Freezing point and sensory quality of skim milk as affected by addition of ultrafiltration permeates for protein standardization

ht. Dairy Journal 6 (1996) 569-519 Copyright 0 1996 Elsevier Science Limited Printed in Ireland. All rights reserved 095%6946/96/$15.00 + 0.00 ELSEVIER

0958-6946(95)00062-3

Freezing Point and Sensory Quality of Skim Milk as Affected by Addition of Ultrafiltration Permeates for Protein Standardization

W. Rattray Department

of Agricultural, University (Received

3 March

& P. Jelen*

Food and Nutritional, 2-06 Agriculture and Forestry of Alberta, Edmonton, Canada, T6G 2P5 1995; revised version accepted

13 September

Centre,

1995)

ABSTRACT The protein content of skim milk was adjusted to values in the range 3.44-2.1% (w/w) by the addition of permeates obtained by ultrafiltration (UF) of skim milk (SMP), sweet whey (SWP) or acid whey produced by direct acidtfication of skim milk to pH 4.6 (A WP) or by fermenting skim milk to pH 4.7 with lactic acid bacteria (FWP). Addition of increasing amounts of SMP or SWP to skim milk caused the freezing point (FP) to increase progressively, but adding A WP or FWP had the opposite effect. Standardizing skim milk with appropriate ratios of UF permeates [A WPjSMP (40/60%), A WPjSWP (60/40%), FWPjSMP (401 60%) or FWP/SWP (50j50°%)/ caused negligible changes in the FP, thus reducing the possibility that such milk would be considered as adulterated or otherwise abnormal. Triangle sensory tests demonstrated that skim milk standardized with SMP or SWP to >2.4% (w/w) protein or A WP to >2.8% (w/w) protein was indistinguishable from normal skim milk, suggesting that these types of permeates would be suitable for industrial standardization. The use of A WP in combination with SMP or SWP (where the FP did not change) appeared also to be suitable The sensory quality of milk standardized with FWP for protein standardization. or a FWP/SMP combination (where the FP did not change) was unsatisfactory; even at 3.2% (w/w) protein or 3.0% (w/w) protein, respectively, protein-standardized milk had an unacceptably strong off-jl avour. Copyright 0 1996 Elsevier Science Limited * To whom correspondence

should be addressed. 569

570

W. Rattray, P. Jelen INTRODUCTION

Standardization of the protein content of milk is currently a topic of considerable interest to the dairy industry (Gungerich, 1994; Jensen, 1994). One proposed method of ‘downward’ standardization involves adding small quantities of ultrafiltration (UF) permeate to milk causing a slight reduction in protein content and negligible changes in pH, lactose and minerals. Apart from the slightly lower protein content, such protein-standardized milk is of nutritional quality similar to normal milk (Smith, 1995). The technological feasibility of continuous standardization of fat and protein in milk was shown by Rarnkilde Poulsen (1978) and Friis (1985, 1986). The heat stability of protein-standardized products does not seem to be compromised when standardization is performed with permeates obtained by UF of skim milk or sweet whey (Rattray & Jelen, 1996). Although the use of acid whey UF permeate was detrimental to heat stability, mild heat treatments, such as ordinary pasteurization, could still be feasible. The sensory quality of protein-standardized milk is crucial, especially if milk is for direct consumption. Rernkilde Poulsen (1978) standardized the protein content of skim milk, 1.8% fat milk and 3.5% fat milk, in the range 1.5-6.5% protein, using reverse osmosis (RO) or UF procedures. When the protein content of milk was altered by RO, the solids-not-fat (SNF) content changed also and this had a strong influence on the sensory quality; at f0.3% SNF the milk elicited either watery or salty/sweet sensations. In contrast, protein standardization by removal or addition of milk UF permeate did not change the SNF, making distinction between normal milk and protein-standardized milk very difficult, especially as the fat content increased; for skim milk, panelists could not detect differences between samples in which the protein content varied in the range 3. l6.5%, while for protein-standardized whole milk, the range was 1.5-6.4% protein. Milk standardized to low protein concentrations was identified by a translucent appearance rather than an altered taste. The freezing point (FP) of protein-standardized milk could be important from the regulatory viewpoint, because if the FP would be altered sufficiently, proteinstandardized milk would fall outside the ‘normal’ range and would be classilied as adulterated or otherwise abnormal. Although the interpretation of FP data can vary due to slight regional and seasonal variations and milk handling factors (van der Berg, 1979; Eisses & Zee, 1980; Sherbon, 1988; Schukker et al., 1992; Covney, 1993), in general, normal milk freezes at about -0.522”C, while a FP > -0.508”C is considered to be definite proof of extraneous water (AOAC, 1990). It should be noted that in much of the literature, FP data were expressed incorrectly as “C, when “H should have been used (Sherbon, 1988). Because the addition of UF permeate is likely to cause small changes in lactose and minerals, in addition to protein, changes in the FP may be anticipated. Presently, we are not aware of any published data on the FP of protein-standardized milk. In the present study, the potential suitability of four types of UF permeate for protein standardization was assessed by comparing the sensory quality and FP of normal skim milk to protein-standardized skim milk. A regular market skim milk was standardized by adding skim milk UF permeate (SMP); sweet whey UF permeate (SWP); and permeates obtained by UF of whey made by direct acidification of skim milk to pH 4.6 (AWP) or by fermentation of skim milk to

Freezing point and sensory quality

571

pH 4.7 with lactic acid bacteria (FWP). Most of the permeates and the proteinstandardized milk products were prepared on at least two separate occasions.

MATERIALS

AND

METHODS

Pasteurized (72°C I5 s), bulk skim milk, obtained from a local dairy company was processed on a pilot scale into four types of UF permeate, namely, SMP, SWP, AWP and FWP. The SMP was produced by direct UF of skim milk. Sweet whey, prepared by addition of rennet to skim milk (0.2 g L-l), was ultraliltered to obtain SWP. Two types of acid whey were prepared and ultrafiltered to obtain AWP or FWP. Direct acidification of skim milk with lactic acid to pH 4.6 produced whey for AWP. Fermentation of skim milk by lactic acid bacteria to pH 4.7 yielded whey for FWP. The fermentation was carried out at 35°C for 20 h using a mixed culture of Lactococcus lactis subsp. lactis and L. lactis subsp. cremoris, used normally in the manufacture of sour cream by Dairyworld Foods, Edmonton, Alberta. For production of all permeates, UF was carried out for 334 h, at 20°C and a cross-membrane pressure of 400 kPa, using a Carbosep system (SFEC Carbosep, Bollene, France), which consisted of a tubular arrangement of zirconium oxide membranes (cut-off, 20 kDa). The compositions of skim milk and UF permeates were determined by standard methods (AOAC, 1990). The protein content of skim milk was standardized to values in the range 3.442.1% (w/w) by adding calculated quantities of SMP, SWP, AWP or FWP or their combinations. The FP data for UF permeates, normal milk and protein-standardized milk were determined in triplicate by the Hortvet method (AOAC, 1990) using a Milk Cryoscope (Advanced Instruments Inc., MA). Cryoscopic FP data were converted from “H to “C according to AOAC (1990). The sensory quality of milk was evaluated by an untrained panel of 15 individuals selected from the personnel and students of the University of Alberta, using the triangle test method (Jellinek, 1985). Skim milk was especially suitable for sensory evaluation of the effects of UF permeate, as the absence of milk fat increases the sensitivity of individuals to changes in lactose and/or salts (Ronkilde Poulsen, 1978). Milk samples (-30 mL) were served at 15fl”C in small plastic cups, which were randomly numbered using a two digit code. On an individual basis, each panelist was presented with three samples and asked to identify the odd sample on the basis of appearance, taste and odour, guessing if necessary. The odd sample consisted of either protein-standardized skim milk or normal skim milk. The triangle test results were analyzed statistically using tables developed by Roessler et al. (1978).

RESULTS

Freezing point of protein-standardized

AND

DISCUSSION

milk

The FP of milk is influenced primarily by the content of lactose and dissolved salts, especially chloride (van der Have et al., 1980; Koops et al., 1989; Mitchell,

W. Rattray, P. Jelen

572

1989). Essentially, lactose and dissolved salts interact with water via polar and ionic groups, reducing the extent of hydrogen bonding between water molecules; thus, a lower temperature is needed to promote sufficient hydrogen bonding to initiate ice nucleation. Proteins interact with water in an extremely complex fashion, via charged, polar and hydrophobic groups (Edsall & McKenzie, 1983) but due to its high molecular weight, the molality of protein in milk is very low. Thus, protein has a finite, but negligible, effect on the FP of milk and protein standardization causing changes strictly to protein only would probably not change appreciably the FP. The FP of SMP or SWP was considerably higher than that of skim milk, obviously due to their lower lactose and ash contents (Table 1); hence, the addition of increasing quantities of these permeates to skim milk caused the FP to increase steadily (Fig. 1). Because about 2/3 of the salts in milk are associated with casein micelles (Green et al., 1984; Holt, 1985) the relatively low ash content of SMP or SWP was probably caused by removal of colloidal calcium phosphate (CCP) upon separation of the casein by UF or through the action of chymosin, respectively. The relatively high salt content and titratable acidity of the AWP and FWP in comparison to the SMP or SWP probably contributed to their low FP (Table 1). Clearly, the high titratable acidity of the acid permeates was caused by direct addition of lactic acid to skim milk or by fermentation of lactose to lactic acid, involved in the production of AWP or FWP, respectively. The high salt content of the permeates could be attributed to the well-known dissolution of colloidal calcium phosphate from casein micelles upon acidification (Holt, 1985). The

-“.46 5 -0.48 -

-0.56 -

-0.58 1 2.0

2.2

2.4

2.6 % (w/w)

2.8

3.0

3.2

3.4

Protein

Fig. 1. Freezing point of normal skim milk (3.44%, w/w, protein; -) or skim milk with protein content standardised to values in the range 3.442.1% (w/w) protein, by addition of ultrafiltration permeates (0) skim milk permeate; (0) sweet whey permeate; (0) acid whey permeate from whey produced by direct acidification of skim milk to pH 4.6 with lactic acid; (m) acid whey permeate from whey produced by fermentation of skim milk to pH4.7 with lactic acid bacteria. A FP > -0.508”C (---) is considered proof of adulteration.

573

Freezing point and sensory quality

TABLE 1 Average (n = 2) composition (%,w/w), pH and freezing point of skim milk or ultratiltration (UF) permeates obtained from skim milk (SMP), sweet whey (SWP), acid whey produced by direct acidification of skim milk (AWP) or acid whey produced by fermentation of skim milk (FWP)

Total solids Protein” Lactose Ash Titratable acidity PH Freezing point (“C)

Skim milk

SMP

SWP

AWP

FWP

9.5 3.44 5.0 0.70 0.168 6.7 -0.521

5.11 0.33 4.4 0.31 0.086 6.7 -0.465

5.19 0.31 4.4 0.32 0.119 6.7 -0.420

5.90 0.36 4.3 0.57 0.511 4.6 -0.593

5.58 0.29 4.1 0.64 0.502 4.7 -0.610

a % Protein = % N x 6.38. The nitrogen content of UF permeates most probably corresponded to low molecular weight peptides and urea capable of passing through the UF membrane. For the purpose of standardization, all permeates were assumed to have 0% true protein.

addition of AWP or FWP to skim milk was expected to increase the concentration of dissolved salts and lactic acid, thus reducing the FP of proteinstandardized milk, as was indeed observed (Fig. 1). Although the total ash of the skim milk was slightly higher than that of the FWP or AWP, most of this would be in the colloidal state and, presumably, have little impact on the FP. In view of the present results, if protein standardization were to become widespread, the FP measurement should be carried out prior to the addition of UF permeate, to avoid misinterpretation of FP data. In practice, this is likely to occur, as protein standardization can be expected to be performed on bulk milk while FP measurements are usually used to test the quality of milk from individual farmers, rather than bulk milk. It has been recommended that the FP of milk should not exceed -0.508”C; otherwise, the milk almost certainly contains extraneous water (AOAC, 1990). If the same criteria were to be applied to the protein-standardized milk, the FP of our milk, standardized with SMP to 62.6 or SWP to d 3.1% protein, would classify the milk as adulterated (Fig. 1). On the contrary, the low FP of milk standardized with AWP or FWP would be indicative of abnormal milk. Because the addition of UF permeates to skim milk may become an accepted industrial practice due to its negligible effects on sensory quality (Ronkilde Poulsen, 1978; also see below), designating protein-standardized milk as ‘adulterated’ or otherwise abnormal could well be unjustified. Nevertheless, under the present regulations valid in many countries, an altered FP might compromise the acceptability of protein-standardized milk. Under certain conditions, protein-standardized milk could have the same FP as normal milk; thus, the use of FP as an indicator of aceptability of the final product would be questionable. It was observed that the increasing addition of AWP to SMP or AWP to SWP caused the FP to increase gradually (Fig. 2) presumably due to increased concentrations of salts and lactic acid. Critical

574

W. Rattray,

P. Jelen

combinations of AWP and SMP (40/60%) or AWP and SWP (60/40%) had a FP close to -0.521”C, i.e. the same as normal skim milk. Hence, by adding these particular combinations of UF permeate to skim milk, the protein content could be standardized down to at least 2.1% (w/w) with negligible changes in FP (Fig. 3). Similarly, appropriate ratios of FWP to SMP (40/60%) or FWP to SWP (50/50%) were established (data not shown). Sensory quality of protein-standardized

milk

The sensory impact of bovine milk, as an interaction of taste, odour and texture, is a very complex phenomenon, especially in the presence of milk fat. In the absence of milk fat, the taste is influenced primarily by lactose and salts, with protein having a relatively minor effect (Pangborn & Dunkley, 1966). An exceedingly complex ‘cocktail’ of compounds contributes to the odour of milk; Badings & Neeter (1980) identified approximately 400 volatile compounds in whole milk, but it was not possible to attribute a milk-like odour to any one compound or restricted group of compounds. It appears likely that if protein standardization involved changes in protein only, the taste or odour would not be altered significantly. In our sensory evaluation trials, protein-standardized milk containing SMP or SWP was difficult to distinguish from unaltered skim milk. Standardization was possible to > 2.4% (w/w) protein before significant differences were observed (Table 2). In agreement with Rsnkilde Poulsen (1978) milk with a low protein content was detected due to a slightly translucent appearance, rather than to any noticeable change in taste or odour. As in previous investigations (Rattray & Jelen, 1996) protein-standardized and normal skim milk had equal pH values

-0.40 -0.42

2

-0.46 -

a ‘Z ;

-0.48 -0.50 -

33

-0.52 -

c

-0.54 -0.56 -

-0.60

’

I

1cOm

80/20 Ratio

7MO (g/g)

60/N

so/50

of AWP/SMP

40/60

3370

20/%0 o/loo

or AWP/SWP

Fig. 2. Freezing point of skim milk (-), combinations of acid whey permeate and skim milk permeate (AWP/SMP) (0) or combinations of acid whey permeate and sweet whey permeate (AWP/SWP) (0). Acid whey was made by direct acidification of skim milk to pH 4.6, using lactic acid.

Freezing point and sensory quality

-0.53

! 2.0

2.2

2.4

2.6

2.8

8 3.0

3.2

575

3.4

% (w/w) Protein

Fig. 3. Freezing protein content a 40/60 ratio of permeate/sweet

point of normal skim milk (3.44%, w/w, protein; -) or skim milk with standardized to values in the range 3.442.1% (w/w) protein by addition of acid whey permate/skim milk permeate (0) or a 60/40 ratio of acid whey whey permeate (a). A FP> -0.508”C (---) is considered proof of adulteration.

(pH 6.7) which may have contributed to their indistinguishability. Clearly, the increased translucency resulted from a lower concentration of casein micelles which reduced the ability of skim milk to scatter light. Although small decreases in the content of lactose and salts occurred on addition of permeates, as evident by the altered FP of protein-standardized milk (Fig. l), these were insufficient to be detected by the taste panel. In practice, it is unlikely that milk will be standardized down to 2.4% (w/w) protein; therefore, industrial standardization with SMP or SWP would not reduce sensory acceptability. Standardization of skim milk with AWP was possible to > 2.8% (w/w) protein before a noticeable alteration of sensory quality was observed (Table 2). At 2.8% (w/w) protein, the presence of AWP was just perceptible; most participants reported that the milk had a salty and slightly acidic taste, probably caused by increased concentrations of dissolved salts and lactic acid. The slight decrease in pH upon addition of AWP also may have affected the sensory quality of the protein-standardized milk. However, standardization to 3 3.0% (w/w) protein produced milk that was essentially indistinguishable from normal skim milk, indicating that AWP might be suitable also for a limited degree of standardization. The FWP was not suitable as a protein standardizing agent under the conditions of this study (Table 2). Even at 3.2% (w/w) protein, where the pH change was negligible, a strong off-flavour in the milk was detectable by olfaction or gustation. Doubtless, the off-flavour was caused by low molecular weight compounds generated during the fermentation of lactose. The principal volatile compounds produced by many lactic acid bacteria include diacetyl, acetaldehyde, dimethyl sulphoxide, acetic acid and lactic acid (Lindsay et ul., 1967) all of which should easily pass through the UF membrane used. The exact compound(s)

W. Rattray, P. Jelen

516

TABLE 2 Sensory and pH differences between normal skim milk (3.44%, w/w, protein; pH 6.7) and skim milk standardized to 3.2-2.4% (w/w) protein by addition of calculated quantities of individual ultrafiltration (UF) permeates or UF permeate combinations, as determined by triangle tests with 15 individuals Permeate typea

% (w/w) Protein of proteinstandardized skim milk

pH of proteinstandardized skim milk

Number of correct resultsb

SMP

3.2 2.8 2.4

6.7 6.1 6.1

4 (n.s.) 5 (n.s.)

SWP

3.2 2.8 2.4

6.7 6.7 6.1

4 (n.s.) 6 (ns.) 10 (**)

AWP

3.2 3.0 2.8 2.4

6.6 6.6 6.5 6.4

3 (n.s.) 3 (n.s.) 9 (*) 13 (***)

FWP

3.2 3.0 2.8

6.6 6.6 6.4

12 (***) 12 (***) 14 (***)

AWPjSMP

2.8 2.4

6.6 6.5

4 (n.s) 10 (**)

3.2 3.0

6.7 6.7

6 (n.s.)

(40160) FWPjSMP (40160)

9 (*)

9 (*)

a Permeates were obtained by UF of skim milk (SMP), sweet whey (SWP), acid whey produced by direct acidification of skim milk (AWP) or acid whey produced by fermentation of skim milk (FWP). Standardization was on the basis that all permeates contained 0% true protein; skim milk (3.44%, w/w, protein) was standardized to 3.2, 3.0, 2.8 or 2.4% (w/w) protein, by addition of UF permeates at a ratio of 7.5, 15, 23 or 43 g, per 100 g of skim milk, respectively. bNo significant difference (n.s.) or significant differences between normal skim milk and protein-standardized skim milk for P < 0.05, 0.01 or 0.001, as indicated by (*), (**) or (***), respectively.

Freezing point and sensory

quality

577

responsible for the development of the off-flavour in the protein-standardized milk were not determined in this study. It is likely that the nature and extent of off-flavour development in milk standardized with FWP would depend on the type of microorganisms used and the fermentation conditions used in the production of the acid whey. In the present study, the use of a sour cream culture could be expected to produce relatively high amounts of flavourful compounds. Perhaps FWP obtained by UF of whey resulting from other types of fermented dairy products would be less deleterious to the sensory quality of proteinstandardized milk. As shown above, we established appropriate combinations of UF permeates [AWP/SMP (40/60%), AWPjSWP (60/40%), FWPjSMP (40/60%) or FWPjSWP (50/50%)] that might be especially suitable for protein standardization, if it was desired to obtain protein-standardized milk with an unaltered FP. To test further the suitability of these combinations for standardization, the sensory quality of normal skim milk was compared to skim milk standardized with the appropriate UF permeate combinations. The results indicated that the sensory quality of milk standardized with mixtures of AWP and SMP (40/60%) was satisfactory (Table 2) which was not surprising since the use of either permeate alone was also satisfactory. Moreover, the addition of SMP to AWP extended the limit of detectability to 62.8% (w/w) protein, compared to the use of AWP alone, possibly due to smaller changes in pH, salts and/or lactic acid. Although FWP/ SMP combinations were less deleterious to the sensory quality than FWP alone, these mixtures were still unsuitable for protein standardization, as a noticeable off-flavour was detected already in milk standardized to 3.0% (w/w) protein.

CONCLUSIONS SMP or SWP appear to be highly suitable materials for standardizing the protein content of skim milk, provided the resultant increase in FP is not interpreted as evidence of adulteration; such an interpretation may not be justified as changes in sensory quality were insignificant at > 2.4% (w/w) protein and nutritional changes, especially at moderate levels of protein down standardization, may be expected to be unimportant. Triangle test results also suggest the suitability of AWP for protein standardization; sensory changes were insignificant at > 2.8% (w/w) protein; however, the substantially decreased FP of milk standardized with AWP could lead to such milk being outside the regulatory limits of acceptability. If the alteration of the FP of milk were to be avoided during standardization, then a possible solution might be to standardize milk with an appropriate combination of UF permeates (AWP/SMP, AWP/ SWP), which caused no change in the FP and sensory properties also appeared acceptable. The use of FWP for protein standardization was unacceptable; apart from possible problems associated with the lower FP of protein-standardized milk, a strong off-flavour was evident even after very small additions of FWP to skim milk. It should be emphasized that the four kinds of UF permeate used in this study were experimental by-products obtained from a limited number of UF trials using skim milk as the primary raw material. Many types of whey are produced industrially, due primarily to the diversity of cheese manufacture (Sienkiewicz &

578

W. Rattray,

P. Jelen

Riedel, 1990). The composition of UF permeates and hence their suitability for protein standardization will depend both on the type of whey and conditions during the subsequent UF process. Thus, the data presented in this study give only an illustration of the issues likely to be involved in determining the suitability of different UF permeate types for protein standardization.

ACKNOWLEDGEMENTS The authors express sincere thanks to Lawerence Roth and other personnel of Alberta Agriculture for assistance with the determination of freezing points. Additional gratitude is conveyed to staff and students at the University of Alberta who participated in sensory evaluation trials.

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