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Acid Hydrolysis of Lactose in Whey Versus Aqueous Solutions
Acid Hydrolysis of Lactose in Whey Versus Aqueous Solutions A. Y. LIN and T. A. NICKERSON Department of Food Science and Technology University of California Davis 95616 ABSTRACT
Hydrolysis was over 90% in a 30% (wt/vol) solution of lactose in 2N sulfuric acid at 60 C for 36 h. The brown color that developed during hydrolysis was removed easily with activated carbon. Slight offflavors were removed completely by treatment with calcium hydroxide to about pH 6 followed by activated carbon, yielding a syrup of excellent color and flavor. This process minimized anions and cations in the hydrolyzed syrup. Neutralization with lime and treatment with activated carbon gave a syrup with equisweetness to 20.8% sucrose (g/100 ml). Acid hydrolysis of concentrated whey with sulfuric acid produced a more severe browning reaction and off-flavor. Treatments with calcium hydroxide and activated carbon that were effected in aqueous lactose hydrolysates were ineffective in hydrolyzed whey. Sodium bisulfate inhibited color development during hydrolysis of whey but slowed hydrolysis, affected flavor, and interfered with analytical methods. INTRODUCTION
The seriousness of the whey disposal problem warrants multiple approaches to a solution. Hydrolysis is one approach. Previous papers (5, 7) have discussed the efficiency of hydrolysis in aqueous lactose solutions by sulfuric acid as measured by modified colorimetric methods [lactose (L) test, glucose-galactose (G-G) test, and Glucostat]. This paper reports the results of neutralizing the H2SO4-treated syrup with Ca(OH)2 and its effect on the flavor quality of the resulting syrups after decolorization since this procedure minimizes the salts resulting from acid hydrolysis and subsequent neutraliza-
tion. Also investigated was sulfuric acid hydrolysis of lactose in concentrated whey since this would be desirable for many end uses. Rates of reactions and changes during hydrolysis were observed. Attempts were made to inhibit browning and development of off-flavor during whey hydrolysis. EXPERIMENTAL
Preparation of H~SO4-Treated Syrup
Only analytical grade chemicals and U.S.P. alpha-lactose hydrate powder (from Foremost Food Company, San Francisco) were used in this study. Lactose was dissolved in 2N H2SOa to give 30% (wt/vol) on an anhydrous lactose basis. The mixture was heated prior to storage in an air oven at 60 C for 36 h. Heat treatment and sampling techniques were described previously (7). Lactose and glucose-galactose were determined by modified methods described by Nickerson et al. (5). After hydrolysis the mixture was cooled and then neutralized by slowly added 5.8 g Ca(OH)2, moistened with 10 ml distilled water for each 100 ml hydrolysate. The mixture was stirred constantly by a magnetic stirrer and after 30 min was filtered. The pH of the filtrate was in the range of 6.0 to 6.5. If the pH was higher, it was adjusted by adding a few drops of unneutralized syrup. The neutralized syrup was decolorized by adding .5% activated carbon, stirring for 20 min at 60 C, and filtering. The syrup then was stored at about 8 C until needed. Preparation of Hydrolyzed Whey Samples
Powders were prepared by freeze-drying cottage cheese whey (from Crystal Cream and Butter Company, Sacramento, CA) and permeate following ultrafiltration treatment in the department's processing laboratory. The powders were dissolved in 2N H2 SO4 to give 20 and 50% TS (wt/vol) respectively, mixed well, and
Received June 29, 1976.
34
ACID HYDROLYSIS OF LACTOSE IN WHEY held for 10 min prior to heating for hydrolysis. A concentrated whey also was prepared by dissolving nonfat dry milk in distilled water to give 30% TS (total solids) and precipitating the casein at pH 4.6 by slowly adding 2N H 2 SO4. The acidity was adjusted to 2N prior to hydrolysis by adding 10N H2SO4 in a ratio of 4 to 1. Measurement of Browning
A 30% TS (wt/vol) hydrolysis mixture w a s prepared by dissolving 30 g freeze-dried whey powder in 2N H2SOa. Sodium bisulfate w a s added to portions of the mixture to give final concentrations of .2, .4, .6, and 1.0 NaHSO3. These were heated to 60 C and stored in an air oven as described under Preparation of H2 SOatreated syrups. Samples were removed periodically during the hydrolysis period, filtered through Whatman No. 2 paper, and absorbance at 450 nm was read on a Beckman Model DB spectrophotometer. Absorbance o f hydrolyzed whey was maximum at 450 nm whereas absorbance of hydrolyzed lactose syrups was at 350 nm (7). RESULTS A N D DISCUSSION Neutralization of H 2SO 4-Treated Syrups
Hydrolyzed syrups must be neutralized to reduce acidity before they become edible. For 30% (wt/vol) lactose-hydrolyzed syrup, 5.8 g Ca(OH)2 were needed to neutralize 100 ml syrup and bring the pH to 6.0 to 6.5. Since the buffer capacity of the syrup was low, small changes in Ca(OH)2 caused a considerable change in pH. Excessive Ca(OH)2 must be avoided since it caused a color darkening that could not be removed by treatment with carbon. Browing increased with increases in pH above 7.
35
testing. The extent of hydrolysis was 78.9% a s measured by G-G test. The variables investigated were pH at time of treatment with activated carbon, neutralization with Ca(OH)2 before and after decolorization, and time allowed for decolorization. The five procedures are summarized in Table 1. The color produced during acid hydrolysis of aqueous lactose solutions was removed with activated carbon under both highly acid and slightly acidic conditions. Flavor evaluations with a few trained judges, however, indicated that flavor was improved when the activatedcarbon treatment was near pH 6 rather than below pH 4.5. These syrups were sweeter than the control sample containing glucose and galactose, probably because of the residual lactose, and were without any detectable aftertaste. Sweetness
A 30% lactose syrup was hydrolyzed to yield 24.12% glucose and galactose (determined by G-G test). Equisweetness (50% response by a select panel) was obtained by plotting responses of the panel. Figure 1 shows that 20.8% sucrose was required to give equisweetness to the hydrolyzed syrup. Sweetness of hydrolyzed syrup was considerably higher than expected from the relative sweetness of sucrose, glucose, galactose, and lactose as described by Cameron (1). Acid Hydrolysis of Lactose in Whey
Several investigators (2, 4, 6) have studied acid hydrolysis of lactose in whey, but none
IOC 9C 8c
Decolorization of Aqueous Lactose Hydrolysate
Treatment with activated charcoal is a routine procedure to remove off-flavors and colors in preparing commercial sugars and syrups. To determine conditions for optimum removal of off-flavor and color in lactose hydrolysates, a large quantity of 30% (wt/vol) lactose solution w a s hydrolyzed with 2N H2 SO4 at 60 C for 24 h and then divided into smaller samples for
~ 7c ~ 6c a:
5c
N 4c
EQUISWEET
"~/ I;.O
1£~.5
2;.0
2;.5
21'.0
% SUCROSE (g/lOOml)
FIG. 1. Relative sweetness of 30% lactose syrup (91.33% hydrolyzed). Journal of Dairy Science Vol. 60, No. 1
36
LIN AND NICKERSON
I00
O0
(L) -J 0
60
"r~
40
g z'4
,~
o
Y6
4~
"
TIME (h) F I G . 2. Percent hydrolysis o f 5 0 % permeate (contained 32.61 g lactose/lO0 ml) by using 2N H 2 SO 4 •
reported on hydrolysis of concentrated whey. Although conditions of hydrolysis and analytical procedures, such as the L test and the G-G test, were the same as those used previously on aqueous lactose solutions, the results were different. This was attributed to the milk salts and whey proteins which were not in the
E
-! 'Nt
~N
~
t~=t ~N
NO NaHSO~) .2M
.8 ,.
e~ el
~oo
f18 ~ e~
o ~
oo
.6
W
v
0 Z < m or" 0 .4 cO m
~
/
.4M
.~_ ~ ttt~tt
6M .2
0
0
e',
+ OM
0
0
0 ~O 0 ,4 .l < [..
J o u r n a l o f D a i r y S c i e n c e Vol. 6 0 , N o . 1
0~
12
24
36
48
"TIME{h) F I G . 3. C o l o r i n h i b i t i o n b y v a r i o u s c o n c e n t r a t i o n s o f N a H S O 3 in 3 0 % T S whey hydrolysis ( c o n t a i n e d 2 0 . 0 2 g l a e t o s e / l O 0 hal).
ACID HYDROLYSIS OF LACTOSE IN WHEY
37
TABLE 2. Effect of milk salts on the absorbance of color reactions. Sample absorbance As4 o nm (methylamine)
A~ t onm (molybdenum blue)
Milk salts
1
2
3
4b
5
6
7
8c
Not added Added
.046 .052
.295 .320
.595 .595
.940 .920
.146 .129
.258 .231
.352 .321
.442 .415
aprepared by method of Jenness and Koops (3). bsamples No. 1 to 4, respectively, contained lactose .5, 1.0, 1.5, and 2.0 mg/ml. CSamples No. 5 to 8, respectively, contained glucose and galactose (in equal amount) at 1.0, 2.0, 3.0, and 4.0 mg/ml.
lactose solutions. The milk-salt mixture gave little or no interference in the L test but uniformly decreased color intensity in the G-G test (Table 2). The decreased color in the G-G test can be explained by the reaction of milk salts with the reagents; light blue-green color developed as soon as the reagents were mixed with samples containing milk salt, even without heating. Table 3 shows rates of lactose hydrolysis in whey containing 7, 20, and 30% total solids (TS) by 2N H2SO4 at 60 C. Whey can be hydrolyzed under the same conditions as aqueous lactose solutions, though more slowly. Regardless of small differences in fat and ash contents, hydrolysis rates were similar in the two kinds of whey (sulfuric acid whey and cottage cheese whey), respectively, containing 11.4% (Table 3) and 12.8% (Table 4) lactose.
Hydrolysis of lactose in permeate from ultrafiltration treatment (low protein content), which contained 32.01 g lactose/100 ml (Fig. 2) was faster than in whey o f similar lactose content. Thus, whey protein may be the reason for the slow hydrolysis in whey, contrary to findings of Ney and Wirotoma (4), who suggested that whey proteins actually accelerate hydrolysis. During hydrolysis of whey, more than 90% glucose and galactose was recovered as measured by the G-G test. This is higher than in aqueous lactose solutions (7). Thus, it may be that whey proteins and salt function to prevent loss of hexoses. On the other hand, the hydrolysis value was lower from the L test than from the G-G test during whey hydrolysis. This was caused by more serious browning during whey hydrolysis. Glucose became brownish instead of light yellow when heated in 2N H 2 SOa (with or
TABLE 3. Effect of whey concentration on lactose hydrolysis by 2N H2SO+ as measured by the L and G-G tests. Hydrolysis L-test
G-G test
Time
7% TS
20% TS
30% TS
(h) 6 12 21 24 30 36 42 48
20.30 37.15 55.41 56.80 61.10 69.96 84.90 84.50
22.22 40.45 59.40 60.77 65.41 79.91 83.19 84.37
22.59 36.66 66.55 68.68 74.06 84.00 88.18 88.67
7% TS
20% TS
30% TS a
26.97 44.66 68.04 79.44 85.69 86.29 88.21 90.16
25.14 38.77 66.67 73.52 78+73 86.45 91.66 97.41
25.08 40.64 65.46 69.86 77.59 78.46 86.59 89.04
(%)
a7% TS = 5.0% lactose; 20dATS = 11.4% lactose; 30% TS = 17.8% lactose. Journal of Dairy Science Vol. 60, No. 1
38
LIN AND NICKERSON
TABLE 4. Rate of hydolysis of 20% TS cottage cheese whey a by 2N H~ SO, as measured by L and G-G tests,
TABLE 5. Effect of NaHSO 3 on 30% TS whey 2 hydrolysis measured by L test.
Hy drolysis L-test Time
Mean
SD
13.58 24.59 74.48 79.04 82.02 83.51
1.35 1.06 .43 .19 .16 .24
(h)
Hydrolysis
G-G test Mean SD Time
(%)
6 12 24 30 36 48
No NaHSO 3 added
(h) ... 23.82 72.97 77.78 87.05 90.81
_.o
1.84 3.01 .88 2.69 2.14
With .4 M NaHSO 3
(%)
6 12 21 24 30 36 48
4.38 8.79 14.85 24.77 28.74 27.25 31.66
9.06 26.70 41.58 57.29 65.11 70.24 77.40
a12.8 g anhydrous lactose/100 ml. a18.14 g anhydrous lactose/100 ml.
without amino acid present) before analysis. Browning was more obvious in the presence of amino acids, particularly at alkaline pH values. Therefore, whey syrups turned brown rapidly when heated if they were overneutralized. Browning was much darker and more complex during whey hydrolysis than during aqueous lactose hydrolysis. Activated carbon did not remove the color developed through sugaramino acid reactions during whey hydrolysis. Procedures similar to those that proved effective on aqueous lactose solutions (Table 5) were ineffective in clarifying hydrolyzed whey solutions; they remained discolored and were salty besides having a pronounced off-flavor (bitter). Sodium bisulfite has been used in many food products to prevent Maillard browning.
The present work found sodium bisulfite also effective in inhibiting browning of hydrolyzed whey. Figure 3 shows the degree of color inhibition by sodium bisulfite. Color development was prevented completely by 1.0 M sodium bisulfite during the hydrolysis of 20.02% whey lactose in 2N H2SO4. For an acceptable final color most of the browning reaction can be inhibited by .4 M sodium bisulfite. Although sodium bisulfite reduced Maillard browning during whey hydrolysis, it contributes other problems, such as developing a strong flavor, slowing hydrolysis, and interfering with analytical methods. Figures 4 and 5 show the effects of sodium bisulfite on both the L test and G-G test. In the presence of .01
I
.61
E zz
I.O
o
tO Z
~ .4 0r.o < .2
LACTOSE WITH
J
WITH I M NaHSO~
O
CONCENTRATION
{mg/ml}
FIG. 4. Effect of NaHSO~ on lactose test.
Journal of Dairy Science Vol. 60, No. 1
I.O
2.O
3 'O
4 iO - -
CONCENTRATION I rn9 / m l }
FIG. 5, Effect of NaHSO 3 on glucose-galactose test.
ACID HYDROLYSIS OF LACTOSE IN WHEY
M sodium bisulfite, color development decreased during the L test, but increased in the G-G test. Table 5 shows that the hydrolysis of whey is retarded in the presence of sodium bisulfite. The possibility of developing acidhydrolyzed syrups from whey with acceptable flavor and color, therefore, seems unlikely although the feasibility of producing such syrups from aqueous lactose solutions has been demonstrated. REFERENCES
Cameron, A. T. 1947. The taste sense and the relative sweetness of sugars and other sweet substances. Sugar Research Foundation Rept. No. 9, 1 to 72, New York.
39
2 Coughlin, J. R., and T. A. Nickerson. 1975. Acid-catalyzed hydrolysis of lactose in whey and aqueous solutions. J. of Dairy Sci. 58:169. 3 Jenness, R., and J. Koops. 1962. Preparation and properties of a salt solution which simulates milk ultrafiltrate. Neth. Milk & Dairy J. 16:153. 4 Ney, K. H., and I. P. G. Wirotoma. 1970. Investigation of lactose hydrolysis. Zeitschrift Vfir Lebensmittelunter-suchung und Forsch. 143 (2), 93; FS&T Abstr. 2 (11), 11 p. 1591 (1970). 5 Nickerson, T. A., I. F. Vujicic, and A. Y. Lin. 1976. Colorimetric estimation of lactose and its hydrolytic products. J. Dairy Sci. 59:386. 6 Ramsdeil, G. A., and B. H. Webb. 1975. The acid hydrolysis of lactose and the preparation of hydrolyzed lactose sirup. J. Dairy Sci. 28:677. 7 Vujicic, I. F., A. Y. Lin, and T. A. Nickerson. 1976. Changes during acid hydrolysis of lactose. J. of Dairy Sci. 60:29.
Journal of Dairy Science Vol. 60, No. 1
Hydrolysis was over 90% in a 30% (wt/vol) solution of lactose in 2N sulfuric acid at 60 C for 36 h. The brown color that developed during hydrolysis was removed easily with activated carbon. Slight offflavors were removed completely by treatment with calcium hydroxide to about pH 6 followed by activated carbon, yielding a syrup of excellent color and flavor. This process minimized anions and cations in the hydrolyzed syrup. Neutralization with lime and treatment with activated carbon gave a syrup with equisweetness to 20.8% sucrose (g/100 ml). Acid hydrolysis of concentrated whey with sulfuric acid produced a more severe browning reaction and off-flavor. Treatments with calcium hydroxide and activated carbon that were effected in aqueous lactose hydrolysates were ineffective in hydrolyzed whey. Sodium bisulfate inhibited color development during hydrolysis of whey but slowed hydrolysis, affected flavor, and interfered with analytical methods. INTRODUCTION
The seriousness of the whey disposal problem warrants multiple approaches to a solution. Hydrolysis is one approach. Previous papers (5, 7) have discussed the efficiency of hydrolysis in aqueous lactose solutions by sulfuric acid as measured by modified colorimetric methods [lactose (L) test, glucose-galactose (G-G) test, and Glucostat]. This paper reports the results of neutralizing the H2SO4-treated syrup with Ca(OH)2 and its effect on the flavor quality of the resulting syrups after decolorization since this procedure minimizes the salts resulting from acid hydrolysis and subsequent neutraliza-
tion. Also investigated was sulfuric acid hydrolysis of lactose in concentrated whey since this would be desirable for many end uses. Rates of reactions and changes during hydrolysis were observed. Attempts were made to inhibit browning and development of off-flavor during whey hydrolysis. EXPERIMENTAL
Preparation of H~SO4-Treated Syrup
Only analytical grade chemicals and U.S.P. alpha-lactose hydrate powder (from Foremost Food Company, San Francisco) were used in this study. Lactose was dissolved in 2N H2SOa to give 30% (wt/vol) on an anhydrous lactose basis. The mixture was heated prior to storage in an air oven at 60 C for 36 h. Heat treatment and sampling techniques were described previously (7). Lactose and glucose-galactose were determined by modified methods described by Nickerson et al. (5). After hydrolysis the mixture was cooled and then neutralized by slowly added 5.8 g Ca(OH)2, moistened with 10 ml distilled water for each 100 ml hydrolysate. The mixture was stirred constantly by a magnetic stirrer and after 30 min was filtered. The pH of the filtrate was in the range of 6.0 to 6.5. If the pH was higher, it was adjusted by adding a few drops of unneutralized syrup. The neutralized syrup was decolorized by adding .5% activated carbon, stirring for 20 min at 60 C, and filtering. The syrup then was stored at about 8 C until needed. Preparation of Hydrolyzed Whey Samples
Powders were prepared by freeze-drying cottage cheese whey (from Crystal Cream and Butter Company, Sacramento, CA) and permeate following ultrafiltration treatment in the department's processing laboratory. The powders were dissolved in 2N H2 SO4 to give 20 and 50% TS (wt/vol) respectively, mixed well, and
Received June 29, 1976.
34
ACID HYDROLYSIS OF LACTOSE IN WHEY held for 10 min prior to heating for hydrolysis. A concentrated whey also was prepared by dissolving nonfat dry milk in distilled water to give 30% TS (total solids) and precipitating the casein at pH 4.6 by slowly adding 2N H 2 SO4. The acidity was adjusted to 2N prior to hydrolysis by adding 10N H2SO4 in a ratio of 4 to 1. Measurement of Browning
A 30% TS (wt/vol) hydrolysis mixture w a s prepared by dissolving 30 g freeze-dried whey powder in 2N H2SOa. Sodium bisulfate w a s added to portions of the mixture to give final concentrations of .2, .4, .6, and 1.0 NaHSO3. These were heated to 60 C and stored in an air oven as described under Preparation of H2 SOatreated syrups. Samples were removed periodically during the hydrolysis period, filtered through Whatman No. 2 paper, and absorbance at 450 nm was read on a Beckman Model DB spectrophotometer. Absorbance o f hydrolyzed whey was maximum at 450 nm whereas absorbance of hydrolyzed lactose syrups was at 350 nm (7). RESULTS A N D DISCUSSION Neutralization of H 2SO 4-Treated Syrups
Hydrolyzed syrups must be neutralized to reduce acidity before they become edible. For 30% (wt/vol) lactose-hydrolyzed syrup, 5.8 g Ca(OH)2 were needed to neutralize 100 ml syrup and bring the pH to 6.0 to 6.5. Since the buffer capacity of the syrup was low, small changes in Ca(OH)2 caused a considerable change in pH. Excessive Ca(OH)2 must be avoided since it caused a color darkening that could not be removed by treatment with carbon. Browing increased with increases in pH above 7.
35
testing. The extent of hydrolysis was 78.9% a s measured by G-G test. The variables investigated were pH at time of treatment with activated carbon, neutralization with Ca(OH)2 before and after decolorization, and time allowed for decolorization. The five procedures are summarized in Table 1. The color produced during acid hydrolysis of aqueous lactose solutions was removed with activated carbon under both highly acid and slightly acidic conditions. Flavor evaluations with a few trained judges, however, indicated that flavor was improved when the activatedcarbon treatment was near pH 6 rather than below pH 4.5. These syrups were sweeter than the control sample containing glucose and galactose, probably because of the residual lactose, and were without any detectable aftertaste. Sweetness
A 30% lactose syrup was hydrolyzed to yield 24.12% glucose and galactose (determined by G-G test). Equisweetness (50% response by a select panel) was obtained by plotting responses of the panel. Figure 1 shows that 20.8% sucrose was required to give equisweetness to the hydrolyzed syrup. Sweetness of hydrolyzed syrup was considerably higher than expected from the relative sweetness of sucrose, glucose, galactose, and lactose as described by Cameron (1). Acid Hydrolysis of Lactose in Whey
Several investigators (2, 4, 6) have studied acid hydrolysis of lactose in whey, but none
IOC 9C 8c
Decolorization of Aqueous Lactose Hydrolysate
Treatment with activated charcoal is a routine procedure to remove off-flavors and colors in preparing commercial sugars and syrups. To determine conditions for optimum removal of off-flavor and color in lactose hydrolysates, a large quantity of 30% (wt/vol) lactose solution w a s hydrolyzed with 2N H2 SO4 at 60 C for 24 h and then divided into smaller samples for
~ 7c ~ 6c a:
5c
N 4c
EQUISWEET
"~/ I;.O
1£~.5
2;.0
2;.5
21'.0
% SUCROSE (g/lOOml)
FIG. 1. Relative sweetness of 30% lactose syrup (91.33% hydrolyzed). Journal of Dairy Science Vol. 60, No. 1
36
LIN AND NICKERSON
I00
O0
(L) -J 0
60
"r~
40
g z'4
,~
o
Y6
4~
"
TIME (h) F I G . 2. Percent hydrolysis o f 5 0 % permeate (contained 32.61 g lactose/lO0 ml) by using 2N H 2 SO 4 •
reported on hydrolysis of concentrated whey. Although conditions of hydrolysis and analytical procedures, such as the L test and the G-G test, were the same as those used previously on aqueous lactose solutions, the results were different. This was attributed to the milk salts and whey proteins which were not in the
E
-! 'Nt
~N
~
t~=t ~N
NO NaHSO~) .2M
.8 ,.
e~ el
~oo
f18 ~ e~
o ~
oo
.6
W
v
0 Z < m or" 0 .4 cO m
~
/
.4M
.~_ ~ ttt~tt
6M .2
0
0
e',
+ OM
0
0
0 ~O 0 ,4 .l < [..
J o u r n a l o f D a i r y S c i e n c e Vol. 6 0 , N o . 1
0~
12
24
36
48
"TIME{h) F I G . 3. C o l o r i n h i b i t i o n b y v a r i o u s c o n c e n t r a t i o n s o f N a H S O 3 in 3 0 % T S whey hydrolysis ( c o n t a i n e d 2 0 . 0 2 g l a e t o s e / l O 0 hal).
ACID HYDROLYSIS OF LACTOSE IN WHEY
37
TABLE 2. Effect of milk salts on the absorbance of color reactions. Sample absorbance As4 o nm (methylamine)
A~ t onm (molybdenum blue)
Milk salts
1
2
3
4b
5
6
7
8c
Not added Added
.046 .052
.295 .320
.595 .595
.940 .920
.146 .129
.258 .231
.352 .321
.442 .415
aprepared by method of Jenness and Koops (3). bsamples No. 1 to 4, respectively, contained lactose .5, 1.0, 1.5, and 2.0 mg/ml. CSamples No. 5 to 8, respectively, contained glucose and galactose (in equal amount) at 1.0, 2.0, 3.0, and 4.0 mg/ml.
lactose solutions. The milk-salt mixture gave little or no interference in the L test but uniformly decreased color intensity in the G-G test (Table 2). The decreased color in the G-G test can be explained by the reaction of milk salts with the reagents; light blue-green color developed as soon as the reagents were mixed with samples containing milk salt, even without heating. Table 3 shows rates of lactose hydrolysis in whey containing 7, 20, and 30% total solids (TS) by 2N H2SO4 at 60 C. Whey can be hydrolyzed under the same conditions as aqueous lactose solutions, though more slowly. Regardless of small differences in fat and ash contents, hydrolysis rates were similar in the two kinds of whey (sulfuric acid whey and cottage cheese whey), respectively, containing 11.4% (Table 3) and 12.8% (Table 4) lactose.
Hydrolysis of lactose in permeate from ultrafiltration treatment (low protein content), which contained 32.01 g lactose/100 ml (Fig. 2) was faster than in whey o f similar lactose content. Thus, whey protein may be the reason for the slow hydrolysis in whey, contrary to findings of Ney and Wirotoma (4), who suggested that whey proteins actually accelerate hydrolysis. During hydrolysis of whey, more than 90% glucose and galactose was recovered as measured by the G-G test. This is higher than in aqueous lactose solutions (7). Thus, it may be that whey proteins and salt function to prevent loss of hexoses. On the other hand, the hydrolysis value was lower from the L test than from the G-G test during whey hydrolysis. This was caused by more serious browning during whey hydrolysis. Glucose became brownish instead of light yellow when heated in 2N H 2 SOa (with or
TABLE 3. Effect of whey concentration on lactose hydrolysis by 2N H2SO+ as measured by the L and G-G tests. Hydrolysis L-test
G-G test
Time
7% TS
20% TS
30% TS
(h) 6 12 21 24 30 36 42 48
20.30 37.15 55.41 56.80 61.10 69.96 84.90 84.50
22.22 40.45 59.40 60.77 65.41 79.91 83.19 84.37
22.59 36.66 66.55 68.68 74.06 84.00 88.18 88.67
7% TS
20% TS
30% TS a
26.97 44.66 68.04 79.44 85.69 86.29 88.21 90.16
25.14 38.77 66.67 73.52 78+73 86.45 91.66 97.41
25.08 40.64 65.46 69.86 77.59 78.46 86.59 89.04
(%)
a7% TS = 5.0% lactose; 20dATS = 11.4% lactose; 30% TS = 17.8% lactose. Journal of Dairy Science Vol. 60, No. 1
38
LIN AND NICKERSON
TABLE 4. Rate of hydolysis of 20% TS cottage cheese whey a by 2N H~ SO, as measured by L and G-G tests,
TABLE 5. Effect of NaHSO 3 on 30% TS whey 2 hydrolysis measured by L test.
Hy drolysis L-test Time
Mean
SD
13.58 24.59 74.48 79.04 82.02 83.51
1.35 1.06 .43 .19 .16 .24
(h)
Hydrolysis
G-G test Mean SD Time
(%)
6 12 24 30 36 48
No NaHSO 3 added
(h) ... 23.82 72.97 77.78 87.05 90.81
_.o
1.84 3.01 .88 2.69 2.14
With .4 M NaHSO 3
(%)
6 12 21 24 30 36 48
4.38 8.79 14.85 24.77 28.74 27.25 31.66
9.06 26.70 41.58 57.29 65.11 70.24 77.40
a12.8 g anhydrous lactose/100 ml. a18.14 g anhydrous lactose/100 ml.
without amino acid present) before analysis. Browning was more obvious in the presence of amino acids, particularly at alkaline pH values. Therefore, whey syrups turned brown rapidly when heated if they were overneutralized. Browning was much darker and more complex during whey hydrolysis than during aqueous lactose hydrolysis. Activated carbon did not remove the color developed through sugaramino acid reactions during whey hydrolysis. Procedures similar to those that proved effective on aqueous lactose solutions (Table 5) were ineffective in clarifying hydrolyzed whey solutions; they remained discolored and were salty besides having a pronounced off-flavor (bitter). Sodium bisulfite has been used in many food products to prevent Maillard browning.
The present work found sodium bisulfite also effective in inhibiting browning of hydrolyzed whey. Figure 3 shows the degree of color inhibition by sodium bisulfite. Color development was prevented completely by 1.0 M sodium bisulfite during the hydrolysis of 20.02% whey lactose in 2N H2SO4. For an acceptable final color most of the browning reaction can be inhibited by .4 M sodium bisulfite. Although sodium bisulfite reduced Maillard browning during whey hydrolysis, it contributes other problems, such as developing a strong flavor, slowing hydrolysis, and interfering with analytical methods. Figures 4 and 5 show the effects of sodium bisulfite on both the L test and G-G test. In the presence of .01
I
.61
E zz
I.O
o
tO Z
~ .4 0r.o < .2
LACTOSE WITH
J
WITH I M NaHSO~
O
CONCENTRATION
{mg/ml}
FIG. 4. Effect of NaHSO~ on lactose test.
Journal of Dairy Science Vol. 60, No. 1
I.O
2.O
3 'O
4 iO - -
CONCENTRATION I rn9 / m l }
FIG. 5, Effect of NaHSO 3 on glucose-galactose test.
ACID HYDROLYSIS OF LACTOSE IN WHEY
M sodium bisulfite, color development decreased during the L test, but increased in the G-G test. Table 5 shows that the hydrolysis of whey is retarded in the presence of sodium bisulfite. The possibility of developing acidhydrolyzed syrups from whey with acceptable flavor and color, therefore, seems unlikely although the feasibility of producing such syrups from aqueous lactose solutions has been demonstrated. REFERENCES
Cameron, A. T. 1947. The taste sense and the relative sweetness of sugars and other sweet substances. Sugar Research Foundation Rept. No. 9, 1 to 72, New York.
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2 Coughlin, J. R., and T. A. Nickerson. 1975. Acid-catalyzed hydrolysis of lactose in whey and aqueous solutions. J. of Dairy Sci. 58:169. 3 Jenness, R., and J. Koops. 1962. Preparation and properties of a salt solution which simulates milk ultrafiltrate. Neth. Milk & Dairy J. 16:153. 4 Ney, K. H., and I. P. G. Wirotoma. 1970. Investigation of lactose hydrolysis. Zeitschrift Vfir Lebensmittelunter-suchung und Forsch. 143 (2), 93; FS&T Abstr. 2 (11), 11 p. 1591 (1970). 5 Nickerson, T. A., I. F. Vujicic, and A. Y. Lin. 1976. Colorimetric estimation of lactose and its hydrolytic products. J. Dairy Sci. 59:386. 6 Ramsdeil, G. A., and B. H. Webb. 1975. The acid hydrolysis of lactose and the preparation of hydrolyzed lactose sirup. J. Dairy Sci. 28:677. 7 Vujicic, I. F., A. Y. Lin, and T. A. Nickerson. 1976. Changes during acid hydrolysis of lactose. J. of Dairy Sci. 60:29.
Journal of Dairy Science Vol. 60, No. 1