deakin research online - home - drodro.deakin.edu.au/eserv/du:30017402/keast-modificationof...new...

Deakin Research Online Deakin University’s institutional research repository

DDeakin Research Online Research Online This is the author’s final peer reviewed version of the item published as: Keast, Russell 2008-07, Modification of the bitterness of caffeine, Food quality and preference, vol. 19, no. 5, pp. 465-472. Copyright : 2008, Elsevier Ltd

http://www.deakin.edu.au/dro/




Suppression of caffeine bitterness

1

Modification of the bitterness of caffeine

Russell SJ Keast (PhD)

The corresponding author may be contacted at the following address:

Dr. RSJ Keast

Senior Lecturer

School of Exercise and Nutrition Sciences

Deakin University

221 Burwood Highway, Burwood, Victoria 3125 Australia.

Phone: 03 9244 6944 International: +61 3 9244 6944

Fax: 03 9244 6017 International: +61 3 9244 6017

e-mail: [email protected]

Keywords

Bitter, taste, smell, inhibition, suppression, flavor modification


2

ABSTRACT

Caffeine is the worlds most consumed pyschoactive chemical and as such is a

valuable commodity to the food and beverage industry. Caffeine also activates the

bitter taste system causing a potential problem for manufacturers wanting to develop

products containing caffeine. In the present study both oral peripheral and central

cognitive strategies were used in an attempt to suppress the bitterness of caffeine.

Subjects (n=33) assessed the influence of sodium gluconate (100 mM), zinc lactate (5

mM), sucrose (125, 250 mM), milk (0, 2, 4% milkfat), and aromas (coffee, chocolate,

mocha) on the bitterness of caffeine (1.5, 3, 4.5 mM). The oral peripheral strategies

proved most effective at suppressing the bitterness of caffeine: zinc lactate (-71%,

p<0.05), non-fat milk (-49%, p<0.05), and sodium gluconate (-31%). Central

cognitive strategies were partially effective: 250 mM sucrose (-47%, p<0.05) and

mocha aroma (-10%) decreased bitterness, while chocolate (+32%) and coffee

(+17%) aromas increased perceived bitterness. Overall, zinc lactate was the most

effective bitterness inhibitor, however the utility of zinc in foods is negated by its

ability to inhibit sweetness.


3

INTRODUCTION

Caffeine is the most commonly ingested psychoactive drug in the world,

naturally occurring in coffee, cocoa (chocolate), and black teas, and as an additive in

soft- and energy-drinks. The stimulant effect of foods that contain caffeine, such as

the coffee bush berry, has been known for hundreds of years (Flament, 2002). The

stimulant effect was due to caffeine and related methyl xanthines such as theophylline

and theobromine whose physiologic mode of action is antagonism of the adenosine

A1 and A2 receptors in the brain (Rainnie, Grunze, McCarley, & Greene, 1994).

As the site of action of caffeine is similar to the site of action of addictive

drugs, it is not surprising that caffeine, at levels in common beverages, has been

proposed to be an addictive drug (Holtzman, 1990). The dose of caffeine required to

modify behaviour in humans is low (~50 mg) (Nehlig, 1999), similar to the dose

delivered in 500 ml common cola soft-drinks (~53-65 mg, 0.55-0.67 mM).

Behavioural studies have shown that the consumption of caffeine promotes a

physiologic and psychologic dependence that is reinforced with repeat consumption

(Garrett & Griffiths, 1998; Hughes, Oliveto, Bickel, & Higgins, 1993; Schuh &

Griffiths, 1997). The common method of repeat caffeine consumption is via

caffeinated foods such as coffee, tea, cocoa, and soft drinks. Caffeine may promote

liking and consumption of these foods via the development of flavor preferences;

where individuals associate (unconsciously) a food/flavor with its post-ingestive

consequences. The mode of action of caffeine in developing flavour preference is not

immediate (Yeomans et al., 2000) as, for example, we experience with a sucrose

solution (sweet and appetitive) but the positive influences occur post-consumption

with increased vigilance and attention, enhanced mood and arousal as well as

enhanced motor activity.


4

Caffeine may elicit bitterness depending on the concentration (Keast & Roper,

2007) and this can be a problem for food and beverage manufacturers. Solving this

problem is complicated by the observation that the human bitter taste system is

complex. It is subserved by approximately two dozen putative G-protein coupled

receptors, the TAS2Rs (Adler et al., 2000; Chandrashekar et al., 2000), and several

post-receptor transduction mechanisms (Huang et al., 1999; Kinnamon & Margolskee,

1996; Rossler, Kroner, Freitag, Noe, & Breer, 1998; Spielman, Huque, Whitney, &

Brand, 1992; Wong, Gannon, & Margolskee, 1996). Moreover, caffeine may modify

flavour at the peripheral level via interference with taste transduction (Peri et al.,

2000; Rosenzweig, Yan, Dasso, & Spielman, 1999).

In general, there are three approaches to suppressing bitterness: physio-

chemical interactions in a food or beverage matrix, oral peripheral physiological

interactions with receptor cells (e.g., via receptor inhibitors), and central cognitive

mixture suppression (e.g., via taste-taste and taste-aroma interactions).

Physio-chemical interactions can change flavour intensity or even generate

new flavours. They occur in a simple aqueous solution: weak attractive forces, such

as hydrogen or hydrophobic bonding, will result in altered structures; precipitation of

the compounds will render them weaker or tasteless. The chemical composition of a

food matrix will influence perceived flavour, changing from an aqueous to emulsion

system (oil-in-water) decreased bitterness (Metcalf & Vicker, 2002).

Sodium and zinc salts inhibit the bitterness of certain compounds whether or

not the salts elicit a taste, indicating that the inhibition is peripheral rather than based

on perceptual interactions (Bartoshuk & Seibyl, 1982; Breslin & Beauchamp, 1995a;

Keast & Breslin, 2002, 2005; Keast, 2003; Kroeze & Bartoshuk, 1985). As well as

lipid having a physio-chemical influence on bitterness, the components of fats, fatty


5

acids, may also modify bitterness via interactions in the oral periphery (Koriyama,

Wongso, Wananabe, & Abe, 2002).

Central cognitive effects can occur when different qualities of taste stimuli are

mixed together and the perceived intensity of one or more of the components is

diminished by the perception of the others. This is labeled mixture suppression

(Pangborn, 1960) and is caused by cognitive interactions among taste qualities. As

one example, mixture suppression occurs when you add sugar to coffee, both the

sweetness of the sugar and bitterness of the coffee are reduced. Also, the combination

of taste and aroma may influence the intensity of both the taste and aroma (Frank &

Byram, 1988; Frank, Ducheny, & Mize, 1989). The primary requirement for an

aroma influencing the perceived intensity of a taste is the congruency of the aroma-

taste pair. For example, strawberry aroma increases perceived sweetness (we

associate strawberry odour with sweetness) whereas peanut butter aroma does not

increase perceived sweetness (we do not associate peanut butter aroma with

sweetness).

The aim of this study was to investigate both central cognitive and oral

peripheral factors that may modify the bitterness of caffeine.


6

MATERIALS AND METHODS

SUBJECTS

Subjects (n=33, 23±4 years old, 28 female) between the ages of 18 and 38

were University students in Melbourne, Australia. All subjects were volunteers and

agreed to participate and provided informed consent on an approved Institutional

Review Board form. The subjects were asked to refrain from eating, drinking or

chewing gum for one hour prior to testing. Not all subjects participated in all

experiments: Experiment 1, n=30; Experiment 2, n=31; Experiment 3, n=33;

Experiment 4, n=32.

SUBJECT TRAINING

Subjects were initially trained in the use of the general Labeled Magnitude

Scale (gLMS) following the published standard procedures (Green et al., 1996; Green,

Shaffer, & Gilmore, 1993) except the top of the scale was described as the strongest

imaginable sensation of any kind (Bartoshuk, 2000). The gLMS is a psychophysical

tool that requires subjects to rate perceived intensity along a vertical axis lined with

adjectives: barely detectable = 1.5, weak = 6, moderate = 17, strong = 35, very strong

= 52, strongest imaginable = 100; the adjectives are placed semi-logarithmically,

based upon experimentally determined intervals to yield data equivalent to magnitude

estimation (Green et al., 1996; Green et al., 1993). The scale only shows adjectives,

not numbers, to the subjects, but the experimenter calculates numerical data from the

scale. The gLMS was chosen for this study as it provides ratio quality data

(Bartoshuk, 2000). A computerized data-collection program (Compusense 5) was

used in all sessions.


7

Subjects were trained to identify each of the five taste qualities by presenting

them with exemplars. Salty taste was identified as the predominant taste quality from

150 mM NaCl, bitterness as the predominant quality from 0.50 mM quinine-HCl,

sweetness as the predominant quality from 300 mM sucrose, sourness as the

predominant quality from 3 mM citric acid, and umami the predominant quality from

a mixture of 100 mM MSG and 50 mM IMP. In addition, astringency was identified

from 0.5mM tannic acid. To help subjects understand a stimulus could elicit multiple

taste quality, a mixture of 0.50 mM quinine-HCl & 3 mM citric acid (bitter and sour)

and a mixture of 150 mM NaCl & 300 mM sucrose (salty and sweet) were employed

as training stimuli. The gLMS was the scale used during taste quality training. All

subjects were able to correctly identify taste and astringent qualities after training.

STANDARDISATION OF gLMS RATINGS WITH HEAVINESS RATINGS

The gLMS standardization methodology followed previously published

methodology (Delwiche, Buletic, & Breslin, 2001) and is employed to minimise

individual scale use bias. Subjects rated the heaviness of five visually identical

weights (opaque, sand-filled jars at levels 52, 294, 538, 789, and 1028 g). All ratings

were made on the gLMS. Subjects were asked to rate the intensity of heaviness and all

judgments were made within the context of the full range of sensations experienced in

life. All weights were presented twice in blocks of ascending order.

To determine a standardization factor, each subject’s average intensity for

heaviness was divided by the grand mean for heaviness across weight levels and

subjects. Each individual’s intensity ratings were multiplied by his or her personal

standardization factor for scale-use bias. The data collected were normally distributed

therefore did not require log transformation.


8

STIMULI

Caffeine, sodium gluconate, zinc lactate, and sucrose were purchased from

Sigma Chemical (St. Louis, MO) and were the highest purity available. Zinc and

sodium were selected as they are both known bitterness inhibitors and the

concentrations were selected to ensure tastes from the salts did not overly interfere

with judgements of bitterness (Keast & Breslin, 2005; Keast, Canty, & Breslin,

2004a; Keast, 2003). Milk of varying fat concentrations was prepared according to

manufacturers instructions (Anchor powdered milk range). Coffee and chocolate

aromas were supplied by IFF and prepared according to manufacturers instructions.

Caffeine concentrations were chosen as they are in the range commonly occurring in

foods and beverages and the chosen concentrations could be perceptually

distinguished in preliminary testing.

All solutions were prepared with deionized (di) filtered water and were stored

in glass bottles at 4°-8°C and were brought to room temperature prior to testing.

Filtered di water was used as a control stimulus in experiments 1 and 2 and the rinsing

agent in all experiments.

STATISTICAL ANALYSIS

Numerical results are expressed as arithmetic means ± standard error.

Statistical analysis of results was determined with repeated measures analysis of

variance (ANOVA) with Bonferroni correction using SPSS version 14. Statistical

analyses of bitterness intensity ratings are included in results presented from

experiment one, three and four, and bitterness and sweetness intensity ratings are


9

presented for experiment two. Ratings of the other qualities were collected to

minimize halo dumping effects (Clark & Lawless, 1994). These ratings were

generally not statistically different across conditions, low in magnitude (less than

weak on the gLMS), and not relevant to the objectives of this research project. P

values <0.05 were considered statistically significant. Significance levels for pairwise

post-hoc tests were determined after applying Bonferroni corrections. The post-hoc

significance level for experiments 1, 2 and 3 were: 0.05/18= p<0.0027; and

experiment 4, 0.05/30= p<0.0016. Calculation of percent bitterness inhibition was

calculated using the formula:

(MB / OB) * 100 = Y % Y – (-100) = Z %

where MB= mean modified bitterness pooled across caffeine concentrations, OB =

original bitterness pooled across caffeine concentrations, Y = percent original

bitterness remaining, Z= percent bitterness inhibition


10

METHODS

Experiment 1: The effect of sodium and zinc ions on bitterness of caffeine

Water (di), three caffeine concentrations (1.5, 3, 4.5 mM), and 100 mM Na-gluconate

(or 5 mM Zn-lactate) were tasted individually and in combination to yield a total of 8

solutions per-session per-subject (full factorial design). There were two different

trays (one for each salt), and each tray was tasted on at least three separate occasions,

resulting in a total of 6 sessions on 6 separate days. The testing protocol was as

follows: Solutions (10 ml) were presented in 30 ml plastic medicine cups (McFarlane

Medical and Scientific, Melbourne) on numbered trays. The solutions were

presented in random order across subjects. Subjects rinsed with di water at least four

times over a 2-minute period prior to testing. The subjects were instructed to pour the

whole sample in their mouth while wearing nose-clips, hold it in their mouth for a few

seconds, and rate the solution for sour, sweet, bitter, salty, umami, and astringent

perceptions prior to expectorating. All subjects rinsed with di water 4 times during

the interstimulus interval of 2min. The gLMS was used as the rating method.

Experiment 2: The effect sweetness on bitterness of caffeine

Water (di), three caffeine concentrations (1.5, 3, 4.5 mM), and two sucrose

concentrations (125 and 250 mM) were tasted individually and in combination to

yield a total of 12 solutions per-session per-subject (full factorial design). Each tray

was presented on three separate occasions on three separate days. The testing

protocol was the same as experiment 1.

Experiment 3: The effect of milk fat on bitterness of caffeine


11

Four caffeine concentrations (0, 1.5, 3, 4.5 mM), and three powdered bovine milks

(0%, 2%, 4% milkfat) were tasted individually and in combination to yield a total of

12 solutions per-session per-subject (full factorial design). Each tray was presented

on three separate occasions on three separate days. The testing protocol was the same

as experiment 1.

Experiment 4: The effect of congruent aroma on bitterness of caffeine

Chocolate and coffee aromas were assessed for bitterness in aqueous solutions with

subjects wearing noseclips. There was no perceivable bitterness elicited by the

solutions. Three caffeine concentrations (1.5, 3, 4.5 mM) in 0% milkfat bovine milk

with four aromas (blank, coffee, chocolate, mocha (½ coffee. ½ chocolate)) were

tasted individually and in combination to yield a total of 12 solutions per-session per-

subject (full factorial design). Each tray was presented on three separate occasions on

three separate days. The testing protocol was the same as experiment 1 except

subjects did not wear noseclips to allow for the influence of aroma on bitterness

perception.


12

RESULTS

Experiment 1: The effect of sodium and zinc ions on bitterness of caffeine

Results from a 3 x 4 (salt v caffeine) two-way repeated measures ANOVA

with bitterness as the dependent variable revealed there was a significant main effect

of salt [F(2,58) = 12, p<0.0005] and a significant main effect of caffeine [F(3,87) =

40.8, p<0.0005]. Both Na-gluconate and Zn-lactate significantly inhibited bitterness

(pooled across caffeine concentrations) (p<0.05), and there were significant

differences in bitterness across all caffeine concentrations (pooled across salts)

(p<0.0005). There was a significant interaction among the salts and caffeine [F(6,83)

= 30, p<0.0005] indicating differences in bitterness intensity of specific combinations

of caffeine and salts. Post hoc analysis with Bonferroni correction revealed that Na-

gluconate did not inhibit the bitterness at any caffeine concentration (p>0.0027).

However Zn-lactate significantly inhibited the bitterness of 3 & 4.5 mM caffeine

(p<0.0027) (Figure 1).

Experiment 2: The effect of sweetness on bitterness of caffeine

BITTERNESS

Results from a 3 x 4 (sucrose v caffeine) two-way repeated measures ANOVA

with bitterness as the dependent variable revealed there was a significant main effect

of sucrose [F(2,89) = 76, p<0.0005] and a significant main effect of caffeine [F(3,88)

= 59, p<0.0005]. There were significant increases in bitterness as caffeine

concentration increased from 1.5 to 3 to 4.5 mM (pooled across sucrose

concentrations) (p<0.05). There were significant decreases in bitterness as sucrose

concentration increased from 0 to 125 to 250 mM (pooled across caffeine

concentrations) (p<0.05). There was a significant interaction between sucrose and


13

caffeine [F(6,85) = 17, p<0.0005] indicating differences in bitterness intensity of

specific combinations of caffeine and sucrose. Post-hoc pairwise analysis revealed a

significant increase in bitterness between 1.5 mM caffeine and 3 & 4.5 mM caffeine

(p<0.0027). Sucrose concentrations did not significantly affect bitterness of 1.5 mM

caffeine (p>0.0027). However 250 mM sucrose did significantly inhibit the bitterness

of 3 mM and 4.5 mM caffeine (p<0.0027), yet 125 mM sucrose did not (Figure 2A).

SWEETNESS

Results from a 3 x 4 (sucrose v caffeine) two-way repeated measures ANOVA

with sweetness as the dependent variable revealed there was a significant main effect

of sucrose [F(2,89) = 190, p<0.0005] and a significant main effect of caffeine [F(3,88)

= 34, p<0.0005]. There were significant differences in sweetness between 1.5 mM

caffeine and 3 & 4.5 mM caffeine (p<0.05) but no difference in sweetness between 3

to 4.5 mM caffeine (pooled across sucrose concentrations) (p=1.0). There were

significant increases in sweetness as sucrose concentration increased from 0 to 125 to

250 mM (pooled across caffeine concentrations) (p<0.0005). There was a significant

interaction between sucrose and caffeine [F(6,85) = 18, p<0.0005] indicating

differences in sweetness intensity of specific combinations of caffeine and sucrose.

Post-hoc analysis revealed caffeine concentration had no effect on the sweetness of

125 mM sucrose (p>0.0027). However, 3 & 4.5 mM caffeine significantly inhibited

the sweetness of 250 mM sucrose (p<0.0027). All 250 mM sucrose solutions were

significantly sweeter than the corresponding 125mM sucrose solution (p<0.0027)

(Figure 2B).

Experiment 3: The effect of milk fat on bitterness of caffeine


14

Results from a 3 x 4 (fat v caffeine) two-way repeated measures ANOVA with

bitterness as the dependent variable revealed there was a significant main effect of fat

[F(2,95) = 20, p<0.0005] and a significant main effect of caffeine [F(3,94) = 39,

p<0.0005]. There were significant increases in bitterness as caffeine concentration

increased from 1.5 to 3 to 4.5 mM (pooled across levels of fat) (p<0.05). Also, there

were significant differences in bitterness between 0% fat and 2% & 4% fat (p<0.05)

but no difference in bitterness between 2 and 4% fat (pooled across caffeine

concentrations) (p=0.09)). There was a significant interaction among fat and caffeine

[F(6,91) = 8, p<0.0005] indicating differences in bitterness intensity of specific

combinations of caffeine and fat. Post-hoc analysis revealed no significant

differences in bitterness between 0-2% milkfat, however at all caffeine concentrations

there were significant differences between 0-4% milkfat (p<0.0027) with 0% milk fat

being less bitter at each caffeine concentration (Figure 3).

Experiment 4: The effect of congruent aroma on bitterness of caffeine

Results from a 4 x 3 (aroma v caffeine) two-way repeated measures ANOVA with

bitterness as the dependent variable revealed there was a significant main effect of

aroma [F(3,91) = 10, p<0.0005] and a significant main effect of caffeine [F(2,92) =

28, p<0.0005]. There were significant increases in bitterness as caffeine

concentration increased from 1.5 to 3 to 4.5 mM (pooled across aromas) (p=0.62).

Mocha aroma was significantly less bitter than either chocolate or coffee aroma

(p<0.05) (pooled across caffeine concentrations) but there was no difference in

bitterness between coffee and chocolate aromas (p=0.99). There was no significant

interaction between aromas and caffeine [F(6,88) = 1.6, p=0.15] (Figure 4).


15

DISCUSSION

The organization of the bitter taste system is complex with multiple putative

receptor mechanisms (Chandrashekar, Hoon, Ryba, & Zuker, 2006), including a

specific G protein coupled receptor for caffeine in Drosophilla (Moon, Kottgen, Jiao,

Xu, & Montell, 2006). Our understanding of how caffeine activates bitter taste is far

from complete and this study shows that the bitterness elicited by caffeine can be

partially inhibited using both oral peripheral (Na, Zn) and central cognitive strategies.

The most effective inhibitor of the bitterness of caffeine appears to be Zn-lactate

(71% bitterness inhibition), followed by non-fat milk (49%), 250 mM sucrose (47%),

Na-gluconate (31%), and mocha (10%).

Sodium and zinc ions as bitterness inhibitors

As previously shown, the utility of zinc ions as a bitterness inhibitor in foods

and pharmaceuticals is compromised by the fact that zinc ions are potent inhibitors of

sweetness (Keast, Canty, & Breslin, 2004b). However, zinc ions are useful to help

our understanding of the bitterness elicited by caffeine. While human psychophysical

studies cannot directly test oral peripheral mechanisms of taste, such studies can

provide information to help understand the taste system (Schiffman, Booth, Sattely-

Miller, Graham, & Gibes, 1999). The influence of zinc ions on bitterness is

presumably in the oral periphery (Keast & Breslin, 2005), and zinc ions are known to

modulate allosterically trans-membrane receptors (GPCRs and ion channels) and can

both activate or inhibit them depending on the receptor system (Swaminath, Lee, &

Kobilka, 2003; Zheng, Gingrich, Traynelis, & Conn, 1998). To inhibit bitterness,

zinc ions may form a complex with the extracellular portions of the bitter taste

receptor/s (TAS2Rs), as zinc ions readily complex with amino acids and proteins and


16

have a high affinity for both thiol and hydroxy groups (Christianson, 1991). If zinc

ions did bind to a TAS2R, the native configuration of the receptor could be altered

and made unavailable for normal reception. Alternatively, zinc ions could form

complexes with the bitter compounds that would render them insoluble and, thus,

unable to access receptors; however, visual inspection of all solutions did not reveal

any precipitation. It is of interest that there was no increase in bitterness as caffeine

concentration increased from 1.5 to 4.5 mM indicating the mechanism responsible for

increasing levels of bitterness was affected by Zn-lactate. It is reasonable to suggest

the zinc ions are modulating an extracellular GPCR’s, presumably a TAS2R but

further studies at a receptor level would be required to verify this. Moreover, there

appears to be a portion of caffeine bitterness that is not affected by Zn-lactate; this

may be due to intracellular activation of bitter taste by caffeine (Peri et al., 2000;

Rosenzweig et al., 1999).

Previous studies have shown sodium ions may or may not inhibit the

bitterness of caffeine (Breslin & Beauchamp, 1995b; Kamen, Pilgrim, Gutman, &

Kroll, 1961; Pangborn, 1960). This study shows Na-gluconate significantly decreases

the bitterness of caffeine and inhibition was greater at 3 and 4.5 mM caffeine than at

lower concentrations (similar to zinc ions).

Bitter-sweet interactions

The mutual suppression of sweet and bitter tastes has been known for some

years (Calvino & Garrido, 1991; Kroeze & Bartoshuk, 1985; Pangborn, 1960). The

suppression is primarily central cognitive and named mixture suppression where both

bitter and sweet taste intensities are mutually suppressed when mixed in binary

solutions (Keast & Breslin, 2003; Kroeze & Bartoshuk, 1985). In this study the


17

sweetness of sucrose had a greater effect on the bitterness of caffeine than the reverse,

similar to the interaction reported by (Kamen et al., 1961). This effect may be due to

250 mM sucrose eliciting a higher intensity of sweetness than the corresponding

bitterness elicited by 4.5 mM caffeine. This does not adequately explain why higher

levels of bitterness failed to significantly suppress 125mM sucrose sweetness.

Milk-fat and caffeine bitterness

Fat emulsions have been shown to decrease bitterness intensity of quinine-HCl

(Metcalf & Vicker, 2002; Valentova & Pokorny, 1998). The authors speculated that

quinine-HCl preferentially partitioned into the fat phase of the emulsion thereby

diluting the concentration in the aqueous phase; tastants in the aqueous phase are less

able to access taste receptors, therefore quinine-HCl was less effective at reaching and

activating bitter taste receptors. Specific fatty acids have also been implicated in

reduction of quinine bitterness (Koriyama, Kohata, Wananabe, & Abe, 2002;

Koriyama, Wongso et al., 2002). Contrary to those previous studies, results from the

present study show that as the milk fat content increases from 0 to 4%, the level of

caffeine bitterness significantly increases. The milk matrix also contained milk

proteins and carbohydrates, so the significant increase in bitterness cannot be directly

attributed to the increasing fat content. Alternatively, milk proteins could form

complexes with caffeine that would render it insoluble and, thus, unable to access

receptors; as the solution was opaque, visual inspection of solutions did not reveal any

precipitation. Of further interest, comparing the bitterness of like concentrations of

caffeine in water (experiments 1& 2) and milk powder we can see there is a decrease

in bitterness (40-50%) when caffeine is in the milk powders: the decrease is greatest

when caffeine is in non-fat milk powder. The finding that milk powder, and in


18

particular non-fat milk powder, inhibits the bitterness of caffeine warrants further

inspection.

Aromas and caffeine bitterness

All aromas were congruent with caffeine bitterness but coffee, chocolate, or

mocha did not significantly inhibit the bitterness of caffeine. Indeed, both chocolate

(32%) and coffee (17%) aromas increased the perceived bitterness of caffeine. This

was expected as aroma can ‘acquire taste’ after a learning phase (Stevenson, Prescott,

& Boakes, 1995), and it is known that congruent aromas can modulate taste (Frank &

Byram, 1988; Small & Prescott, 2005). This result supports a study by Labbe et al.,

where a cocoa flavoured caffeinated beverage was perceived more bitter when tasted

with olfactory input than without (Labbe, Damevin, Vaccher, Morgenegg, & Martin,

2006). Chocolate was rated the most bitter of the three aromas at 1.5 & 3 mM

caffeine, while coffee was most bitter at 4.5 mM caffeine. Of interest was that mocha

(½ chocolate, ½ coffee) was perceived as significantly less bitter than coffee or

chocolate aromas indicating that the potential flavour association of mocha with

bitterness might not be as strong as with chocolate or coffee aromas.


19

CONCLUSION

Zinc ions were most effective at inhibiting the bitterness of caffeine

(presumably the mode of action is in the oral periphery) but the use of zinc in food

formulations has additional problems as zinc ions also inhibits sweetness. Other

methods to inhibit caffeine bitterness were only partially successful with 0% milk fat

milk the next most successful, although the mechanism of bitterness suppression

remains unknown. Finally, the cognitive strategy of mixture suppression using the

sweetness of 250 mM sucrose also proved successful at inhibiting the bitterness of

caffeine. Due to the complexity of the bitter taste system, the bitterness of caffeine

remains an ongoing problem for manufacturers wanting to add caffeine to food

formulations.


20

ACKNOWLEDGEMENTS

The author thanks Jessica Roper for technical assistance with this study. This work

was supported by The Institute of Biotechnology, Deakin University and the RPA

cluster funding from Health, Medicine, Nursing and Behavioural Sciences Faculty,

Deakin University.


21

REFERENCES

Adler, E., Hoon, M., Mueller, K., Chandrashekar, J., Ryba, N., & Zuker, C. (2000). A novel family of mammalian taste receptors. Cell, 100(6), 693-702.

Bartoshuk, L. (2000). Comparing sensory experience across individuals: Recent psychophysical advances illuminate genetic variation in taste perception. Chemical Senses, 25, 447-460.

Bartoshuk, L., & Seibyl, J. P. (1982). Suppression of QHCL in mixtures: possible mechanisms. Paper presented at the AChems, Sarasota 4th Annual Meeting.

Breslin, P., & Beauchamp, G. (1995a). Suppression of bitterness by sodium: variation among bitter taste stimuli. Chemical Senses, 20(6), 609-623.

Breslin, P. A., & Beauchamp, G. K. (1995b). Suppression of bitterness by sodium: variation among bitter taste stimuli. Chem Senses, 20(6), 609-623.

Calvino, A., & Garrido, D. (1991). Spatial and temperol behaviour of bitter sweet mixtures. Perceptual and Motor Skills, 73, 1216.

Chandrashekar, J., Hoon, M., Ryba, N., & Zuker, C. (2006). The receptors and cells for mammalian taste. Nature, 444(7117), 288-294.

Chandrashekar, J., Mueller, K., Hoon, M., Adler, E., Feng, L., Guo, W., et al. (2000). T2Rs function as bitter taste receptors. Cell, 100(6), 703-711.

Christianson, D. (1991). Structural biology of zinc. Advances in Protein Chemistry, 42, 281-355.

Clark, C., & Lawless, H. (1994). Limiting response alternatives in time-intensity scaling: an examination of the halo-dumping effect. Chemical Senses, 19(6), 583-594.

Delwiche, J. F., Buletic, Z., & Breslin, P. A. S. (2001). Covariation in individuals' sensitivities to bitter compounds: Evidence supporting multiple mechanisms. Perception and Psychophysics, 63(5), 761-776.

Flament, I. (2002). Coffee Flavor Chemistry. West Sussex: John Wiley & Sons. Frank, R., & Byram, J. (1988). Taste-smell interactions are tastant and odorant

dependent. Chemical Senses, 13, 445-455. Frank, R., Ducheny, K., & Mize, S. (1989). Strawberry odor, but not red color,

enhances the sweetness of sucrose solutions. Chemical Senses, 14, 371-377. Garrett, B. E., & Griffiths, R. R. (1998). Physical dependence increases the relative

reinforcing effects of caffeine versus placebo. Psychopharmacology (Berl), 139(3), 195-202.

Green, B., Dalton, P., Cowart, B., Shaffer, G., Rankin, K., & Higgins, J. (1996). Evaluating the 'Labeled Magnitude Scale' for measuring sensations of taste and smell. Chemical Senses, 21(3), 323-334.

Green, B. G., Shaffer, G. S., & Gilmore, M. M. (1993). Derivation and evaluation of a semantic scale of oral sensation magnitude with apparent ratio properties. Chemical Senses, 18, 683-702.

Holtzman, S. (1990). Caffeine as a model drug of abuse. Trends in Pharmacological Science, 11(9), 355-356.

Huang, L., Shanker, Y., Dubauskaite, J., Zheng, J., Yan, W., Rosenzweig, S., et al. (1999). Ggamma13 colocalizes with gustducin in taste receptor cells and mediates IP3 responses to bitter denatonium. Nature Neuroscience, 2(12), 1055-1062.

Hughes, J., Oliveto, A., Bickel, W., & Higgins, S. (1993). Indication of caffeine dependence in a population based study. In L. Harris (Ed.), Problems of drug dependence. Washington DC: US Government Printing Office.


22

Kamen, J. M., Pilgrim, F. J., Gutman, N. J., & Kroll, B. J. (1961). Interactions of suprathreshold taste stimuli. Journal of Experimental Psychology, 62, 348-356.

Keast, R., & Roper, J. (2007). A complex relationship among chemical concentration, detection threshold, and suprathreshold intensity of bitter compounds. Chemical Senses, 32(3), 245-253.

Keast, R. S., & Breslin, P. A. (2002). Modifying the bitterness of selected oral pharmaceuticals with cation and anion series of salts. Pharmaceutical Research, 19(7), 1019-1026.

Keast, R. S., & Breslin, P. A. (2005). Bitterness suppression with zinc sulfate and na-cyclamate: a model of combined peripheral and central neural approaches to flavor modification. Pharmaceutical Research, 22(11), 1970-1977.

Keast, R. S., Canty, T. M., & Breslin, P. A. (2004a). The influence of sodium salts on binary mixtures of bitter-tasting compounds. Chemical Senses, 29(5), 431-439.

Keast, R. S. J. (2003). The effect of zinc on human taste perception. Journal of Food Science, 68(5), 1871-1877.

Keast, R. S. J., & Breslin, P. A. S. (2003). An overview of binary taste-taste interactions. Food Quality and Preference, 14(2), 111-124.

Keast, R. S. J., Canty, T., & Breslin, P. A. S. (2004b). Oral zinc solutions inhibit sweet taste perception. Chemical Senses, 29(6), 513-521.

Kinnamon, S., & Margolskee, R. (1996). Mechanisms of taste transduction. Current Opinion in Neurobiology, 6(4), 506-513.

Koriyama, T., Kohata, T., Wananabe, K., & Abe, H. (2002). Effects of docosahexaenoic acid content in triacylglycerol on human taste perception. Journal of Food Science, 67(6), 2352-2356.

Koriyama, T., Wongso, S., Wananabe, K., & Abe, H. (2002). Fatty acid compositions of oil species affect the 5 basic taste perceptions. J Food Sci, 67(2), 868-873.

Kroeze, J. H. A., & Bartoshuk, L. M. (1985). Bitterness suppression as revealed by split-tongue taste stimulation in humans. Physiology & Behavior, 35, 779-783.

Labbe, D., Damevin, L., Vaccher, C., Morgenegg, C., & Martin, N. (2006). Modulation of perceived taste by olfaction in familar and unfamilar beverages. Food Quality and Preference, 17, 582-589.

Metcalf, K., & Vicker, Z. (2002). Taste intensities of oil-in-water emulsions with varying fat content. Journal of Sensory Studies, 17, 379-390.

Moon, S., Kottgen, M., Jiao, Y., Xu, H., & Montell, C. (2006). A taste receptor required for the caffeine response in vivo. Current Biology, 16, 1812-1817.

Nehlig, A. (1999). Are we dependent upon coffee and caffeine? A review on human and animal data. Neuroscience and Biobehavioral Reviews, 23(4), 563-576.

Pangborn, R. M. (1960). Taste interrelationships. Food Research, 25, 245-256. Peri, I., Mamrud-Brains, H., Rodin, S., Krizhanovsky, V., Shai, Y., Nir, S., et al.

(2000). Rapid entry of bitter and sweet tastants into liposomes and taste cells: implications for signal transduction. [In Process Citation]. The American Journal of Physiology, 278(1), C17-C25.

Rainnie, D., Grunze, H., McCarley, R., & Greene, R. (1994). Adenosine inhibition of mesopontine cholinergic neurons: implications for EEG arousal. Science, 263, 689-692.

Rosenzweig, S., Yan, W., Dasso, M., & Spielman, A. (1999). Possible novel mechanism for bitter taste mediated through cGMP. Journal of Neurophysiology, 81(4), 1661-1665.


23

Rossler, P., Kroner, C., Freitag, J., Noe, J., & Breer, H. (1998). Identification of a phospholipase C beta sub-type in rat taste cells. European Journal of Cell Biology, 77, 253-261.

Schiffman, S., Booth, B., Sattely-Miller, E., Graham, B., & Gibes, K. (1999). Selective inhibition of sweetness by the sodium salt of +/-2-(4-methoxyphenoxy)propanoic acid. Chemical Senses, 24, 439-447.

Schuh, K. J., & Griffiths, R. R. (1997). Caffeine reinforcement: the role of withdrawal. Psychopharmacology (Berl), 130(4), 320-326.

Small, D., & Prescott, J. (2005). Odor/taste integration and the perception of flavor. Experimental brain research, 166(3-4), 345-357.

Spielman, A., Huque, T., Whitney, G., & Brand, J. G. (1992). The diversity of bitter taste transduction mechanisms. In D. P. Corey & S. D. Roper (Eds.), Sensory Transduction (pp. 307-324). New York: Rockefeller Press.

Stevenson, R., Prescott, J., & Boakes, R. (1995). The acquisition of taste properties by odors. Learning & Motivation, 26, 1-23.

Swaminath, G., Lee, T., & Kobilka, B. (2003). Identification of an allosteric binding site for Zn2+ on the beta2 adrenergic receptor. Journal of Biological Chemistry, 278, 352-356.

Valentova, H., & Pokorny, J. (1998). Effect of edible oils and oil emulsions on the perception of basic tastes. Die Nahrung, 42(6), 406-408.

Wong, G., Gannon, K., & Margolskee, R. (1996). Transduction of bitter and sweet taste by gustducin [published erratum appears in Nature 1996 Oct 10; 383(6600):557]. Nature, 381(6585), 796-800.

Yeomans, M., Jackson, A., Lee, M., Steer, B., Tinley, E., Durlach, P., et al. (2000). Acquisition and extinction of flavour preferences conditioned by caffeine in humans. Appetite, 35(2), 131-141.

Zheng, F., Gingrich, M., Traynelis, S., & Conn, P. (1998). Tyrosine kinase potentiates NMDA receptor currents by reducing tonic zinc inhibition. Nature Neuroscience, 1, 185-191.


24

FIGURES

Figure 1 The influence of sodium gluconate and zinc lactate on the bitterness of

caffeine.

Each bar represents the average bitterness intensity of the compounds listed along the

X-axis. The Y-axis represents the average bitterness rating (arithmetic mean ±

standard error) on the gLMS (general Labeled Magnitude Scale) for caffeine

solutions. Abbreviations and concentrations of the compounds were: Caffeine (Caff)

(1.5, 3, 4.5 mM), Sodium Gluconate (NaGluc) (100 mM), Zinc lactate (Zn) (5 mM).

Differences among letters over bars indicate that means are statistically different

(p<0.0027) in bitterness intensity among relevant pairwise compounds only. For

example, grouping within vertical dashed lines, or the same salt at different caffeine

concentrations. Water is not included in the graph as the magnitude of bitterness was

<1gLMS unit.

Figure 2A The influence of the sweetness of sucrose on the bitterness of caffeine

Refer to Figure 1 for graph description. Abbreviations and concentrations of the

compounds were: Caffeine (Caff) (1.5, 3, 4.5 mM) and Sucrose (Suc) (125, 250 mM).

Figure 2B The influence of the bitterness of caffeine on the sweetness of sucrose

Each bar represents the average sweetness intensity of the compounds listed along the

X-axis. The Y-axis represents the average sweetness rating (arithmetic mean ±

standard error) on the gLMS (general Labeled Magnitude Scale) for sucrose solutions.

Abbreviations and concentrations of the compounds were: Caffeine (Caff) (1.5, 3, 4.5

mM) and Sucrose (Suc) (125, 250 mM). Refer to Figure 1 for graph description.


25

Figure 3 The influence of the milk fat on the bitterness of caffeine


compounds were: Caffeine (Caff) (1.5, 3, 4.5 mM) and Milk Fat (Fat) (0, 2, 4%).

Figure 4 The influence of congruent flavours on caffeine bitterness


compounds were: Caffeine (Caff) (1.5, 3, 4.5 mM) and coffee (coff), chocolate

(choc).


26

1.5m

M C

aff

1.5m

M C

aff N

aGlu

c

1.5m

M C

aff Z

n

3mM

Caf

f3m

M C

aff N

aGlu

c

3mM

Caf

f Zn

4.5m

M C

aff

4.5m

M C

aff N

aGlu

c

4.5m

M C

aff Z

n

gLM

S b

itter

ness

inte

nsity

0

2

4

6

8

10

12

aa

a

b

a,b,c

a,c

b

b,c

a


27

1.5m

M C

aff

1.5m

M C

aff 1

25 S

uc1.

5mM

Caf

f 250

Suc

3mM

Caf

f3m

M C

aff 1

25 S

uc3m

M C

aff 2

50 S

uc

4.5m

M C

aff

4.5m

M C

aff 1

25 S

uc4.

5mM

Caf

f 250

Suc

gLM

S bi

ttern

ess

inte

nsity

0

2

4

6

8

10

12

a

b

b

d

a,c,da

a

a,b,c

b,c,d


28

125m

M S

uc1.

5mM

Caf

f 125

Suc

3mM

Caf

f 125

Suc

4.5m

M C

aff 1

25 S

uc

250m

M S

uc1.

5mM

Caf

f 250

Suc

3mM

Caf

f 250

Suc

4.5m

M C

aff 2

50 S

uc

gLM

S sw

eet i

nten

sity

0

2

4

6

8

10

12

14

16

b

cc

b,c

a

a a a


29

1.5m

M C

aff 0

% F

at1.

5mM

Caf

f 2%

Fat

1.5m

M C

aff 4

% F

at3m

M C

aff 0

% F

at3m

M C

aff 2

% F

at3m

M C

aff 4

% F

at4.

5mM

Caf

f 0%

Fat

4.5m

M C

aff 2

% F

at4.

5mM

Caf

f 4%

Fat

gLM

S b

itter

ness

inte

nsity

0

2

4

6

8

10

a

a a

a,b

a,b

a,b

b

b b


30

1.5m

M C

aff m

ilk1.

5mM

Caf

f Cof

f1.

5mM

Caf

f Cho

c1.

5mM

Caf

f Moc

ha3m

M C

aff m

ilk3m

M C

aff C

off

3mM

Caf

f Cho

c3m

M C

aff M

ocha

4.5m

M C

aff m

ilk4.

5mM

Caf

f Cof

f4.

5mM

Caf

f Cho

c4.

5mM

Caf

f Moc

ha

gLM

S b

itter

ness

inte

nsity

0

2

4

6

8

deakin research online - home - drodro.deakin.edu.au/eserv/du:30017402/keast-modificationof...new...

Documents