Meat Science, Vol. 42, No. 1, 3748, 1996 Copyright 0 1995 Elsevier Science Ltd

Printed-in Great Britain. All rights reserved 0309-1740/96/$9.50 + .OO

0309-1740(95)00017-8

Potassium Chloride, Potassium Lactate and Glycine as Sodium Chloride Substitutes in Fermented Sausages and in

Dry-cured Pork Loin

P. Gou,* L. Guerrero, J. Gelabert & J. Arnau

Institut de Recerca i Tecnologia Agroalimentiries, Centre de Tecnologia de la Carn, Granja Camps i Armet s/n, 17121, Monells, Spain

(Received 19 January 1995; revised version received 10 April 1995; accepted 21 April 1995)

ABSTRACT

Salt is essential in the elaboration of dry meat products, contributing to their tex- ture andjlavour development. The effect brought about by substituting NaCl with KC1 (MO%), potassium lactate (&lOO%) andglycine (C&100%) on the texture, flavour and colour characteristics of fermented sausages and dry-cured pork loins was evaluated. Texture profile analysis and a sensory analysis were performed. Importantjavour defects were detected with substitutions above 40% for the three substituents in both products, and with substitutions above 30% for glycine in dry- cured loin. A loss of cohesiveness was detected by the sensory analysis in fermented sausages at substitution levels higher than 30% with potassium lactate (K-lactate) and higher than 50% with glycine. Although the instrumental analysis detected texture changes in dry-cured loin, the sensory analysis did not detect any substitution effect on texture.

INTRODUCTION

There is a positive relationship between high sodium intake and the incidence of hypertension (Frost et al., 1991; Law et al., 1991a,b). However, NaCl is an essential ingredient in processed meat products, contributing to the water-holding capacity, colour, fat binding and flavour. Moreover, salt decreases water activity (uw), and this significantly affects the shelf-life (Wirth, 1989).

The ways in which the sodium content of meat products can be reduced were reviewed by Terre11 (1983). The approaches generally involved: (1) reducing the addition of NaCl; (2) substitution of NaCl with other ingredients; and/or (3) alteration of processing techniques.

The majority of studies on NaCl reduction have been performed on cooked meat products, such as frankfurters or cooked ham (Sofos, 1983a,6; Whiting, 1984). In order to improve the results, some authors propose modifications in the process, such as tumbling (Frye et al., 1986; Lin et al., 1991), emulsion coating (Thiel et al., 1986), or adding potassium sorbate (Sofos, 1985, 1986) or certain

*Author to whom correspondence should be addressed.

37

38 P. Gou et al.

protein hydrolysates on a collagen basis (Hofmann and Maggrander, 1989), which compensate for the loss of binding produced by a reduction of NaCl. In Spain cooked ham without added NaCl is already commercialized.

In dry-fermented sausages and dry-cured ham reduction of NaCl requires modifications to the curing process, such as a reduction of the ripening tempera- ture (Baldini et al., 1983; Wirth, 1989). Substitution of NaCl with KC1 can be undertaken without functional loss, but metallic and astringent tastes could limit its use (Terre11 and Olson, 1981; Pasin et al., 1989). KC1 and LiCl have been studied as NaCl substitutes in country-style hams (Hand et al., 1982; Keeton, 1984). However, LiCl is toxic and KC1 substitution at a level of 33.3% presented a slight bitter taste, being unacceptable at a level of 50% and above. Annemiche et al. (1990) noted the preservative qualities of the lactate ion, which suggests that it may be possible to use potassium lactate (K-lactate) as a substitute for salt. However, as far as potassium is concerned, the abnormal taste encountered could limit its use. Recently, Askar et al. (1994) stated that the substitution of NaCl with KC1 and K-lactate at 40% or below does not affect the sensory character- istics in pasterma.

Glycine, which reduces the a, (Chen and Karmas, 1980), may improve micro- biological stability and could be used as a NaCl substitute, although it is necessary to determine its effect on the sensory characteristics of meat products before ascertaining its feasibility as a substitute.

The aim of this study was to evaluate the effect of substitution of NaCl with KCl, K-lactate and glycine on the texture, flavour and colour characteristics of fermented sausages and dry-cured pieces of loin, which could be considered as models of dry-cured ham.

MATERIALS AND METHODS

In both products, fermented sausage and dry-cured loin, the treatments for each substituent corresponded to molar substitutions of NaCl progressively by lo%, from 0 to 60% with KC1 and from 0 to 100% with K-lactate and glycine. Eight gilt carcasses with a pH (measured on the Semimembranosus muscle at 45 min post mortem) of above 6 were selected and frozen at -20°C.

Fermented sausage preparation

Frozen meat from the shoulders and bellies was thawed at 4°C and ground in a meat grinder by passing through a 6 mm plate. Three batches were prepared, one for each substituent. A shoulder:belly proportion of 70:30 was mixed with the common additives (g per kg of meat): dextrose 10, sodium pyrophosphate (solution l%, pH= 5.0) 2, black pepper 2, sodium ascorbate 0.5, starter (Amerex, SAGA L) 0.3, KN03 0.2 and NaN02 0.1. The mixture was divided into portions of 2 kg each. Each portion was assigned to a substitution treatment and the correct amount of salt and/or salt substitute was added. Twenty-six grams per kilogram were added to the control. The stuffed sausages (diameter 40 mm) were hung in a chamber for fermentation at a temperature of 18-20°C and a relative humidity of 9&95% until pH decreased to 5.0, or for 3 days maximum. Drying was carried out at 12-14°C with a relative humidity of

Sodium chloride substitutes in fermented sausages 39

7&80% for 15 days. The pH was measured at different intervals throughout the fermentation, in order to determine the end of fermentation (pH < 5.0), and at the end of the whole process.

Dry-cured loin preparation

Fifteen frozen loins were thawed at 4°C. Three loins were cut into seven pieces each, which were then put aside for the six substitutions with KC1 and a control. The rest of the loins, six for K-lactate substitutions and six for glycine substitu- tions, were divided into six pieces each, which were then put aside for a control and five substitutions (from 10 to 50% or from 60 to 100%). The three pieces with the same substitution were sampled from different zones of the loin: the head, centre and back. Salting was carried out using a surface massage. Thirty- five grams of NaCl and 0.5 g of KNOs per kg of loin were added to the control. After 8 days, the loins were stuffed into regenerated collagen casings and kept for 5 days at a temperature of 2-3°C. They were then stored for 2 days in a drying room at 9-l 1°C at a relative humidity of 8&90%, and then for 21 days at 1 l-13°C at a relative humidity of 70-80%.

Instrumental texture analysis

A Texture Analyser (model TA.XT2 of Stable Micro Systems Ltd.) was used to carry out a texture profile analysis (TPA: Bourne, 1978), on three sausages and three pieces of loin for each substitution. The samples (1 x 1 x 1 cm) were com- pressed to 60% rather than 80% in order to avoid overloading of the load cell. Crosshead speed was 5 mm/seg. The following parameters were calculated: hardness (kg), springiness (%), cohesiveness (%), gumminess (kg) and chewiness (kg). The mean of six replicates was recorded for each sample.

Sensory analysis

Five selected and trained assessors (ASTM, 1981) undertook the sensory analysis on 2 mm slices. The generation and selection of the descriptors was performed by an open discussion in three previous sessions.

Evaluation of the substitution effect on fermented sausages was undertaken in six sessions with KC1 and seven sessions with potassium lactate and glycine. Three sausages, each with a different level of substitution, were selected using an incomplete block design and compared to a control in each session. The descrip- tors were: fat fluidity, hardness, cohesiveness, acid taste, saltiness, piquantness, off-flavour (bitterness with KCl, potassium lactate flavour with K-lactate and sweetness with glycine), colour intensity and colour uniformity.

To study the substitution effect in dry-cured loin, three pieces, each with a different substitution level, were evaluated in each session using an incomplete block design. The descriptors were: pastiness, hardness, crumbliness, saltiness, off-flavour (bitterness with KCl, potassium lactate flavour with K-lactate and sweetness with glycine), colour intensity and colour uniformity.

A non-structured lo-point scoring scale (Amerine et al., 1965) was used, where 0 means absence of off-flavours, saltiness, acid taste and piquantness or very low intensity of other descriptors and 10 means very high intensity.

40 P. Gou et al.

Statistical analysis

The data were analysed for each substitute by variance analysis, using the GLM procedure of SAS (SAS, 1985). In fermented sausages the model for instrumental texture parameters included the substitution level as a fixed effect. The model for sensory characteristics included the substitution level and the combination assessor by session as fixed effects and the sausage as a random effect. The substitution level effect was tested using the sausage effect as the error term.

In dry-cured loin, the model for instrumental texture parameters included the substitution level and the loin as fixed effects. The model for sensory character- istics included the substitution level, the loin and the combination assessor by session as fixed effects and the piece as a random effect. The substitution level effect was tested using the piece effect as the error term.

Differences between each of the substitution levels and the control were tested and tabulated.

RESULTS

Fermented sausages

Substitution with KC1 No substitution level showed differences in pH reduction during fermentation or at the end of the process with respect to the control.

TABLE 1 Differences in Instrumental Texture Parameters and in Sensory Characteristics Between

Each of the KC1 Substitutions (l&60%) and the Control (C) in Fermented Sausages

10%-C 20%-C 30%-C 40%-C 50%-C 60%-C

Instrumental texture analysis (TPA) Springiness (%) 0.87 Hardness (kg) -2.93 Cohesiveness (%) -0.59 Gumminess (kg) -1.48 Chewiness (kg) -0.75

Sensory analysis” Fat fluidity 0.03 Hardness -0.66 Acid taste -0.18 Saltiness 0.23 Piquantness -0.25 Bitterness 0.10 Colour intensity -0.45 Colour uniformity -0.64

2.31 2.69 1.81 -1.09 -0.62 -0.84 0.22 1.01 -0.72

-0.46 -0.19 -0.51 -0.12 0.06 -0.19

-0.31 0.23 0.09 -0.34 -0.15 0.75* -0.02 -0.44 0.08 0.01 -0.43 -0.31

-0.13 -0.41 0.53 0.36 1.11* 1.37*

-0.59 -0.09 0.04 -0.55 -0.08 0.47

-2.59 -0.82 -0.29 -1.16 -2.45 0.85 -0.41 -0.47 -0.36 -0.29

0.35 -0.07 0.61* 0.71*

-0.43 -0.49 -0.90* -1.02* -0.53 -0.55 3.28* 3.84* 0.03 -0.14 0.24 0.18

*Significant difference (P < 0.05). OOriginal scales (before the control value was subtracted): 0 means absence of acid taste, saltiness, piquantness and bitterness or very low intensity of other parameters and 10 means very high intensity.

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.

Sodium chloride substitutes in fermented sausages 43

The TPA analysis did not show any difference in texture and the sensory analysis only detected a slight increase in hardness at 40, 50 and 60% levels of substitution (Table 1).

The bitter taste was the most important defect. It was detected at a 30% level of substitution, though the assessors did not consider its intensity important until the 50% level was reached. Below this level the other taste characteristics and the colour parameters were unaffected by the substitution. Saltiness decreased at 50 and 60% levels of substitution.

Substitution with K-lactate The drop in pH to a level below 5 took place within 22 hr in the control sample. In the samples with a substitution of 30 or 40% the process took about 2 days, and with greater substitutions the pH did not drop below 5 even after 3 days.

The TPA analysis showed a decrease in chewiness, gumminess and hardness from a 10% substitution level; this reduction being more important from a 40% substitution level (Table 2). Also, from this level some other reductions were found: in springiness by TPA analysis, and in fat fluidity and in hardness by sensory analysis. The loss in cohesiveness was first detected by sensory analysis (from a 40% level) and afterwards by instrumental analysis (from 60% level).

The acid taste diminished from a 30% substitution level and saltiness from a 20% substitution level. A slight K-lactate flavour from 30% was detected, though this defect became much more important from 50%. A decrease in colour uni- formity from 60% and in colour intensity from 70% were also detected.

Substitution with glycine The drop in pH below 5.0 during fermentation took more than a day only at substitutions of 90 and loo%, and there was no effect of substitution on pH at the end of the process.

Low substitution levels affected the texture of the product (Table 3). The TPA results showed reductions in springiness from a 10% substitution level, in cohesiveness and chewiness from 20%, and in gumminess from 50%. The sensory analysis detected a reduction of fat fluidity and hardness from 30% and a loss of cohesiveness from 50%.

The sensory analysis detected slight reductions in acid taste and salty taste from a 20% substitution level and in piquantness from 30%, and a slight increase in sweetness from 30%. These effects became more noticeable from 50%. Colour was not affected by glycine substitutions.

Dry-cured loins

Substitution with KC1 Substitution of NaCl with KC1 in dry-cured loin did not affect the texture or the colour, except for a slight reduction in springiness in substitutions of 50 and 60% (Table 4). A noticeable bitter taste was detected in substitutions of 50 and 60%.

Substitution with K-lactate The TPA in dry-cured loins showed a reduction in springiness and cohesiveness when substitutions of NaCl with K-lactate at levels of 20% and above were used (Table 5). However, no substitution effect on the texture was detected by sensory

44 P. Gou et al.

TABLE 4 Differences in Instrumental Texture Parameters and in Sensory Characteristics Between Each of the KC1 Substitutions (1060%) and the Control (C) in Dry-cured Loin

10%-C 20%-C 30%-C 40%-C 50%-C 60%-C

Instrumental texture analysis (TPA) Springiness (%) 2.31 Hardness (kg) -2.59 Cohesiveness (%) -0.56 Gumminess (kg) -1.41 Chewiness (kg) -0.66

Sensory analysis* Pastiness -0.18 Hardness -0.05 Crumbliness 0.13 Saltiness 0.51 Bitterness -0.19 Colour intensity 0.84 Colour uniformity 0.19

-0.45 -1.77 0.39 -3.11* -4.06* -2.08 -0.85 0.19 0.37 0.07 0.28 -2.81 0.33 -1.23 -1.98

-1.11 -0.70 0.25 0.10 -0.14 -0.66 -0.44 0.23 -0.11 -0.28

0.56 0.05 0.54 0.23 0.38 -1.16 0.12 -0.91 -0.56 0.19 0.72 -0.27 -0.20 -0.17 -0.22

-0.18 0.51 -0.35 0.40 -0.31 0.34 1.52 1.35 2.28* 3.81* 1.08 1.20 1.20 0.20 0.73 0.01 0.52 -0.04 0.33 -0.17

*Significant difference (P < 0.05). “Original scales (before the control value was subtracted): 0 means absence of saltiness and bitterness or very low intensity of other parameters and 10 means very high intensity.

analysis. A slight decrease in saltiness was noted from a 20% substitution level. From a 50% substitution level, a K-lactate flavour was detected. Colour was not affected by any of the substitutions.

Substitution with glycine The TPA analysis showed a substitution effect on the texture of dry-cured loin when glycine was used (Table 6). At a 40% substitution level and above a sig- nificant reduction in springiness was detected. Other instrumental texture para- meters were seen to be affected in greater substitutions: cohesiveness from levels of 50% and chewiness from 60%. However, no effect on the texture was detected by sensory analysis. Substitutions below 40% reduced saltiness only slightly. From the 40% substitution level a noticeable reduction in saltiness was noted, as well as the presence of an undesirable sweet taste. Colour was not affected by the substitutions.

DISCUSSION

Fermented sausages

Bitterness is the only limiting factor in KC1 substitutions. Although a bitter taste is detected at substitutions levels as low as 30%, it would appear acceptable to substitute 40% of the NaCl content with KCl. Askar et al. (1994) also regarded the 40% level as the maximum at which the bitterness could be acceptable in pasterma.

TA

BL

E

5 D

iffe

renc

es

in I

nstr

umen

tal

Tex

ture

Pa

ram

eter

s an

d in

Sen

sory

C

hara

cter

istic

s B

etw

een

Eac

h of

the

K

-lac

tate

Su

bstit

utio

ns

(l&

100%

) an

d th

e C

ontr

ol

(C)

in D

ry-c

ured

L

oin

10%

-C

20%

-C

30%

-C

40%

-C

50%

-C

60%

-C

70%

-C

80%

-C

90%

-C

100%

-C

3 B

Inst

rum

enta

l te

xtur

e an

alys

is

(TPA

) s

Spri

ngin

ess

(%)

-3.6

5 -8

.85*

-7

.02*

-7

.69*

-1

1.96

* -1

4.91

* -1

0.67

* -1

7.16

* -1

6.80

* -1

5.33

* g

Har

dnes

s (k

g)

-0.2

9 -2

.33

0.61

-3

.51

-0.3

5 -1

.34

-1.4

1 0.

64

1.28

&

- -1

.70

Coh

esiv

enes

s (%

) 2.

-2

.83

-5.1

1*

-7.6

0*

-6.9

6*

-6.4

5*

-6.1

0*

-8.1

2*

-8.9

1*

-9.9

7*

-10.

15*

&

Gum

min

ess

(kg)

-0

.37

-1.8

5 -0

.32

-2.4

0 -0

.64

-1.3

5 -1

.27

-0.6

2 -0

.14

-1.7

8 B

C

hew

ines

s (k

g)

-0.3

5 -1

.50

-0.4

6 -1

.78

-0.7

8 -1

.46

-1.0

6 -1

.22

-0.7

0 -1

.64

F

Sens

ory

anal

ysis

” z.

z

Past

ines

s -0

.29

0.53

0.

02

0.00

0.

53

0.82

0.

98

-0.1

8 0.

52

0.85

5

Har

dnes

s -0

.11

-0.0

5 0.

35

-0.0

4 0.

05

0.32

-0

.29

-0.5

0 -0

.48

0.10

9’

C

rum

blin

ess

0.30

-0

.49

-0.5

1 -0

.19

-0.5

5 -0

.47

0.20

-0

.42

0.53

0.

01

&

Salti

ness

-0

.11

-0.7

7*

-0.5

0*

-0.9

1*

-1.2

7*

-1.6

7*

-2.2

9*

-2.4

2*

-3.1

7*

-3.4

3*

3 K

-lac

tate

fl

avou

r 0.

08

0.34

-0

.05

0.78

3.

72*

3.40

* 3.

58*

5.23

* 6.

15*

6.82

* 3

Col

our

inte

nsity

0.

10

-0.8

2 -0

.51

-0.0

8 0.

80

-0.5

2 -1

.24

-1.0

4 -0

.58

-1.7

6 g

Col

our

unif

orm

ity

0.17

-0

.81

0.09

0.

32

-0.0

8 -0

.37

-0.3

3 -1

.07

-0.3

4 -1

.69

2 E

*Sig

nifi

cant

di

ffer

ence

(P

<

0.

05).

&

“Ori

gina

l sc

ales

(b

efor

e th

e co

ntro

l va

lue

was

su

btra

cted

):

0 m

eans

ab

senc

e of

sal

tines

s an

d K

-lac

tate

fl

avou

r or

ve

ry

low

in

tens

ity

of o

ther

6

para

met

ers

and

10 m

eans

ve

ry

high

in

tens

ity.

TA

BL

E

6 D

iffe

renc

es

in I

nstr

umen

tal

Tex

ture

Pa

ram

eter

s an

d in

Sen

sory

C

hara

cter

istic

s B

etw

een

Eac

h of

the

G

lyci

ne

Subs

titut

ions

(l

Crl

OO

%)

and

the

Con

trol

(C

) in

Dry

-cur

ed

Loi

n

10%

-C

20%

-C

30%

-C

40%

-C

50%

-C

60%

-C

70%

-C

80%

-C

90%

-C

100%

-C

Inst

rum

enta

l te

xtur

e an

alys

is

(TPA

) Sp

ring

ines

s (%

) 0.

81

-7.7

2 1.

12

-9.6

4*

-9.6

3*

-16.

10*

-13.

74*

-18.

68*

-16.

12*

-19.

81*

Har

dnes

s (k

g)

0.04

-0

.11

-0.2

0 -0

.37

0.01

0.

03

-0.1

8 -0

.64

-0.2

4 -0

.98

Coh

esiv

enes

s (%

) -0

.60

-4.7

8 -3

.46

-4.2

3 -8

.24*

-1

1.80

* -1

1.22

* -1

3.19

* -1

4.64

* -1

4.01

*

Gum

min

ess

(kg)

-0

.02

-0.3

1 -0

.31

-0.4

8 -0

.47

-0.6

9 -0

.78

-1.0

4 -1

.00

-1.2

2 C

hew

ines

s (k

g)

0.00

-0

.41

-0.1

7 -0

.61

-0.5

8 -0

.87*

-0

.86*

-1

.11*

-1

.03*

-1

.21*

.?

Sens

ory

anal

ysis

” Q

Past

ines

s B

0.

17

0.03

0.

20

0.46

0.

12

0.17

0.

52

0.37

0.

60

0.60

H

ardn

ess

9 0.

27

0.41

0.

46

-0.5

0 0.

49

-0.7

4 -1

.29

-1.3

4 -0

.89

-1.4

9 C

rum

blin

ess

-0.8

3 -1

.04

-1.0

2 -0

.31

-1.3

3 -0

.29

0.98

0.

3.0

-0.1

8 0.

05

F

Salti

ness

-0

.41

-0.9

3*

-1.2

1*

-1.8

5*

-2.0

4*

-2.5

5*

-2.6

8*

-3.5

1*

-3.6

8*

-4.2

2*

Swee

tnes

s 0.

89

1.92

1.

79

3.55

* 3.

06*

3.67

* 5.

29*

5.82

* 6.

66*

7.81

* C

olou

r in

tens

ity

0.36

-0

.04

0.86

0.

91

0.87

-0

.09

-0.7

0 -0

.07

0.39

-0

.03

Col

our

unif

orm

ity

0.89

1.

38

0.93

1.

03

0.82

-0

.70

-0.6

7 0.

44

0.11

0.

49

*Sig

nifi

cant

di

ffer

ence

(P

<

0.

05).

“O

rigi

nal

scal

es

(bef

ore

the

cont

rol

valu

e w

as

subt

ract

ed):

0

mea

ns

abse

nce

of

salti

ness

an

d sw

eetn

ess

or

very

lo

w

inte

nsity

of

ot

her

para

- m

eter

s an

d 10

mea

ns

very

hi

gh

inte

nsity

.

Sodium chloride substitutes in fermented sausages 47

High K-lactate substitution levels present problems in texture, colour and flavour. Moreover, the delay in the pH drop represents a spoilage risk, which may be compensated in part by the antimicrobial activity of ion lactate (De Koos, 1993).

Consistency is probably the most important texture characteristic in this product because it determines the slicing ability. The drop in pH and the drying process promote gelling and consequently consistency. Moreover, a pH below 5.5 is desirable for the reactions involved in the formation of red colour from the nitrite. At these pH levels the reactions of reduction are more rapid and allow a higher colour stability (Frey, 1993). Therefore, the delay in pH drop in K-lactate substitutions could explain the modifications detected in texture from the 40% substitution level and in colour from the 60% substitution level.

As with KC1 substitutions it seems acceptable, from a flavour point of view, to substitute 40% of NaCl with K-lactate, although at a 30% level a slight K-lactate flavour can be detected. Askar et al. (1994) obtained the same maximum level of substitution with K-lactate in pasterma. However, at this level (40%) it would be necessary to apply some modifications in the process in order to correct the texture.

NaCl substitution with glycine could be acceptable at a 40% level. Above this level, an unacceptable sweet taste and an inconsistent texture appear. In this case the differences in consistency cannot be explained by the drop in pH, because it was similar to the control. It is probable that glycine has less capacity to dissolve proteins than NaCl, and this might explain the loss of consistency.

Dry-cured loins

The main problem in NaCl substitution with any of the three substituents studied is the flavour. None of the substitutes affected the colour. The instrumental tex- ture analysis shows some differences, though they were not detected by sensory analysis.

It would appear that KC1 and K-lactate could substitute 40% of NaCl without any significant defect in flavour. These results agree with those of Keeton (1984), who concluded that a substitution of NaCl for KC1 of one-third is possible without significantly altering the product characteristics in country-style hams. However, he did not use substitution levels of between 33.3% and 66.7%. Sweetness in substitutions with glycine is detected at a low level (40%) so 30% would seem to be the maximum substitution level.

Substitution improvement

The drying process affects NaCl distribution in the product, increasing the con- centration in the inner part of dried sausage (Riidel, 1985) and dry-cured ham (Arnau et al., 1995); and it is probably similar for the distribution of KCl, K-lactate and glycine. High local concentrations of these products increase off-flavour (bitterness, potassium lactate flavour or sweetness), which seems to be the principal impediment to NaCl substitution. Therefore, it would be interesting to study process modifications in order to improve the substitute distributions.

A flavour improvement could also be obtained by mixing substitutes since, in mixtures of substances with different tastes, suppression of each taste tends to be

48 P. Gou et al.

the rule; but whether the total taste intensity is less than the sum of the perceived taste intensities of the components is not easily determined (Shallenberger, 1993). This possibility is being studied by the authors.

ACKNOWLEDGEMENTS

We would like to thank J. Arbonb for his collaboration in the manufacturing process. Part of this study has been supported with funds from the Spanish Ministry of Agriculture, Fishing and Feeding (Project INIA SC93-126).

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