Vol 62 International Journal of Dairy Technology 1 ORIGINAL RESEARCH *Author for correspondence. E-mail: desmond.morgan@ucd.ie © 2008 Society of Dairy Technology doi: 10.1111/j.1471-0307.2008.00435.x Blackwell Publishing Ltd Oxford, UK IDT International Journal of Dairy Technology 1364-727X 1364-0307 Society of Dairy Technology 2008 XXX ORIGINAL RESEARCH ORIGINAL RESEARCH Effect of high intensity pulsed electric fields on enzymes and vitamins in bovine raw milk JOERG RIENER, FRANCESCO NOCI, DENIS A CRONIN, DESMOND J MORGAN* and JAMES G LYNG UCD School of Agriculture Food Science and Veterinary Medicine, College of Life Sciences, UCD Dublin, Belfield, Dublin 4, Ireland Pulsed electric fields (PEF) were applied to fresh bovine raw milk using a laboratory-scale continuous PEF system to study the impact on selected native enzyme activities and on vitamin degradation. The activities of protease, lipase, alkaline phosphatase and lactoperoxidase following PEF treatment at electric field strengths ranging from 15 to 35 kV/cm for treatment times of 12.5 μs to 75 μs were examined. Reductions in enzyme activity, even under the most severe conditions employed, were quite modest, amounting to 14%, 37% and 29% for lipase, protease and alkaline phosphatase, respectively, while the activity of lactoperoxidase remained unchanged. The PEF treatment had no effect on the levels of thiamin, riboflavin, retinol and α-tocopherol in the milk. Keywords Bovine milk, Enzymes, Pulsed electric fields, Vitamins. *Author for correspondence. E-mail: desmond.morgan@ucd.ie INTRODUCTION Generally, fresh unpasteurized raw milk is heat treated to guarantee a microbiologically safe and shelf-stable product. While thermal treatments are very efficient, some undesirable organoleptic and chemical changes can occur. The latter may include the formation of lactulose, furosine or hydroxymethylfurfural, as well as some protein degradation (Morales et al. 1996; Contarini et al. 1997; Lopez-Fandino and Olano 1999; Villamiel et al. 1999; Calligaris et al. 2004). An interesting alternative to heat treatment is the application of pulsed electric fields (PEF), because of its effectiveness against most microorganisms, while flavour, colour and vitamins are only minimally or nonaffected (Sizer and Balasubramaniam 1999; Cserhalmi et al. 2003). The effect of PEF on enzyme inactivation has not been widely investigated and there is a lack of published information. In milk there are around 70 indigenous enzymes and several of these are significant in relation to the quality of milk and milk products (Kelly and Fox 2006). For the present study, four enzymes that have important roles in milk quality were selected. Lipases are enzymes that catalyse the hydrolysis of triglycerides, the major lipid components of milk. The significance of lipolysis in milk is twofold: flavour production and altered functionality (Ha and Lindsay 1993). Free fatty acids, particularly those of short and medium chain length, have strong flavours that are generally considered as undesirable (Balcao and Malcata 1998; Deeth 2006). The application of 20 pulses at 21.5 kV/cm to lipase in milk led to a complete inactivation (Grahl and Märkl 1996) (Table 1). Bendicho et al. (2002b) observed a reduction of 62.1% maximum activity when it was treated at 27.4 kV/cm and 80 pulses. However, when PEF treatments were applied in a continuous flow mode by the same research group, an inactivation rate of just 13% was achieved, after applying 80 pulses at 37.3 kV/cm and 3.5 Hz. Proteases are enzymes that degrade proteins. Raw milk may have proteolytic activity of endogenous or indigenous origin, which can adversely affect milk flavour (Chen et al. 2003). It was found that PEF reduced protease activity by 62.7% after an 896 μs treatment at 35.5 kV/cm and 111 Hz (Bendicho et al. 2003b). An 80% reduction in protease activity was obtained in tryptic soy broth with yeast extract after 20 pulses of 2 μs and 18 kV/cm at 0.25 Hz. When the protease was treated in sterilized skim milk, PEFs of 14 and 15 kV/cm and pulsing rates of 1 and 2 Hz achieved 40% and 60% inactivation after 32 and 98 pulses of 2 μs duration, respectively (Vega- Mercado et al. 1995; Vega-Mercado et al. 1997). Alkaline phosphatase (ALP) is a native enzyme found in raw milk. It is a rather nonspecific mono- phosphoester hydrolase that catalyses the removal of phosphate groups. Conventionally, the effective- ness of milk pasteurization processes is monitored by the inactivation of ALP, which is more heat resistant than the pathogens present in milk (Mussa