magazinelogo

International Journal of Food Science and Agriculture

ISSN Online: 2578-3475 ISSN Print: 2578-3467 CODEN: IJFSJ3
Frequency: quarterly Email: ijfsa@hillpublisher.com
Total View: 3743626 Downloads: 618891 Citations: 1084 (From Dimensions)
Google-based citation data
  • citations

    2157
  • h-index

    20
  • i10-index

    66
ArticleOpen Access http://dx.doi.org/10.26855/ijfsa.2026.09.001

The Effect of Milk Replacers Containing Fermented Plant Protein and High Starch Content on the Growth Performance of Holstein Bull Calves

E. E. Kriel1,*, J. H. C. van Zyl2, G. V. Kriel3

1Department of Animal Science, University of the Free State, Bloemfontein 9301, Free State, South Africa.

2Department of Animal Science, Stellenbosch University, Stellenbosch 7600, South Africa.

3Independent researcher, Oudtshoorn 6625, South Africa.

*Corresponding author: E. E. Kriel

Published: July 27, 2026

Abstract

This study investigated the effects of fermented protein and increased starch levels in milk replacers on calf growth performance. Fermented protein was developed as an alternative raw material to enhance the amino acid composition of plant-based substrates in milk replacers, with the aim of improving growth outcomes. Two milk replacers were used, one containing fermented protein and the other a commercial milk replacer using a conventional protein source. To assess the impact of increased starch levels, two additional milk replacers were used, one with fermented protein and one without, both containing 20% pre-cooked maize. A total of 32 Holstein bull calves were randomly assigned to four treatments (n = 8) in a 63-day trial, during which milk replacers were fed. Milk replacer intake was controlled, while starter meal intake was offered ad libitum, and all intakes and refusals were recorded. Growth performance parameters, including weight gain, dry matter intake, average daily gain, dry matter intake as a percentage of body weight, and feed conversion ratio, were determined and analysed using factorial ANOVA. Except for controlled milk replacer intake, fermented protein-containing milk replacers generally resulted in poorer performance compared to standard protein milk replacers. This was attributed to higher levels of trypsin inhibitors in the fermented protein, which negatively impacted performance. The high starch milk replacers promoted greater starter meal intake. Based on these results, it is recommended that milk replacers with higher starch content can be beneficial to stimulate starter meal intake, while the use of fermented protein as a raw material should only be considered after the fermentation process is better optimized to reduce trypsin inhibitor levels to below 4 mg/g protein.

Keyword

Diarrhoea; enzymatic digestion; pre-ruminants; starter meal; trypsin inhibitor

References

[1] Porter JWG. Digestion in the pre-ruminant animal. Proc Nutr Soc. 1969;28(1):115-21.

[2] Ørskov ER. Reflex closure of the oesophageal groove and its potential application in ruminant nutrition. S Afr J Anim Sci. 1972; 2(2):169-76.

[3] Amado L, Berends H, Leal LN, et al. Effect of energy source in calf milk replacer on performance, digestibility, and gut permeability in rearing calves. J Dairy Sci.2019;102(5):3994-4001.

[4] Johnson KF, Vinod Nair ABR, Cand A, Wathes DC. Comparison of the effects of high and low milk-replacer feeding regimens on health and growth of cross bred dairy heifers. Anim Prod Sci. 2019;59(8):1648-59.

[5] Brown EG, VandeHaar MJ, Daniels KM, et al. Effect of increasing energy and protein intake on body growth and carcass composition of heifer calves. J Dairy Sci. 2005;88(2):585-94.

[6] Bach A, Terré M, Pinto A. Performance and health responses of dairy calves offered different milk replacer allowances. J Dairy Sci. 2013;96(12):7790-7.

[7] Hu W, Hill TM, Dennis TS, et al. Effects of milk replacer feeding rates on growth performance of Holstein dairy calves to 4 months of age, evaluated via a meta-analytical approach. J Dairy Sci. 2020;103(3):2217-32.

[8] Kitadai N, Maruyama S. Origins of building blocks of life: a review. Geosci Front. 2018;9(4):1117-53.

[9] Jacob F, Monod J. Genetic regulatory mechanisms in the synthesis of proteins. J Mol Biol. 1961;3:318-56.

[10] Numata K. How to define and study structural proteins as biopolymer materials. Polym J.2020;52(8):1043-56.

[11] Morris R, Black KA, Stollar EJ. Uncovering protein function: from classification to complexes. Essays Biochem.2022;66(3):255-85.

[12] Katz DL, Doughty KN, Geagan K, et al. Perspective: the public health case for modernizing the definition of protein quality. Adv Nutr. 2019;10(5):755-64.

[13] Schubert DC, Chuppava B, Hoffmans S, et al. Impacts of reducing protein content in milk replacer on growth performance and health of young calves. Animals (Basel). 2022;12(14):1756.

[14] Huang K, Tu Y, Si B, et al. Effects of protein sources for milk replacers on growth performance and serum biochemical indexes of suckling calves. Anim Nutr. 2015;1(4):349-55.

[15] Andrén A. Rennets and coagulants. In: Fuquay CC, Fox PF, McSweeney PLH, editors. Encyclopedia of Dairy Sciences. 2nd ed. San Diego: Academic Press/Elsevier; 2011. p. 566-73.

[16] Miyazaki T, Okada K, Yamashita T, Miyazaki M. Temporal changes of abomasal contents and volumes in calves fed milk diluted with oral rehydration salt solution. J Vet Med Sci. 2019;81(2):256-62.

[17] Cruywagen CW, Brisson GJ, Meissner HH. Casein curd-forming ability and abomasal retention of milk replacer components in young calves. J Dairy Sci. 1990;73(6):1578-85.

[18] Petit HV, Ivan M, Brisson GJ. Duodenal flow of digesta in preruminant calves fed clotting or nonclotting milk replacer. J Dairy Sci. 1987;70(12):2570-6.

[19] Longenbach JI, Heinrichs AJ. A review of the importance and physiological role of curd formation in the abomasum of young calves. Anim Feed Sci Technol. 1998;73(1-2):85-97.

[20] Rivera Del Rio A, Keppler JK, Boom RM, Janssen AEM. Protein acidification and hydrolysis by pepsin ensure efficient trypsin-catalyzed hydrolysis. Food Funct.2021;12(10):4570-81.

[21] Kurz A, Seifert J. Factors influencing proteolysis and protein utilization in the intestine of pigs: a review. Animals (Basel). 2021;11(12):3551.

[22] Van der Velden VHJ, Hulsmann AR. Peptidases: structure, function and modulation of peptide-mediated effects in the human lung. Clin Exp Allergy. 1999;29(4):445-56.

[23] López-Otín C, Bond JS. Proteases: multifunctional enzymes in life and disease. J Biol Chem. 2008;283(45):30433-7.

[24] National Research Council (US). Nutrient requirements of dairy cattle. 8th rev ed. Washington (DC): National Academies Press; 2021.

[25] Beever DE. Meeting the protein requirements of ruminant livestock. S Afr J Anim Sci. 1996;26(1):20-6.

[26] Ertl P, Knaus W, Zollitsch W. An approach to including protein quality when assessing the net contribution of livestock to human food supply. Animal. 2016;10(11):1883-9.

[27] Wolfe RR, Rutherfurd SM, Kim IY, Moughan PJ. Protein quality as determined by the digestible indispensable amino acid score: evaluation of factors underlying the calculation. Nutr Rev. 2016;74(9):584-99.

[28] Jahan-Mihan A, Luhovyy BL, El Khoury D, Anderson GH. Dietary proteins as determinants of metabolic and physiologic functions of the gastrointestinal tract. Nutrients. 2011;3(5):574-603.

[29] Akande KE, Fabiyi EF. Effect of processing methods on some antinutritional factors in legume seeds for poultry feeding. Int J Poult Sci. 2010;9(10):996-1001.

[30] El-Shemy H, Abdel-Rahim E, Shaban O, et al. Comparison of nutritional and antinutritional factors in soybean and fababean seeds with or without cortex. Soil Sci Plant Nutr. 2000;46(2):515-24.

[31] Sánchez-García J, Muñoz-Pina S, García-Hernández J, et al. Protein digestibility and ACE inhibitory activity of fermented flours in older adults and standard gastrointestinal simulation. Food Res Int. 2024;180:114080.

[32] El-Hag ME, El-Tinay AH, Yousif NE. Effect of fermentation and dehulling on starch, total polyphenols, phytic acid content and in vitro protein digestibility of pearl millet. Food Chem.2002;77(2):193-6.

[33] Singh Alka, Yadav N, Sharma S. Effect of fermentation on physicochemical properties & in vitro starch and protein digestibility of selected cereals. Int J Agric Food Sci. 2012;2(3):66-70.

[34] Ali MAM, El-Tinay AH, Abdalla AH. Effect of fermentation on the in vitro protein digestibility of pearl millet. Food Chem. 2003;80(1):51-4.

[35] Pranoto Y, Anggrahini S, Efendi Z. Effect of natural and Lactobacillus plantarum fermentation on in vitro protein and starch digestibilities of sorghum flours. Food Biosci.2013;2:46-52.

[36] Hamad AM, Fields ML. Evaluation of the protein quality and available lysine of germinated and fermented cereal. J Food Sci. 1979;44(2):456-9.

[37] Wakil SM, Onilude AA. Microbiological and chemical changes during production of malted and fermented cereal-legume weaning foods. Adv Food Sci. 2009;31(2):139-45.

[38] Adeyemo SM, Onilude AA. Enzymatic reduction of anti-nutritional factors in fermenting soybeans by Lactobacillus plantarum isolates from fermenting cereals. Niger Food J. 2013;31(2):84-90.

[39] Shekib LA. Nutritional improvement of lentils, chick pea, rice and wheat by natural fermentation. Plant Foods Hum Nutr. 1994;46(3):201-5.

[40] Reyes-Moreno C, Cuevas-Rodríguez EO, Milán-Carrillo J, et al. Solid state fermentation process for producing chickpea (Cicer arietinum L) tempeh flour. Physicochemical and nutritional characteristics of the product. J Sci Food Agric. 2004;84(3):271-8.

[41] Chandra-Hioe MV, Wong CHM, Arcot J. The potential use of fermented chickpea and faba bean flour as food ingredients. Plant Foods Hum Nutr. 2016;71(1):90-5.

[42] Lopez Y, Gordon DT, Fields ML. Release of phosphorous from phytate by natural lactic fermentation. J Food Sci. 1983;48(3): 935-9.

[43] Hemalatha S, Platel K, Srinivasan K. Influence of germination and fermentation on bioaccessibility of zinc and iron from food grains. Eur J Clin Nutr. 2007;61(3):342-8.

[44] Emambux MN, Taylor JN. Sorghum kafirin interaction with various phenolic compounds. J Sci Food Agric. 2003;83(4):402-7.

[45] Nkhata SG, Ayua E, Kamau EH, Shingiro JB. Fermentation and germination improve nutritional value of cereals and legumes through activation of endogenous enzymes. Food Sci Nutr. 2018;6(8):2446-58.

[46] Liang J, Han BZ, Nout MJR, Hamer RJ. Effects of soaking, germination and fermentation on phytic acid, total and in vitro soluble zinc in brown rice. Food Chem. 2008;110(3):821-8.

[47] Çabuk B, Nosworthy MG, Stone AK, et al. Effect of fermentation on the protein digestibility and levels of non-nutritive compounds of pea protein concentrate. Food Technol Biotechnol. 2018;56(2):257-64.

[48] Leenhardt F, Levrat-Verny M, Chanliaud E, Rémésy C. Moderate decrease of pH by sourdough fermentation is sufficient to reduce phytate content of whole wheat flour through endogenous phytase activity. J Agric Food Chem. 2005;53(1):98-102.

[49] Vargas-Rodriguez CF, Engstrom M, Azem E, Bradford BJ. Effects of dietary amylase and sucrose on productivity of cows fed low-starch diets. J Dairy Sci. 2014;97(7):4464-70.

[50] Hua D, Hendriks WH, Xiong B, Pellikaan WF. Starch and cellulose degradation in the rumen and applications of metagenomics on ruminal microorganisms. Animals (Basel). 2022;12(21):3020.

[51] Gambelli L. Milk and its sugar-lactose: a picture of evaluation methodologies. Beverages. 2017;3(3):35.

[52] Dollar AM, Porter JWG. Utilization of carbohydrates by the young calf. Nature. 1957;179(4573):1299-300.

[53] Blaxter K. Book review: Animal nutrition by A.A. Bondi. Livest Prod Sci. 1988;18(3-4):315-6.

[54] Roy JHB. The calf. Vol 1: Management of health. 5th ed. London: Butterworths; 1990.

[55] Juers DH, Matthews BW, Huber RE. LacZ β-galactosidase: structure and function of an enzyme of historical and molecular biological importance. Protein Sci. 2012;21(12):1792-807.

[56] Coombe NB, Siddons RC. Carbohydrases of the bovine small intestine. Br J Nutr. 1973;30(2):269-76.

[57] Toofanian F, Hill FWG, Kidder DE. The mucosal disaccharidases in the small intestine of the calf. Ann Rech Vet. 1973;4(1):57-69.

[58] Le Huerou-Luron I. Production and gene expression of brush border disaccharidases and peptidases during development in pigs and calves. In: Tolkamp BJ, Kyriazakis I, editors. Biology of Growing Animals. Vol 1. Amsterdam: Elsevier; 2006. p. 491-513.

[59] Walker DM. The development of the digestive system of the young animal III. Carbohydrase enzyme development in the young lamb. J Agric Sci. 1959;53(3):374-80.

[60] Estévez CA, Isasi JR, Larrañeta E, Vélaz I. Release of β-galactosidase from poloxamine/α-cyclodextrin hydrogels. Beilstein J Org Chem. 2014;10:3127-35.

[61] Salema-Oom M, Valadão Pinto V, Gonçalves P, Spencer-Martins I. Maltotriose utilization by industrial Saccharomycesstrains: characterization of a new member of the α-glucoside transporter family. Appl Environ Microbiol. 2005;71(9):5044-9.

[62] Damager I, Engelsen SB, Blennow A, Møller BL, Motawia MS. First principles insight into the alpha-glucan structures of starch: their synthesis, conformation, and hydration. Chem Rev. 2010;110(4):2049-80.

[63] Le Huerou I, Guilloteau P, Wicker C, et al. Activity distribution of seven digestive enzymes along small intestine in calves during development and weaning. Dig Dis Sci. 1992;37(1):40-6.

[64] Parada J, Aguilera JM. Review: starch matrices and the glycemic response. Food Sci Technol Int. 2011;17(3):187-204.

[65] Serena A, Jørgensen H, Knudsen KEB. Digestion of carbohydrates and utilization of energy in sows fed diets with contrasting levels and physicochemical properties of dietary fiber. J Anim Sci. 2008;86(9):2208-16.

[66] Velu JG, Kendall KA, Gardner KE. Utilization of various sugars by the young dairy calf. J Dairy Sci. 1960;43(4):546-52.

[67] Ballard FJ, Oliver IT. Carbohydrate metabolism in liver from foetal and neonatal sheep. Biochem J. 1965;95(1):191-200.

[68] Trotta RJ, Harmon DL, Matthews JC, Swanson KC. Nutritional and physiological constraints contributing to limitations in small intestinal starch digestion and glucose absorption in ruminants. Ruminants. 2022;2(1):1-26.

[69] Krehbiel CR, Britton RA, Harmon DL, et al. Effects of varying levels of duodenal or midjejunal glucose and 2-deoxyglucose infusion on small intestinal disappearance and net portal glucose flux in steers. J Anim Sci. 1996;74(3):693-700.

[70] Cordy PA. A descriptive study on the production of microbial derived protein [master’s thesis]. Stellenbosch (South Africa): University of Stellenbosch; 2024.

[71] Miyashige T, Yahata S. Development of intestinal disaccharidase activities in nursing calves [Japanese]. Jpn J Zootech Sci. 1980;51(1):58-68.

[72] Harmon DL. Nutritional regulation of postruminal digestive enzymes in ruminants. J Dairy Sci. 1993;76(7):2102-11.

[73] AOAC International. Official methods of analysis. 17th ed. Arlington (VA): AOAC International; 2002. Method 990.03.

[74] Moneeb AHM, Hammam ARA, Ahmed AKA, et al. Effect of fat extraction methods on the fatty acids composition of bovine milk using gas chromatography. Food Sci Nutr. 2021;9(6):2936-42.

[75] Evers JM, Wightman LM, Crawford RA, et al. A precise method to measure the total fat content of spreadable fats. Int Dairy J. 2001;10(12):815-27.

[76] Fiorentini G, Carvalho IP, Messana JD, et al. Effect of lipid sources with different fatty acid profiles on intake, nutrient digestion and ruminal fermentation of feedlot Nellore steers. Asian-Australas J Anim Sci. 2015;28(11):1583-91.

[77] Bionaz M, Vargas-Bello-Pérez E, Busato S. Advances in fatty acids nutrition in dairy cows: from gut to cells and effects on performance. J Anim Sci Biotechnol. 2020;11:110.

[78] Nitsan Z, Nir I. Accentuated response to soybean inhibitors by meal-feeding in various species. In: Friedman M, editor. Nutritional and Toxicological Significance of Enzyme Inhibitors in Foods. Adv Exp Med Biol. Vol 199. Boston: Springer; 1986. p. 177-81.

[79] Uskenov R, Yengsebek T, Kurzhykaev Z, et al. The relationship between dry matter intake and the average daily gain. BIO Web Conf.2024;85:01006.

[80] Avilés-Gaxiola S, Chuck-Hernández C, Serna Saldívar SO. Inactivation methods of trypsin inhibitor in legumes: a review. J Food Sci. 2018;83(1):17-29.

[81] Chavan JK, Kadam SS. Nutritional improvement of cereals by fermentation. Crit Rev Food Sci Nutr. 1989;28(5):349-400.

[82] Duodu KG, Taylor JRN, Belton PS, Hamaker BR. Factors affecting sorghum protein digestibility. J Cereal Sci.2003;38(2):117-31.

[83] Gao Y, Wang C, Zhu Q, Qian G. Optimization of solid-state fermentation with Lactobacillus brevisand Aspergillus oryzaefor trypsin inhibitor degradation in soybean meal. J Integr Agric. 2013;12(5):869-76.

[84] Emkani M, Oliete B, Saurel R. Effect of lactic acid fermentation on legume protein properties, a review. Fermentation. 2022;8(6):244.

[85] Lallès JP, Tukur HM, Toullec R, Müller BG. Analytical criteria for predicting apparent digestibility of soybean protein in pre-ruminant calves. J Dairy Sci. 1996;79(3):475-82.

[86] Nielsen NC. The structure and complexity of the 11S polypeptides in soybeans. J Am Oil Chem Soc. 1985;62(12):1680-6.

[87] Iwabuchi S, Yamauchi F. Determination of glycinin and β-conglycinin in soybean proteins by immunological methods. J Agric Food Chem. 1987;35(2):200-5.

[88] Lallès JP, Benkredda D, Toullec R. Influence of soya antigen levels in milk replacers on the disruption of intestinal motility patterns in calves sensitive to soya. J Vet Med A Physiol Pathol Clin Med. 1995;42(7):467-78.

[89] Gemede HF, Ratta N. Antinutritional factors in plant foods: potential health benefits and adverse effects. Int J Food Sci Nutr. 2014;3(4):284-9.

[90] Nisley MJ, Spencer JD, Mendoza OF, et al. 137 Impact of dietary soybean-mediated trypsin inhibitor concentrations on nursery pig performance. J Anim Sci. 2024;102(Suppl 2):109-10.

[91] Kakade ML, Thompson RD, Engelstad WE, et al. Failure of soybean trypsin inhibitor to exert deleterious effects in calves. J Dairy Sci. 1976;59(8):1484-9.

[92] Ansia I, Drackley JK. Graduate student literature review: the past and future of soy protein in calf nutrition. J Dairy Sci. 2020;103(8):7625-38.

[93] Mir PS, Burton JH, Willkie BN. Reduction of β-conglycinin antigenicity and rate of acid-pepsin proteolysis of proteins in extruded or rumen fluid-treated soybean meal. Can J Anim Sci. 1989;69(3):727-34.

[94] Barratt ME, Strachan PJ, Porter P. Antibody mechanisms implicated in digestive disturbances following ingestion of soya protein in calves and piglets. Clin Exp Immunol. 1978;31(3):305-12.

[95] Diao Q, Zhang R, Fu T. Review of strategies to promote rumen development in calves. Animals (Basel). 2019;9(8):490.

[96] Nejad JG, Hosseindoust A, Shoae A, et al. Effects of feeding levels of starter on weaning age, performance, nutrient digestibility and health parameters in Holstein dairy calves. Asian-Australas J Anim Sci. 2013;26(6):827-30.

[97] Van Hoeij RJ, Kok A, Bruckmaier RM, et al. Relationship between metabolic status and behavior in dairy cows in week 4 of lactation. Anim. 2019;13(3):640-8.

[98] Colucci PE, Chase LE, Van Soest PJ. Feed intake, apparent diet digestibility, and rate of particulate passage in dairy cattle. J Dairy Sci. 1982;65(8):1445-56.

[99] Ganai AM, ul Haq Z, Beigh YA, Sheikh GG. Bypass nutrient technology with recent advances for enhancing animal production: a review. J Pharmacogn Phytochem. 2019;8(Spec Issue 5):269-75.

[100] Huber JT, Natrajan S, Polan CE. Varying levels of starch in calf milk replacers. J Dairy Sci. 1968;51(7):1081-4.

[101] Bernard JK, Castro JJ, Kertz AF, Murphy MR. Effect of milk replacer carbohydrate source on performance and health of dairy calves. Prof Anim Sci. 2013;29(5):463-8.

Copyright

© 2026 by the author(s).
This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution-NonCommercial-NoDerivatives (CC BY-NC-ND) license, which permits non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited and is not modified or adapted.
https://creativecommons.org/licenses/by-nc-nd/4.0/

How to cite this paper

The Effect of Milk Replacers Containing Fermented Plant Protein and High Starch Content on the Growth Performance of Holstein Bull Calves

How to cite this paper: E. E. Kriel, J. H. C. van Zyl, G. V. Kriel. (2026) The Effect of Milk Replacers Containing Fermented Plant Protein and High Starch Content on the Growth Performance of Holstein Bull CalvesInternational Journal of Food Science and Agriculture10(3), 197-207.

DOI: http://dx.doi.org/10.26855/ijfsa.2026.09.001