of the infant animals. Milk is an ideal source of macroelements such as Ca, K, P, in addition to microelements such as Mn, Cu, Fe, Se and Zn and even heavy metals can be found (Sikiric et al. 2003; Qin et al. 2009). The amount of metals in uncontaminated milk is minute, but their contents may be significantly altered through manufacturing and packaging as well as metals that may be contaminated from different cattle feeds and environment such as Pb, Cd, Cr, Ni and Co (Enb et al. 2009).

World-wide contamination of milk with undesirable substances via animal feeds, heavy metals, mycotoxins, dioxins and similar pollutants is considered to be of great concern to public health due to their toxic effects on human and wildlife. Despite the essential benefits of consuming milk, the contamination of milk from moderate agricultural practices, industrial pollutants in the environment, animal feeds and use of sewage sludge in agriculture is increasing and therefore requires urgent attention (Jigam et al. 2011).

Heavy metals are described as those metals which, in their standard state, have a specific gravity (density) of more than 5 gm/cm³, and atomic weight of 63.546-200.590 (Aslam et al. 2011). The heavy metals are responsible for many pernicious effects on human health such as saturnisme (lead contamination), immunodepression and skin diseases (zinc and copper contamination), cancer (cadmium), hyperkeratosis (arsenic), neurological disorders (manganese) or blood disorders (iron) (Konuspayera et al. 2009).

Many studies indicate the presence of heavy metals in milk such as Cu, Zn, Pb, Cd, Cr, Co, Mo and As (Sikiric et al. 2003; Vidal et al. 2004; Dobranski et al. 2005; Valiukenaite et al. 2006; Jigam et al. 2011; Ogabiela et al. 2011). The toxic metal content of milk and dairy products is due to such factors as environmental conditions and manufacturing processes (Anastasio et al. 2006).

Foodstuffs grown on agricultural soil or irrigated with impure water accumulate metal contents and are a big source of heavy metals exposure to the animals and humans. These metals are toxic in nature and even at relatively low concentrations can cause adverse effects (Aslam et al. 2011). Many dangerous elements or compounds such as metals and metalloids accumulate along the food chain, and their concentration in the environment grows with the increase of urban, agricultural and industrial emissions (Birghila et al. 2008). Most of our water resources are gradually becoming polluted due to the addition of foreign materials from the surroundings, and these include organic matter of plant and animal origin, land surface washing and industrial and sewage effluents (Lokeshwari and Chandrappa 2006). Due to the growing environmental pollution it is necessary to determine and monitor the levels of toxic metals in waste water and milk , such as lead and cadmium, because they can significantly influence the human and animal health (Sikiric et al. 2003).

Sugar industry in the Sudan started with the establishment of the Guneid sugar plant in 1962. There are now five sugar plants in the country, four of these are state-owned; the Guneid, the New Halfa, the Sinnar and the Assalaya plants, while the fifth, the Kenana plant, is a joint venture with Sudanese, Arab and other capital; and a new project (White Nile sugar project) is under establishment. Sugar cane plants dispose waste water from cane processing into either main canals or rivers, and while some plants started to treat this water before getting rid of it, others do not carry out any treatment practices to the water, and if this water is disposed in irrigated areas might cause serious hazards to human and animals grazing on the vegetation.

The objective of the study is to evaluate the heavy metals content (Cu, Pb, Zn, Co, Cr, Cd) of waste water from the sugar cane factories and the milk of animals grazing around the factories.

Materials and methods

Study area

The study area included the following five subareas where the sugar cane plants are located. The milk sample collection was from an area of approximately 1-2 Km radius within each sub-area.

All these plants are located in low rainfall savanna which comprises 18.5% of the Sudan’s area with mean annual rainfall ranging from 300 to 800 mm. Irrigated agriculture, rain-fed traditional cultivation and mechanized farming are practiced in this zone.

Sample collection

A total of 106 samples of waste water and 94 samples of milk were collected from gazing areas around sugar cane plants as follows:

The samples were aseptically collected in sterile glass bottles and transported to the laboratory in ice box at ≤5°C and kept in the refrigerator at this temperature till analysis was carried out within 24 hr.

Milk sample preparation for heavy metals determination

Five grams of milk samples were heated on a steam bath, followed by oven drying at 105ºC for 3 hr to remove the moisture from the sample. The dried sample was weighed and placed in a muffle furnace at 550^oC for 4 hr till the sample was converted into ash, and 5 ml hydrochloric acid (HCl, 5N) were added and heated on sand bath for 10 min, dissolved in 30 ml deionized water and filtered with Whatman filter paper No. 42. The solution was made up to the volume in 100 ml flask and stored in glass bottle at ≤5ºC till analysis (Ryan et al. 2001).

Sample analysis

Heavy metals analysis of milk and waste water

Heavy metals were determined by flame atomic absorption spectrophotometer (model 3110, Perkin Elmer Corporation, USA) in a flame acetylene-air. Standard atomic absorption conditions, wavelength and detection limits of the heavy metals are presented in Table 1.

Statistical analysis

The statistical analysis was performed using Statistical Analysis Systems (SAS, ver. 9). The results were evaluated by a multiple-variance analysis, using ANOVA. Means were separated by Duncan multiple range test at P≤0.05.

Table 1: Standard atomic absorption conditions and detection limits of heavy metals
Heavy metal	Wavelength (nm)	Characteristic concentration (mg/L)	Linear range (mg/L)	Detection limit (mg/L)
Cu	328	0.770	5.0	0.001
Zn	214	0.018	1.0	0.001
Pb	283	0.45	20.0	0.01
Cd	229	0.028	2.0	0.001
Cr	357	0.078	5.0	0.003
Co	241	0.120	3.5	0.01

Results and discussion

Heavy metals concentration of waste water from sugar cane plants

The concentrations of heavy metals (Cu, Zn, Pb, Cd, Cr, Co) in waste water from sugar cane plants (Guneid, Sinnar, Assalaya, Kenana, New Halfa) are presented in Table 2. The concentration of Cu was high (0.107 mg/L) in Guneid and low (0.0013 mg/L) in New Halfa, and below detection level (BDL) in Kenana. Zinc (Zn) was in its highest concentration (0.723 mg/L) in Sinnar and BDL in Assalaya. Lead (Pb) was highest in Guneid (0.0385 mg/L) and BDL in Kenana and New Halfa. Cadmium (Cd) was only detected in Sinnar (0.0138 mg/L) and Guneid (0.0040 gm/L) and Chromium (Cr) was BDL in all plants. Cobalt (Co) was only detected in Guneid (0.0100 mg/L).

The results of heavy metal concentration in this study are far below that reported by Selladurai et al. (2010) who reported the heavy metal concentrations of distillery sludge from sugar cane industry to be as follows: 2.72±0.1 – 2.94±0.4 mg/L for Zn, 1.53±3.5 -1.94±1.5 for Cu, 0.29±0.5 – 0.83±0.1 for Cr, 0.18±0.9 – 0.20±0.5 for Cd and 0.24±0.6 – 0.29±0.8 for Co, and that even after treatment of the sludge in bioreactor inoculated with Eudrilus eugeniae and Eiseria foetida earthworms, the concentration was not decreased significantly. This study is in accordance with previous researchers who detected Zn, Cu, Cd and Co in industrial effluents which h resulted in increased level in both the drain and groundwater (Kadem et al. 2012). Lokeshwari and Chandrappa (2006) reported that the average heavy metal concentration (μg/L) in the rainy and dry seasons in lake water was 179 and 113 (Zn), 17 and 16 (Cu), 18 and 5 (Cr), 13 and 6 (Pb) and 0.2 and 3.3 (Cd), respectively.

Table 2: Heavy metals concentration (mg/L) in waste water from sugar cane plants
Sugar cane plant	Heavy metals (mg/L)
Sugar cane plant	Cu	Zn	Pb	Cd	Cr	Co
Guneid	0.107^a	0.138^a	0.0385^a	0.0040^b	BDL	0.0100
Sinnar	0.0352^b	0.723^a	0.0209^ab	0.0138^a	BDL	BDL
Assalaya	0.0076^c	BDL	0.0057^b	BDL	BDL	BDL
Kenana	BDL	0.366^a	BDL	BDL	BDL	BDL
New Halfa	0.0013^c	0.487^a	BDL	BDL	BDL	BDL
SEM	0.014	0.626	0.024	0.0005	-	0.0002
p	<0.0001	0.482	0.0202	0.0003	-	-
Mean values in the same column without common letter differ at P<0.05 SEM. = Standard error of means BDL = Below detection level

Heavy metals concentration of milk from animals grazing around sugar cane plants

The milks of cows, goats and sheep grazing in the areas around sugar cane plants were subjected to heavy metals determination. The results show that Cu concentration ranged between 0.0033 mg/L in Kenana to 0.384 mg/L in Guneid; Zn ranged between 0.269 mg/L in New Halfa and 2.12 mg/L in Assalaya; Pb was below detection level in New Halfa and 0.156 mg/L in Guneid; Cd was below detection level in Kenana and New Hallfa and 0.0085 mg/L in Assalaya; Cr was only detected in Guneid (0.0045 mg/L); Co was detected in Sinnar (0.0273 mg/L) and Guneid (0.0135 mg/L) (Table 3). Although there was no significant difference in the concentration of Cu and Cr in the milk of three species under investigation, the highest concentration of Cu was in cow milk, while the highest concentration of Cr was in goat milk. Zn was significantly higher in goat milk; Pb was higher in cow milk; Cd was higher in sheep milk; Co was higher in cow milk (Table 4).

There are important factors influencing the occurrence and concentration of the elements including the toxic ones in milk and these are; the environmental pollution, industrial emission, waste water, some other factors associated with the contamination of vegetation from soil and water and feeding and factors related to animal handling by humans (Sikiric et al. 2003; Selladurai et al. 2010; Kadem et al. 2012).

The concentration of copper (Cu) was high in cow and goat milks and low in sheep milk. Similar results were reported by Enb et al. (2009) in cow milk, while the results are lower than the findings reported in previous studies (Hejtmankova et al. 2002; Sikiric et al. 2003; Vidal et al. 2004; Dobrzanski et al. 2005; Qin et al. 2009; Jigam et al. 2011). Abnormal concentration of Cu in tissues and blood causes Wilson disease, and acute exposure to Cu causes vomiting, bloody diarrhea, hypertension and cardiovascular collapse (Ogabiela et al. 2011), and the occurrence of Cu in milk is likely to be influenced by the environment (Qin et al. 2009).

The concentration of zinc (Zn) was in the range of 0.237 mg/L in sheep milk to 1.24 mg/L in goat milk. These results are similar to the findings of Sikiric et al. (2003) who reported that the average concentration of Zn from 15 dairy farms in Croatia was 0.51±0.30 mg/Kg, and higher than that reported by Jigam et al. (2011) in Nigeria (0.33±0.10 mg/L). However, many researchers reported Zn concentration higher than that reported in this investigation (Enb et al. 2009; Qin et al. 2009; Ogabiela et al. 2011). Chronic Zn exposure results in anaemia, leucopemia, gastrointentinal diseases and diarrhea (Ogabiela et al. 2011).

Table 3: Heavy metals concentration (mg/L) in milk collected from animals grazed around sugar cane plants
Heavy metal (gm/L)	Sugar cane plants					SEM	p
Heavy metal (gm/L)	Sinnar	Guneid	Assalaya	Kenana	New Halfa	SEM	p
Cu	0.0677^b	0.384^a	0.0321^bc	0.0033^c	0.0588^bc	0.0404	<0.0001
Zn	0.814^ab	0.172^b	2.12^a	1.37^ab	0.269^b	0.897	0.0155
Pb	0.0536^b	0.156^a	0.0295^b	0.018^b	BDL	0.0375	<0.0001
Cd	0.0027^b	0.0035^ab	0.0085^a	BDL	BDL	0.0004	0.0079
Cr	BDL	0.0045	BDL	BDL	BDL	-	-
Co	0.0273^a	0.0135^b	BDL	BDL	BDL	0.0002	<0.0001
Mean values in the same row without common letter differ at P<0.05 BDL = below detection level

Table 4: Heavy metals concentration (mg/L) in cow’s, goat’s and sheep’s milk grazed around sugar cane plants
Heavy metal (gm/L)	Animal species			SEM	p
Heavy metal (gm/L)	Cow	Goat	Sheep	SEM	p
Cu	0.124^a	0.111^a	0.0918^a	0.0521	0.25
Zn	0.971^a	1.24^a	0.237^a	1.14	0.29
Pb	0.502^a	0.0748^a	0.0307^a	0.0485	0.23
Cd	0.0006^b	0.0059^a	0.0071^a	0.0005	0.019
Cr	0.0006^a	0.0021^a	BDL	0.0003	0.16
Co	0.0122^a	0.0062^b	0.005^b	0.0002	<0.000
Mean values in the same row without common letter differ at P<0.05 BDL = Below detection level

Lead (Pb) concentration in milk ranged between 0.502 mg/L in cow milk and 0.0307 mg/L in sheep milk. Lead is a toxic element often associated with traffic pollution. Similar results were reported by Sikiric et al. (2003), while Enb et al. (2009) and Qin et al. (2009) reported Pb concentration higher than that reported in this investigation. However, our findings are far below the level reported by Vidal et al. (2004), Jigam et al. (2011) and Ogabiela et al. (2011). Aslam et al. (2011) reported that goat milk contained 43.414 mg/L of Pb, and Hejtmankova et al. (2002) reported higher heavy metal content in goat milk from farms located near the natural reserve than in milk from farms located near areas of high contamination from industrial sources. They attributed this to the geological composition of the specific area especially the feed seems to be more decisive for the resultant contents of contaminating chemical elements in milk than the amount of industrial and traffic emissions in the area. Cadmium (Cd) content was as low as 0.0006 mg/L in cow milk and 0.0071 mg/L in sheep milk. Previous studies reported higher concentration than those reported in this investigation [Hejtmankova et al. 2002 (1.84 mg/Kg for goat milk), Enb et al. 2009 (0.086±0.062 mg/Kg), Aslam et al. 2011 (0.147±0.003 mg/L for cow milk and 0.145±0.002 mg/L for goat milk), Ogabiela et al. 2011 (0.131 mg/L)]. Chromium (Cr) content was between 0.021 in goat milk and BDL mg/L in sheep milk. The concentration of Cr in this study is below that reported by Dobranski et al. (2005), Enb et al. (2009), Qin et al. (2009) and Aslam et al. (2011). Ogabiela et al. (2011) reported that the high concentration of Cr in cow milk could be attributed to discharges released from tanneries. Chromium is responsible for stimulating the activities of insulin and also helps to control blood cholesterol level. Cobalt (Co) content was between 0.005 mg/L in sheep milk and 0.0122 mg/L in cow milk. The concentration of Co reported in this study is higher than that reported by Enb et al. (2009) who reported a concentration of 0.004±0.001 mg/Kg, and lower than that reported by Dobrzanski et al. (2005).

Comparison of heavy metals concentration in milk and waste water

All heavy metals were significantly higher in milk compared to waste water, except Cd which was slightly higher in waste water although the difference was not significant (Table 5). The results may clearly indicate that the source of contamination of milk by heavy metals is not only the waste water from sugar cane industry, it rather may be due to the contamination of vegetation from either fertilizers, insecticides or from pollution of car combustion. Qin et al. (2009) reported that the content of Cu, Cd and Pb in milk is likely to be through environmental and atmospheric deposits-soil-cattle feed-milk chain. Combustion of Pb and its emission into the atmosphere can be responsible for high concentration of Pb in some vegetation, roadside, soil, air, water and plants, in addition, contamination may come from industrial wastes as well as application of fertilizers (Enb et al. 2009).

Table 5: Heavy metals concentration (mg/L) in waste water and milk of animals grazed around sugar cane plants
Heavy metals	Waste water	Milk	SEM	p
Cu	0.0289^b	0.116^a	0.0514	<0.0001
Zn	0.348^b	0.946^a	1.19	0.0137
Pb	0.0126^b	0.0549^a	0.0511	0.0003
Cd	0.0035^a	0.0032^a	0.0007	0.68
Cr	BDL	0.0009	-	-
Co	0.0019^b	0.0093^a	0.0005	<0.0001
Mean values in the same row without common letter differ at P<0.05 BDL = Below detection level

Conclusions

Acknowledgements

The financial support of the Ministry of Higher Education and Scientific Research is appreciated. The authors would like to thank the officials in the sugar cane plants and the owners of the animals for their help during the collection of water and milk samples.

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