Effect of Different Levels of Egg Shell Powder on Gas Production, Fermentation Parameters and Digestibility of Aflatoxin B1 Contaminated Diet in vitro

Document Type : Ruminant Nutrition

Authors

• Rashid Safari ¹
• Zabihollah Nemati ²
• Mohammadreza Sheikhlou ³

¹ Department of Animal Science, Ahar Faculty of Agriculture and Natural Resources, University of Tabriz, Tabriz, Iran

² Animal Science Department,Ahar Faculty of Agriculture and Natural Resources, University of Tabriz, Tabriz, Iran.

³ Animal Science Department, Ahar Faculty of Agriculture and Natural Resources, University of Tabriz, Tabriz, Iran.

10.22067/ijasr.2021.38309.0

Abstract

Introduction Mycotoxins are secondary metabolites of fungi that are produced under stress condition. Aflatoxin is one of several extremely toxic, mutagenic and carcinogenic compounds produced by Aspergillus Flavus and Aspergillus Parasiticus. Numerous agricultural commodities such as Forages, cereal grains, oilseeds, and cotton seeds are potential sources of aflatoxins in ruminant diets. Many studies indicate that ruminants, like other animals, are affected by aflatoxins. Aflatoxin B₁ reduces ruminal digestion, animal production, and in high doses causes liver damage and death in ruminants. Several chemical, biological, and physical strategies developed in order to, detoxification of aflatoxins or minimizing the production of aflatoxins and inhibiting the absorption of them in the gastrointestinal tract. Recently, many researchers are focused on aflatoxin adsorbents to reduce the bioavailability of aflatoxins in the diet. Eggshell has a porous structure and on the other hand has significant amounts of pure calcium carbonate, which has the ability to absorb toxins. Due to limit information on the ability of egg shell powder to absorb aflatoxin, the present study was designed to investigate the effect of adding egg shell powder as toxin binder in diets containing aflatoxin B₁ on fermentation parameters and ruminal digestibility and gas production in vitro.
 Materials and Methods To produce the aflatoxin required for the experiment, a standard strain of Aspergillus Parasiticus NRLL 2999 used and cultured on potato dextrose agar. In other to obtain proper amount of Aflatoxin, 2 ml of spore suspension with a concentration of 6.5× 10⁶ grown fungi was prepared and added to a flask containing sterile culture medium. After 5 days, the culture medium was dried in an oven. Culture medium contained of 250.9 mg/kg aflatoxin B₁. This experiment was performed in a completely randomized design with 5 treatments and 4 replications in each treatment. Experimental treatments consist of , control with methanol, control without methanol, Aflatoxin (800 ng/mg of rumen fluid), Aflatoxin+ level 1 egg shell powder (7 mg per 200 mg of diet), Aflatoxin +level 2 egg shell powder(75 mg per 200 mg of diet), Aflatoxin +level 3 egg shell powder(150 mg per 200 mg of diet). Rumen fluid was collected before the morning feed from three fistulated Moghani male sheep with 46 ± 3 kg live weight. Sheep fed with basal diet used in this experiment at a concentration of 50:50 forage to concentrate for 15 days before rumen fluid collection. In vitro gas production was measured in 4 replicate with 200 mg DM for each. The bottles were filled with 30 ml of incubation medium that consisted of 10 ml of rumen fluid plus 20 ml of buffer solution and placed in a water bath at 39 °C. Gas production was recorded at 2, 4, 8, 16, 24, 48, 72 and 96 h. Gas values corrected for blank incubation. The gas production and rate of gas production measured through 96 h incubation. A procedure similar to gas production with 4 replicate for each treatment was used for rumen batch culture system to measure NH3-N and in vitro digestibility after 24 h incubation. Contents of each glass bottle were filtered through three layers of cheesecloth and rumen fluid used to determination of NH3-N using the distillation method. Finally, all remaining contents oven dried at 60 °C for 48 h and analyzed for IVDMD and IVOMD. Metabolizable energy (ME), net energy for lactation (NE_L), short chain fatty acids (SCFA), partitioning factor (PF), Microbial mass and Efficiency of microbial synthesis calculate throughout 24h incubation.
 Results and Discussion Results indicate a significant decrease in GP in aflatoxin treatment compared to treatments without aflatoxin, so that the amount of gas production decreased from 295.05 and 294.38 in control with methanol and control without methanol to 228.48 ml/g of DM. This change in GP was associated with significant reduction in IVDMD and IVOMD, ME, NE_L, SCFA, PF, microbial mass and Efficiency of microbial synthesis. Addition of different levels of egg shell powder as aflatoxin binder improved fermentation conditions which was significant in level 2 treatment compare to aflatoxin treatment. There was no significant difference in GP, IVDMD and IVOMD, ME, NE_L, SCFA, PF, microbial mass and efficiency of microbial synthesis between control treatments and level 2 egg shell powder as toxin binder.
 Conclusion Considering all the results of experiment, egg shell could be considered as an adsorbent of aflatoxin in ruminal conditions. Egg shell powder suggested as toxin binder to reduce the negative effects of aflatoxin B₁ on microbial activity and degradability in ruminal conditions.

Keywords

• Aflatoxin
• Digestibility
• Egg Shell
• Gas production
• Partitioning Factor

20.1001.1.20083106.1400.13.1.8.2

---

References

1. Abdulrazak, S. A., J. Nyangaga., and T. Fujihara. 2001. Relative palatability to sheep of some browse species, their in sacco degradability and in vitro gas production characteristics. Asian-Australasian Journal of Animal Science, 14(11) 1580–1584.

2. Akhtar, S., M. A. Shahzad., S. H. Yoo., and A. Ismail. 2017. Determination of aflatoxin M1 and heavy metals in infant formula milk brands available in Pakistani markets. Korean Journal for Food Science of Animal Resources, 37(1) 79–86.

3. Assadzadeh, S., A. Tahmasbi., A. A. Naserian., and R. Valizadeh. 2018. The effect of organic and inorganic aflatoxin B₁ absorbents on in vitro digestibility and rumen fermentation characteristics. Iranian Journal of Animal Science Research, 9(4) 413–423.

4. Cassel, E., B. Campbell., M. Draper., and B. Epperson. 2001. Aflatoxins hazards in grain (Aflatoxicosis and Livestock).South Dakota State University Extention.

5. Elliott, C. T., L. Connolly., and O. Kolawole. 2020. Potential adverse effects on animal health and performance caused by the addition of mineral adsorbents to feeds to reduce mycotoxin exposure. Mycotoxin Research, 36(1) 115–126.

6. Gallo, A., G. Giuberti., J. C. Frisvad., T. Bertuzzi., and K. F. Nielsen. 2015. Review on mycotoxin issues in ruminants: Occurrence in forages, effects of mycotoxin ingestion on health status and animal performance and practical strategies to counteract their negative effects. Toxins, 7(8) 3057–3111.

7. Getachew, G., H. P. S. Makkar., and K. Becker. 2002. Tropical browses: Contents of phenolic compounds, in vitro gas production and stoichiometric relationship between short chain fatty acid and in vitro gas production. Journal of Agricultural Science, 139(3) 341–352.

8. Gurung, N. K., D. L. Rankins., and R. A. Shelby. 1999. In vitro ruminal disappearance of fumonisin B₁ and its effects on in vitro dry matter disappearance. Veterinary and Human Toxicology, 41(4) 196–199.

9. Hall, M. B., T. D. Nennich., P. H. Doane., and G. E. Brink. 2015. Total volatile fatty acid concentrations are unreliable estimators of treatment effects on ruminal fermentation in vivo. Journal of Dairy Science, 98(6) 3988–3999.

10. Helferich, W. G., R. L. Baldwin., and D. P. H. Hsieh. 1986. Aflatoxin B₁ metabolism in lactating goats and rats. Journal of Animal Science, 62(3) 697–705.

11. Jeong, J., J. Lee., Y. Simizu., and H. Tazaki. 2010. Effects of the Fusarium mycotoxin deoxynivalenol on in vitro rumen fermentation. Animal Feed Science and Technology, 162(3–4) 144–148.

12. Jiang, Y. H., H. J. Yang., and P. Lund. 2012. Effect of aflatoxin B₁ on in vitro ruminal fermentation of rations high in alfalfa hay or ryegrass hay. Animal Feed Science and Technology, 175(1–2) 85–89.

13. Jiang, Y., I. M. Ogunade., D. H. Kim., and X. Li. 2018. Effect of adding clay with or without a Saccharomyces cerevisiae fermentation product on the health and performance of lactating dairy cows challenged with dietary aflatoxin B₁. Journal of Dairy Science, 101(4) 3008–3020.

14. Jouany, J., and A. Yiannikouris. 2009. Risk assessment of mycotoxins in ruminants and ruminant products. Options Mediterraneennes, 85(1) 205–224.

15. Kotinagu, K., T. Mohanamba., and L. Rathna Kumari. 2015. Assessment of aflatoxin B₁ in livestock feed and feed ingredients by high-performance thin layer chromatography. Veterinary World, 8(12) 1396–1399.

16. Makkar, H. P. S. 2004. Recent advances in the in vitro gas method for evaluation of nutritional quality of feed resoures. In Assessing quality and safety of animal feeds. 55–88.

17. Mauricio, R. M., F. L. Mould., M. S. Dhanoa., E. Owen., K. S. Channa., and M. K. Theodorou. 1999. A semi-automated in vitro gas production technique for ruminant feedstuff evaluation. Animal Feed Science and Technology, 79(1) 321–330.

18. Menke, K. H., L. Raab., A. Salewski., H. Steingass., D. Fritz., and W. Schneider. 1979. The estimation of the digestibility and metabolizable energy content of ruminant feedingstuffs from the gas production when they are incubated with rumen liquor in vitro. The Journal of Agricultural Science, 93(1) 217–222.

19. Menke, K., and H. Steingass. 1988. Estimation of the energetic feed value obtained from chemical analysis and in vitro gas production using rumen fluid. animal research, 287–55.

20. Mojtahedi, M. 2013. Effect of aflatoxin B₁ on in vitro rumen microbial fermentation responses using batch culture. Annual review & Research in Biology, 3(4) 686–693.

21. Nemati, Z., H. Janmohammadi., A. Taghizadeh., H. Maleki Nejad., and Gh. Mogaddam. 2015. Effect of Bentonite as a natural adsorbent to ameliorate the adverse effects of aflatoxin B₁ on performance and immune systems in broiler chicks. Animal Production Research, 4(3) 67–78.

22. Nemati, Z., A. Karimi., and M. Besharati. 2015. Impact of aflatoxins contaminating feed and yeast cell wall supplementation on immune system in broiler chickens. International Conference on Innovations in Chemical & Agricultural Engineering, 14–17.

23. Nemati, Z., and R. Safari. 2020. Effect of activated charcoal on the fermentation parameters and degradability of contaminated diet with aflatoxin B₁ by microorganisms isolated from rumen in vitro condition. Research On Animal Production, 11(29) 27–29.

24. Nemati, Z., R. Safari., N. Kamrani., M. Sheikhlou., and M. Besharati. 2020. Effect of Aflatoxin B₁ on In Vitro Digestibility and Ruminal Fermentation of Sheep Diet. Research On Animal Production, 11(28) 75–83.

25. Orskov, E. R., and I. Mcdonald. 1979. The estimation of protein degradability in the rumen from incubation measurements weighted according to rate of passage. The Journal of Agricultural Science, 92(2) 499–503.

26. Park, H. J., J. Seong Wook., Y. Jae Kyu., K. Boo Gil., and L. Seung Mok. 2007. Removal of heavy metals using waste eggshell. Journal of Environmental Sciences, 19(12) 1436–1441.

27. Reynal, S. M., I. R. Ipharraguerre., M. Liñeiro., and A. F. Brito. 2007. Omasal flow of soluble proteins, peptides, and free amino acids in dairy cows fed diets supplemented with proteins of varying ruminai degradabilities. Journal of Dairy Science, 90(4) 1887–1903.

28. Safari, R., Z. Nemati., N. Kamrani., and A. Karimi. 2020. Effect of diatoms on the fermentation parameters and digestibility of contaminated diet with aflatoxin B₁ by microorganisms isolated from rumen in vitro condition. Journal of Veterinary Microbiology, 16(1) 50–60.

29. SAS. 2002. SAS User’s Guide. 9.1 Edition. Statistics SAS Institute, Cary, NC, U.

30. Souza, N. K. P., E. Detmann., S. C. Valadares Filho., V. A. C. Costa., D. S. Pina., D. I. Gomes., A. C. Queiroz., and H. C. Mantovani. 2013. Accuracy of the estimates of ammonia concentration in rumen fluid using different analytical methods. Arquivo Brasileiro de Medicina Veterinaria e Zootecnia, 65(6) 1752–1758.

31. Tapia, M. O., M. D. Stern., A. L. Soraci., and R. Meronuck. 2005. Patulin-producing molds in corn silage and high moisture corn and effects of patulin on fermentation by ruminal microbes in continuous culture. Animal Feed Science and Technology, 119(3–4) 247–258.

32. Udomkun, P., A. N. Wiredu., M. Nagle., J. Müller., B. Vanlauwe., and R. Bandyopadhyay. 2017. Innovative technologies to manage aflatoxins in foods and feeds and the profitability of application – A review. Food Control, 76 127–138.

33. Varadyova, Z., M. Baran., and I. Zelenak. 2005. Comparison of two in vitro fermentation gas production methods using both rumen fluid and faecal inoculum from sheep. Animal Feed Science and Technology, 123 81–94.

34. Vercoe, P. E., H. P. S. Makkar., and A. C. Schlink. 2010. In vitro screening of plant resources for extra-nutritional attributes in ruminants: Nuclear and related methodologies. Springer Netherlands.

35. Westlake, K., R. I. Mackie., and M. F. Dutton. 1989. In vitro metabolism of mycotoxins by bacterial, protozoal and ovine ruminal fluid preparations. Asian-Australasian Journal of Animal Sciences, 25 169–178.