Effect of nano zinc supplementation on serum biochemical and mineral levels in Malabari kids fed on complete rations with varying energy levels #

Reddy ,


____________________________________________________________________________Reddy et al.
Zinc is the second most prevalent trace element in the body and is essential for normal physical, physiological and disease-free status of an animal . Zinc is also an important co-factor for more than 200 zinc-metalloenzymes like alcohol dehydrogenase, carbonic anhydrase, glutamic dehydrogenase, carboxypeptidase, glyceraldehyde-3-phosphate dehydrogenase, alkaline phosphatase, lactic dehydrogenase and many others enzymes (Riordan, 1976) which have a vital role in carbohydrate, protein and lipid metabolism. Several studies have shown that zinc supplementation in the feed has increased nutrient digestion because of improved secretion and activation of digestive enzymes (Hu et al., 2012;Brugger and Windisch, 2016;Lee et al., 2021). It also helps in improving osteoblastic activity, calcium deposition, cell division, bone growth and proper functioning of endocrine system and hormonal regulation.
The prevalence of zinc deficiency among livestock is twenty-five per cent (Goklaney et al., 2019) in India. Poor bioavailability, interaction with minerals and poor storage makes it more critical and a continuous supply through feed is required. Zinc absorption is also affected by excess calcium (Adham et al., 1980), copper (Prasad, 1993) or cadmium (Ahokas et al., 1980). Through nanotechnology, the particle size could be reduced and their quantum properties could be changed and hence converting zinc to nano form improves its bio availability (Li et al., 2016;Swain et al., 2019;Kumar et al., 2020). Objective of this study was to assess the zinc status in Malabari kids fed on complete ration having different energy levels on supplemention with nano zinc. The aim was also to evaluate the effect of prolong supplementation of nano zinc on mineral status, blood biochemistry and serum enzyme levels in growing Malabari kids.

Experimental animals
Fifteen Malabari weaned kids of 2-3 months of age were procured from University Goat and Sheep Farm, Mannuthy. They were dewormed and maintained at standard condition in an experimental shed for a period of 90 days. The kids were randomly allotted into three groups of five animals each as uniformly as possible with regard to age, sex and body weight.

Experimental treatments
The kids in all three experimental groups were fed ad libitum with iso-nitrogenous complete feed with varying energy levels. The dietary composition and chemical composition of three complete ration are given in Table 1 and 2 respectively. Three experimental rations were Treatment III: -Complete feed with 14% CP 60% TDN + 20 ppm nano zinc supplementation. Blood was collected at the end of experiment period and analyzed for haemoglobulin (Cyanmethaemoglobin method), blood glucose (GOD-POD Method), serum total protein (Biurette method), serum albumin (Bromo cresol green method), serum creatinine (modified Jaffe's method) and serum enzymes like aspartate aminotransferase (AST), serum alanine aminotransferase (ALT) (Modified IFCC method) and creatinine by using test kits.
Serum calcium concentration (Modified Arsenazo III method) and serum phosphorous (Phosphomolybdate method) were estimated with semi-automatic blood analyzer. Micro minerals like zinc, copper, manganese in serum were estimated by atomic absorption spectrophotometer (Perkin Elmer pinAAcle 500).

Statistical analysis
The experimental data were analysed using one-way ANOVA (SPSS 24.0) and means were compared using Duncan's multiple range test. by adopting standard statistical procedures (Snedecor and Cochran, 1994). Vitamin AD2EK was supplemented @ 20g/ 100 kg.
T2 and T3 supplemented with 20 ppm of nano zinc oxide.

Haemoglobin
Haemoglobin values of kids at the end of the experiment were 9.20±0.49, 8.89±0.15 and 9.15±0.19 g/dL respectively and is shown in Table 3. The values were in normal physiological ranges (Kaneko et al., 2008). Similar observations were also reported by Sethy et al. (2016) and Swain et al. (2019) who observed no significant effect on haemoglobin levels with zinc supplementation either inorganic or organic or nano form. However, Sobhanirad et al. (2014) reported supplementation of zinc had significantly increased the haemoglobin concentration in Baluchi goats. Chavan et al. (2016) and Ulutas et al.,(2020) suggested that, increased haemoglobin levels with zinc supplementation could be due to its role in erythropoiesis along with iron, folate and cyanocobalamin.

Blood Glucose
The blood glucose levels of kids estimated at end of experiment were 59.69±2.95, 56.20±1.81 and 57.35±2.65 mg/dl for the three treatments T1, T2 and T3 respectively shown in Table 3. The values obtained were in accordance with Kaneko et al., (2008). Comparable values of serum glucose levels in the experimental animals could be due to insulin-like activity of zinc which helps in regulating serum glucose levels (Attia et al., 2015) .The values were in concurrence with findings of Anil et al., (2020) who observed no difference in serum glucose levels in calves supplemented with inorganic or nano zinc (5 or 10 ppm respectively). The improved blood glucose concentration were also reported in Nubian goat (Elamin et al, 2013), Black Bengal goats (Sethy et al., 2016) and Osmanabadi goat (Govardhan, 2019) suggestive of its important role in carbohydrate metabolism.
324 Assessment of nano zinc supplementation in Malabari kids _______________________________________

Serum biochemical profile
The serum albumin, globulin and albumin to globulin ratio were found to be normal among the treatment groups and was in concurrence with values reported by Jasmine et al. (2017) in growing Malabari kids and Singh et al. (2018) in Jalauni kids. Serum enzymes, Alanine transaminase (ALT) levels were 18.40±0.95, 19.01±0.54 and 17.65±0.47 and Aspartate aminotransferase (AST) levels were 79.30±1.54, 82.14±3.11 and 75.98±1.59 in T1, T2 and T3, respectively and creatinine levels were 0.45±0.07, 0.61±0.03 and 0.51±0.08 in T1, T2 and T3, respectively. The ALT, AST and creatinine levels where in normal range and did not differ significantly (p>0.05) among treatment groups showing that long term supplementation of nano zinc had no significant effect on serum biochemical parameters of Malabari kids. Similar reports were also observed by Dhruw (2017), Swain et al. (2019) and Govardhan (2019) showing the supplemention of nano zinc at various levels in goats had no significance effect on serum enzymes such as serum ALT, AST and creatinine levels. In contrary, supplementation of nano zinc had increased AST levels in rats (Jung et al., 2010;Sharma et al., 2012), rabbits (Ismail andEl-Araby, 2017) and lambs (Najafzadeh et al., 2013) and lowered AST in Mehraban lambs (Alimohamady et al., 2018). Studies also showed a higher ALT, ALP and creatinine levels in rats (Jung et al., 2010;Sharma et al., 2012), and in lamb (Najafzadeh et al., 2013) with higher level of nano zinc supplementation.

Serum mineral profile
Serum calcium, phosphorous, copper and manganese were found to be similar in all groups and is shown in Table 4 and Fig. 1 and  Fig 2. Serum zinc levels in T1, T2 and T3 were 0.826±0.07, 1.651±0.23 and 1.736±0.31 ppm respectively found to be significantly (p<0.05) higher in nano zinc supplemented groups (T2 and T3) over T1.

Fig. 2.
Concentration of serum zinc, copper and manganese at end of kids maintained on three experimental ration, ppm ranged from 0.69 to 0.78 ppm and 1.44 to 1.48 ppm respectively and values were found within normal physiological range in growing Malabari kids. Lower copper levels in serum (Jia et al., 2009) and plasma (Salama et al., 2013) in goats were reported on zinc supplementation suggestive of their interaction.
In present study, serum zinc levels were higher (p<0.05) in nano zinc supplemented groups (T2 and T3) compared to T1 could be due to higher zinc retention and increased serum zinc levels did not affect the serum mineral status of calcium, phosphorous, copper and manganese.Similarly, supplementation of inorganic zinc to buffalo (Jadhav et al., 2005), organic zinc to calves (Mandal et al., 2007), Muzzafanagari lambs (Garg et al., 2008 and Mehraban lambs (Aliarabi et al., 2015) and nano zinc supplementation in non-descriptive 326 Assessment of nano zinc supplementation in Malabari kids _______________________________________ goats (Dhruw, 2017) and Osmanabadi goats (Govardhan, 2019) were observed to have improved serum zinc levels without affecting serum mineral status of calcium, phosphorous, copper and iron levels.In contrast to our findings, serum calcium levels were increased significantly in growing calves (Khan, 1978;Bedi, 1979) and decreased in buffaloes (Daghash and Mousa, 1999) and West African goats (Phiri et al., 2009). Serum zinc levels were not improved by supplementation of inorganic zinc in Markhoz goat kids (Zaboli et al., 2013) and Nubian goats (Elamin et al., 2013).

Conclusion
The results of the study on kids maintained on complete feed with lower energy levels (65 or 60 TDN) with supplementation of nano zinc at 20 ppm were at par with growing kids fed on higher energy level (70 TDN) without any change in haematological and serum biochemical profile. Serum glucose levels were in normal range and comparable between treatment groups fed on different energy levels.
The minerals status of all groups were similar except serum zinc levels were higher in nano zinc supplemented groups showing higher zinc bio availability. Thus, it can be concluded that nano zinc oxide at 20 ppm could be incorporated safely in complete feed having low energy (60 % TDN) to growing Malabari kids.