This study aimed to perform lactose hydrolysis using free and immobilised β-galactosidase in cow and sheep milk with different fat levels, followed by lactulose synthesis through the isomerisation of glucose to fructose using immobilised glucose isomerase. There was no significant difference (P > 0.05) in lactose hydrolysis (>95%) using the free enzyme in the different types of milk. In contrast, significant differences were observed for the hydrolysis of whole cow milk (WCM) and skimmed cow milk (SCM) with the immobilised enzyme (IE). WCM required a longer hydrolysis time (43 h). Thus, the fat content may have affected the lactose hydrolysis of bovine milk with immobilised β-galactosidase. No difference was observed for whole sheep milk and skimmed sheep milk hydrolysed by IE, possibly due to the presence of smaller fat globules, which did not affect the lactose hydrolysis. No significant differences were observed for lactulose synthesis in whole milk from the different species evaluated. The lactulose synthesis in cow and sheep milk by the enzymatic route may be a promising alternative, though further studies are required to optimise the production of this prebiotic in situ.
AOAC (2016). Official methods of analysis of AOAC international, 20th ed. AOAC International, Rockville.
Balthazar, C.F., Silva, H.L.A., Vieira, A.H., Neto, R.P.C., Cappato, L.P., Coimbra, P.T., Moraes, J., Andrade, M.M., Calado, V.M.A., Granato, D., Freitas, M.Q., Tavares, M.I.B., Raices, R.S.L., Silva, M.C., and Cruz, A.G. (2017). Assessing the effects of different prebiotic dietary oligosaccharides in sheep milk ice cream. Food Research International, 91: 38–46, https://doi.org/10.1016/j.foodres.2016.11.008.
Barbosa, O., Ortiz, C., Berenguer-Murcia, Á., Torres, R., Rodrigues, R.C., and Fernandez-Lafuente, R. (2015). Strategies for the one-step immobilization-purification of enzymes as industrial biocatalysts. Biotechnology Advances, 33(5): 435456, https://doi.org/10.1016/j.biotechadv.2015.03.006.
Bellé, A.S., Hackenhaar, C.R., Spolidoro, L.S., Rodrigues, E., Klein, M.P., and Hertz, P.F. (2018). Efficient enzyme-assisted extraction of genipin from genipap (Genipa americana L.) and its application as a crosslinker for chitosan gels. Food Chemistry, 246: 266–274, https://doi.org/10.1016/j.foodchem.2017.11.028.
Boitz, L.I. and Mayer, H.K. (2016). Analytical assessment of the intense heat load of whipped cream, coffee cream and condensed milk at retail in Austria and Germany. Dairy Science Technology, 96: 677–692, https://doi.org/10.1007/s13594-016-0295-0.
de Albuquerque, T.L., Gomes, S.D.L., D’almeida, A.P., Fernandez-Lafuente, R., Gonçalves, L.R.B., and Rocha, M.V.P. (2018). Immobilization of β-galactosidase in glutaraldehyde-chitosan and its application to the synthesis of lactulose using cheese whey as feedstock. Process Biochemistry, 73: 65–73, https://doi.org/10.1016/j.procbio.2018.08.010.
Klein, M.P., Fallavena, L.P., Schöffer, J.D.A.N., Ayub, M.A.Z., Rodrigues, R.C., Ninow, J.L., and Hertz, P.F. (2013). High stability of immobilized β-d-galactosidase for lactose hydrolysis and galactooligosaccharides synthesis. Carbohydrate Polymers, 95(1): 465–470, https://doi.org/10.1016/j.carbpol.2013.02.044.
Lane, J.H. and Eynon, L. (1923). Determination of reducing sugars by Fehling’s solution with methylene blue indicator. Journal of the Society of Chemical Industry, 42: 32–37.
Lima, P.C., Gazoni, I., Carvalho, A.M.G. de, Bresolin, D., Cavalheiro, D., Oliveira, D., and Rigo, E. (2022). β-galactosidase from Kluyveromyces lactis in genipin-activated chitosan: an investigation on immobilization, stability, and application in diluted UHT milk. Food Chemistry, 349: 129050, https://doi.org/10.1016/j.foodchem.2021.129050.
Neto, C.A.C.G., Silva, N.C.G., Costa, T.O., Albuquerque, T.L., Gonçalves, L.R.B., Fernandez-Lafuente, R., and Rocha, M.V.P. (2021). The β-galactosidase immobilization protocol determines its performance as catalysts in the kinetically controlled synthesis of lactulose. International Journal of Biological Macromolecules, 176: 468478, https://doi.org/10.1016/j.ijbiomac.2021.02.078.
Raza, A., Iqbal, S., Shah, F.H., Ahmad, Z., Rehman, M.A., Waseem, M., and Usman, M. (2021). Conversion of milk lactose to galacto-oligosaccharides by enzymes to produce prebiotic enriched cheese. Future Foods, 4: 100097, https://doi.org/10.1016/j.fufo.2021.100097.
Rentschler, E., Schuh, K., Krewinkel, M., Baur, C., Claaben, W., Meyer, S., Kuschel, B., Stressler, T., and Fischer, L. (2015). Enzymatic production of lactulose and epilactose in milk. Journal of Dairy Science, 98(10): 6767–6775, https://doi.org/10.3168/jds.2015-9900.
Rodrigues, R.C., Ortiz, C., Berenguer-Murcia, A., Torres, R., and Fernandez-Lafuente, R. (2013). Modifying enzyme activity and selectivity by immobilization. Chemical Society Reviews Journal, 42: 62906307, https://doi.org/10.1039/c2cs35231a.
Shakoor, S., Singh, G., and Singh, M. (2018). Glucose isomerase production and its application in various field. International Journal of Scientific & Technology Research, 4(5): 690699, https://doi.org/10.32628/IJSRST1845102.
Singh, P., Arora, S., Rao, P.S., Kathuria, D., Sharma, V., and Singh, A.K. (2022). Effect of process parameters on the β-galactosidase hydrolysis of lactose and galactooligosaccharide formation in concentrated skim milk. Food Chemistry, 393: 133355, https://doi.org/10.1016/j.foodchem.2022.133355.
Song, Y.S., Lee, H-U., Park, C., and Kim, S-W. (2013). Optimization of lactulose synthesis from whey lactose by immobilized β-galactosidase and glucose isomerase. Carbohydrate Research, 369: 1–5, https://doi.org/10.1016/j.carres.2013.01.002.
Vera, C., Córdova, A., Aburto, C., Guerrero, C., Suárez, S., and Illanes, A. (2016). Synthesis and purification of galacto-oligosaccharides: state of the art. World Journal of Microbiology & Biotechnology, 32: 197, https://doi.org/10.1007/s11274-016-2159-4.
Wang, G., Zhu, J., Liu, L., Yaqoob, M.U., Pei, X., Tao, W., Xiao, Z., Sun, W., and Wang, M. (2020). Optimization for galactooligosaccharides synthesis: a potential alternative for gut health and immunity. Life Sciences, 245: 117353, https://doi.org/10.1016/j.lfs.2020.117353.