Elsevier

International Dairy Journal

Volume 85, October 2018, Pages 137-143
International Dairy Journal

Addition of proline-rich whey peptides during dehydration increases solubility of rehydrated milk protein concentrates

https://doi.org/10.1016/j.idairyj.2018.05.010Get rights and content

Abstract

Protein solubility of milk protein concentrate (80% protein; MPC80) is compromised by condition-dependent processing and storage due to both physical and chemical self-association of caseins. Solubility of MPC80 was studied following storage at 4 °C to selectively focus on efficacy of adding the whey-proline rich peptide hydrolysate, ‘DISSPEP’, in regulating physical protein aggregation processes. A range of concentrations of total solids [5% (control), 50%, 90%, w/w], DISSPEP [0% (control), 5%, 10%, 25% of solids substituted], storage and rehydration pH (6.0, 6.5, 7.5) and storage time [0 (control), 7, 14 days], were examined. Relative to controls, the addition of DISSPEP generally improved MPC80 protein solubility by factors of 50–600%, dependent on storage time and pH. DISSPEP was particularly effective for rehydrating MPC80 following concentration and drying to 50%–90% solids, respectively. The results suggest potential industrial usefulness of DISSPEP for improving solubility of MPCs, particularly at low temperatures.

Introduction

Australia's growing dairy industry, representing 7% of world trade, exports around 38% of its produce (Dairy Australia Limited, 2014) necessitating efficient processing into transportable solids and market-oriented product functionality. Australian processors market a diverse range of dried dairy products reflecting fractionation into the different macro-nutrient chemical classes of milk, including whey and casein sub-fractions of milk proteins (Dairy Australia Limited, 2014). In particular, milk protein concentrates (MPC) represent a protein-enriched variant of skim milk produced by membrane-concentration and spray-drying the total protein fraction from skim milk (Mao et al., 2012, Singh, 2007). MPCs, while maintaining the whey and casein proportions of milk and micellar casein structure, are characterised by the degree of protein enrichment (e.g., MPC35, MPC80) and consequent depletion of lactose and minerals.

While representing a potentially versatile ingredient for reconstruction of dairy products and food ingredient (Mao et al., 2012), MPCs manifest compromised solubility due to hydrophobic interactions, particularly between caseins (Mimouni, Deeth, Whittaker, Gidley, & Bhandari, 2010) that develop during concentration. Solubility is further compromised by chemical reactivity between proteins, including non-reducible cross-linkages and lactosylation, that can occur as a function of storage time (Le et al., 2011, Mimouni et al., 2010, Schokker et al., 2011), pH (Mao et al., 2012), humidity and temperature (Le et al., 2011). As a result, MPCs display poor solubility at low pH (De Castro-Morel & Harper, 2002) and ambient or chilled temperatures, and require heating in most food applications (Singh, 2016).

Apart from driving milk protein cross-linking and aggregation as a consequence of heating (Singh, 2007), the concentration of MPCs is accompanied by elimination of minerals and lactose that promotes specific hydrophobic and chemical interactions between caseins which accounts for their compromised solubility (Anema, Pinder, Hunter, & Hemar, 2006). Casein insolubility in MPCs increases with protein concentration (Baldwin & Truong, 2007) and also with spray-drying temperature (Fang, Rogers, Selomulya, & Chen, 2012).

Efforts to manage and improve protein solubility of MPCs include the use of physical treatments such as high pressure processing (Udabage, Puvanenthiran, Yoo, Versteeg, & Augustin, 2012), use of ultrasound (Bhandari and Zisu, 2016, Sun et al., 2014), regulating drying conditions (Augustin, Sanguansri, Williams, & Andrews, 2012) and porosification of concentrates before drying (Bouvier, Collado, Gardiner, Scott, & Schuck, 2013). Alternately, chemical interventions including agglomeration with emulsifiers (Gaiani, Schuck, Scher, Desobry, & Banon, 2007), addition of milk minerals (Schuck, 2014) and salts (Mao et al., 2012), may also be effective. However, non-protein additives dilute the nutritional quality of MPCs and may introduce undesirable sensory characteristics. In general, there is further opportunity to find alternative solutions to mitigate the poor solubility of MPCs and reproduce the protein functionality of fresh milk.

The structural organisation of caseins into micelles is driven by co-operative hydrophobic and H-bonding and specific interactions of proline-glutamine-rich sequences (Holt, Carver, Ecroyd, & Thorn, 2013). Furthermore, these interactions are also operative in individual caseins and can drive either self-assembly of fibril structures in the case of κ- and αS2-caseins or alternately, the fibrillar assembly of these caseins can be inhibited by the ‘chaperone’ interactions with either β-casein or αS1-caseins (Holt et al., 2013).

The understanding that protein segments containing proline-glutamine-rich primary sequence were important for chaperone interactions, led to the investigation of efficacy of whey protein hydrolysates, also rich in these amino acids, on inhibition of fibril development (Bennett et al., 2009). A whey proline-rich peptide (wPRP) hydrolysate was subsequently demonstrated to inhibit in vitro fibril assembly and toxicity of human beta amyloid peptides, whose accumulation in the brain is a key pathology of Alzheimer's disease (Bharadwaj et al., 2013). Furthermore, the same wPRP product was also demonstrated to prevent aggregation and thereby promote solubility of heated milk proteins, mainly by regulating physical aggregation processes, with an additive effect following storage at 4 °C (Selby-Pham et al., 2013).

A simplified process was subsequently developed involving fewer enzymes (3 versus 5 previously) and did not involve hydrophobic peptide enrichment by C18 solid-phase extraction chromatography. The product from the modified process is referred to here as ‘DISSPEP’ (‘DISSolving PEPtide’) and the aim of this study was to determine its efficacy on regulating solubility of MPC80 during storage at 4 °C, by including the following experimental parameters: MPC solids concentration during storage, storage time, concentration of DISSPEP and pH.

Section snippets

Materials

Micellar milk protein concentrate (MPC) was obtained from MyoPure (81.5% protein, Petersham, NSW, Australia). Buffer salt 2-N-morpholino-ethanesulfonic (MES) acid was from Sigma–Aldrich (St Louis, MO, USA). All other reagents were analytical grade.

Proline-rich peptide product (DISSPEP) was prepared by enzyme hydrolysis of whey protein isolate (WPI, Natrapro (lecithin-free), Murray Goulburn, Southbank, Victoria, Australia) using the pilot plant facilities of CSIRO Agriculture and Food (Werribee,

Effect of pH and extent of dehydration on solubility of MPC

Total solubility of milk proteins in MPC (PSOL assay) included correction for light scatter (Selby-Pham et al., 2013) and also for the contribution of added DISSPEP, using a calibration profile prepared at respective experimental pH values. Regression analysis of the calibration curves for DISSPEP reconstituted at 3 experimental pH values (6.0, 6.5 and 7.5) indicated that the slopes were linear and equivalent (p > 0.05), and therefore that the absorbance of DISSPEP was not pH dependent and the

Factors affecting solubility of MPCs

Concentration and drying of total milk proteins generates a similar range of products to concentrated forms of whey proteins, including ‘concentrates’ (MPC, containing 35–80% milk proteins) and isolates (MPI, containing > 80% milk proteins). Unlike whey proteins, which are typically subjected to low heat processing, total milk protein concentrate products may be subjected to higher heat treatments leading to protein physical and chemical aggregation prior to concentration and drying, and appear

Conclusions

The presence of DISSPEP prevented the loss of solubility of MPC proteins during storage to extents that depended on pH and concentration of total solids. The relationship to substitution level of DISSPEP was generally minimal suggesting that lower levels of DISSPEP addition might achieve similar results. Sustained improvement in MPC protein solubility after 14 days at 4 °C with added DISSPEP ranged from 200 to 500%, which suggests that the benefits of added DISSPEP on protein solubility were

Acknowledgements

Technical assistance from Drs Roderick Williams and Hema Jegasothy are gratefully acknowledged. Amino acid analysis was conducted at the Australian Proteome Analysis Facility Ltd using infrastructure provided by the Australian Government through the National Collaborative Research Infrastructure Strategy (NCRIS).

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