Chem. Senses 27: 653-659,
2002
© Oxford University Press 2002
Turbidity as a Measure of Salivary Protein Reactions with Astringent Substances
Department of Food Science, Cornell University, Ithaca, NY 14853, USA
Correspondence to be sent to: Harry Lawless, Department of Food Science, Stocking Hall, Cornell University, Ithaca, NY 14853, USA. e-mail: htl1{at}cornell.edu
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Abstract |
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 Top Abstract IntroductionExperiment 1. Instrumental...Experiment 2. Correlation of...General discussionReferences |
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Binding of tannins to proline-rich proteins has been proposed as an initial step in the development of astringent sensations. In beer and fruit juices, formation of tannin—protein complexes leads to the well-known effect of haze development or turbidity. Two experiments examined the development of turbidity in human saliva when mixed with tannins as a potential in vitro correlate of astringent sensations. In the first study, haze was measured in filtered human saliva mixed with a range of tannic acid concentrations known to produce supra-threshold psychophysical responses. The second study examined relationships among individual differences in haze development and the magnitude of astringency ratings. Mostly negative correlations were found, consistent with the notion that high levels of salivary proteins protect oral tissues from the drying effects of tannic acid.
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Introduction |
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TopAbstractIntroductionExperiment 1. Instrumental...Experiment 2. Correlation of...General discussionReferences |
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Oral astringency is a complex group of sensations involving dryness, roughness of oral surfaces and tightening, drawing or puckering sensations of the mucosa and muscles around the mouth (Lee and Lawless, 1991
;
Corrigan Thomas and Lawless,
1996; Gawel et al.,
2000). Astringency may also involve the classical taste pathways,
as demonstrated by recordings from taste nerves
(Schiffman et al.,
1992; Critchley and Rolls,
1996). However, much of what human observers report is tactile in
nature and is perceived in areas of the mouth innervated only by trigeminal
nerve endings, a `diffuse sensation' in the words of Bate Smith
(Bate Smith, 1954;
Breslin et al., 1993).
Astringency is produced by a variety of oral chemical stimuli, including
tannins and other polyphenols (Gawel,
1998), tannin substituents such as catechins
(Thorngate and Noble, 1995),
acids (Rubico and McDaniel,
1992; Hartwig and McDaniel,
1995; Lawless et al.,
1996) and aluminum salts (Lee
and Lawless, 1991).
Polyphenolic compounds such as tannins form complexes with salivary
proteins and mucopolysaccharides. Protein—tannin complexes result in the
precipitation and/or aggregation of salivary proteins causing them to lose
their lubricating properties. These interactions have led to a theory that
astringency is at least initially due to delubrication via removal of the
slippery coating on oral surfaces. Thus the mouth feels rough, tightened and
dry. Support for the delubrication theory comes from recent work showing
increases in coefficients of friction when tannins are added to saliva
(Prinz and Lucas, 2000).
Thorngate and Noble (Thorngate and Noble,
1995) pointed out that mechano-receptors are likely involved in
some of the astringent sensations, which seems reasonable given the movement
involved in appreciating sensations of roughness or dryness.
A large family of proline-rich proteins (PRPs) is found in human saliva
(Hay and Oppenheim, 1974;
Kaufman and Keller, 1979) that
are thought to serve functions of wetting, lubrication and protection of the
oral epithelium. Due to the disruption of 
-helical structures by
proline and exposed sites such as the carbonyl groups in the keto-imide
linkages, the PRPs provide a good substrate for the formation of hydrogen
bonds with tannins and for hydrophobic interactions
(McManus et al.,
1981; Murray et al.,
1994). Animals on high tannin diets may produce increased levels
of salivary proteins, which may in turn modulate their sensory responses to
astringency (Glendinning,
1992). This increase in secretions might represent a protective
mechanism since high concentrations of tannins have deleterious effects on
nutrient uptake (Mehansho et al.,
1983; Kim, 1994).
However, Prinz and Lucas (Prinz and Lucas,
2000) proposed that the increased friction and its related
sensations of dryness and roughness may serve as a sensory warning cue to
foraging animals to avoid plant substances that are high in tannins.
Repeated stimulation with astringent tannins shows a pattern of partial
recovery and build-up of intensity in a stepwise manner
(Guinard et al.,
1986). From this perspective, astringency may be viewed as a kind
of `temporarily induced xerostomia' caused by delubrication of oral tissues.
Salivary flow rate has recently been reported to modulate astringent reactions
(Fischer et al., 1994;
Ishikawa and Noble, 1995).
These studies showed slower onset and slower decay of astringent sensations
among groups with low salivary flow rates. Support for the involvement of
salivary proteins in astringency comes from recent work by Kallikathraka
et al. (Kallikathraka et
al., 1998), showing changes in the chromatographic profile of
salivary constituents following stimulation with tannins, possibly due to
formation of protein—tannin complexes. Further work by this group showed
correlations between the time parameters of astringent sensations and specific
peaks in a salivary chromatogram
(Kallikathraka et al.,
2001). Direct action of astringents on the oral epithelium and/or
taste receptors is also possible.
Recent publications in food chemistry bring to light a surprising parallel
between the salivary binding/delubrication hypothesis for astringency and the
association of polyphenols and proteins to form `chill haze' in cider, beer
and other beverages (McMurrough et
al., 1992). PRPs, particularly hordein, a barley protein,
bind with polyphenols to produce turbidity
(Asano et al., 1982).
This reaction is largely specific to PRPs. Both hydrogen bonding and
hydrophobic interactions are likely mechanisms for these associations
(Asano et al., 1982;
Siebert et al.,
1996). A model of polyphenol—protein crosslinking, similar
to that proposed for tannin binding to oral proteins
(McManus et al.,
1981; Haslam and Lilley,
1988), was recently proposed for chill haze
(Siebert et al.,
1996).
This literature substantiates the possibility that binding of tannins to
PRPs is a likely mechanism contributing to astringent sensations.
Protein-binding assays such as simple turbidity measurements may be useful as
a measure of tannin—protein association in the saliva of experimental
subjects who are simultaneously generating psychophysical data. Imm and
Lawless (1996) found lowered
astringency responses amongst individuals with higher overall protein content.
However, establishing relationships between salivary protein content and
perceived astringency has not always met with success. Kallikathraka et
al. (Kallikathraka et al.,
2001) found no relationship between total protein and astringency
response and Guinard et al.
(Guinard et al.,
1998) found no relationship between parotid protein content and
astringency. However, Kallikathraka et al. did find significant
correlations between the magnitude of specific protein fractions and
astringency. An assay that mimics the type of chemical interactions that occur
between salivary proteins and tannins might provide a better correlate with
sensory responses. Based on the literature from beverage research on chill
haze, the development of turbidity in saliva—tannin mixtures could be
such a correlate. A preliminary report from our laboratory showed development
of haze in saliva—tannic acid mixtures over a range of tannic acid
concentrations known to elicit astringent sensations
(Lawless et al.,
1999). The studies reported here follow up with a larger
group.
The objective of the first study was to determine whether human saliva would show measurable haze development over a range of concentrations known to be astringent. A second experiment examined whether individuals with varying haze development would co-vary in their astringency responses. Such a result would implicate salivary proteins as a contributor or modulator of astringent sensations. Flow rate was also examined as a potential correlate of astringency. Subjects with a higher resting rate of salivary flow would be expected to have increased levels of salivary protein available. Increased levels or protein might have a protective coating effect, reducing astringency, and a thus a negative correlation between resting flow and astringency as well as a negative relationship between haze and astringency.
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Experiment 1. Instrumental measurement of salivary turbidity |
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TopAbstractIntroductionExperiment 1. Instrumental...Experiment 2. Correlation of...General discussionReferences |
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Methods
Subjects were 25 volunteers from the Cornell University campus in Ithaca, NY (13 females, 12 males, age range 18-56 years, age median 24 years). Subjects were paid a token cash incentive for their participation.
Whole-mouth saliva was collected by having each panelist expectorate into a
60 ml cup. Panelists were permitted to chew on a square of Parafilm in order
to evoke saliva. Panelists reported not having consumed any food or drink
other than water for at least 1 h before saliva collection. Panelists were
instructed to provide 30-35 g of saliva and had access to a balance to monitor
their progress. Saliva from each panelists was individually filtered in a
vacuum (2 mm Hg) through Whatman no. 1 filter paper. Aliquots ranging from 20
to 31 ml (median 26 ml) were obtained and stored in individual sterile
screw-top vials. All saliva was used and discarded within 48 h of collection.
Saliva held overnight refrigerated at 4°C was allowed to equilibrate to
room temperature (
20°C) before analysis. Four milliliters of saliva
from each individual was combined with 4 ml of tannic acid solution [lot
93H0269, formula weight (FW) 1701.18; Sigma, St Louis, MO] at the following
concentrations: 0, 0.5, 0.89 and 1.58 g/ml. All solutions were placed in
capped 10 ml clear borosilicate glass test tubes and mixed once by
inversion.
Turbidity was measured using a Hach Laboratory Turbidimeter (model 2100N; Hach Co., Loveland, CO). The optical system in this instrument measures light scattering in nephelometric turbidity units (NTUs) and is comprised of a tungsten filament lamp, lenses and apertures to focus the light, a 90° detector, forward scatter detector and a transmitted light detector. The instrument was used for this experiment in ratio measurement mode in which it gathers data from all three detectors and determines a ratio from the signals. The instrument was calibrated prior to the experiment with Formazin primary standards prepared from a 4000 NTU stock (Hach) and HPLC grade deionized water. The calibration of the instrument was verified periodically during the experiment using Gelex secondary turbidimetry standards.
Turbidity (NTUs) was measured immediately after mixing (0 min) for each
test tube and then at 15 min intervals for 105 min. Baseline data were
collected by measuring the turbidities (NTUs) of 4 ml of deionized distilled
water mixed with 4 ml of the following concentrations of tannic acid: 0, 0.5,
0.89 and 1.58 g/l. Concentrations were chosen to correspond to levels of
tannic acid used in previous psychophysical studies
(Lee and Lawless, 1991).
A four-way repeated measures ANOVA with tannic acid concentration and time as within subject factors, gender as a between subjects factor and panelists as a random effect was used to detect differences in the data. Statistical analyses were performed using Statistica v.5.1 (Statsoft Inc., Tulsa, OK). Results reported below are significant at P < 0.01.
Results and discussion
Figure 1 shows the turbidity
development with increasing concentrations of tannic acid. Turbidity increased
with increasing concentrations of tannic acid [F(3,69) = 71.2] and
increased slightly over incubation time, as shown in
Figure 2, except for the no
tannin control [interaction F(21, 483) = 18.7]. Turbidity of
solutions with tannin only and no saliva was .5 NTUs (not plotted). No gender
effects were observed. The haze development confirmed earlier observations on
the potential use of turbidity measurements as an indicator of the
interactions of salivary proteins and tannins
(Lawless et al.,
1999). Measurable haze development is seen over a range of
concentrations that are physiologically active in the dynamic range of sensory
intensity ratings (Guinard et
al., 1986; Lee and
Lawless, 1991; Lawless et
al., 1994). The method is rapid and simple to execute. This
further supports the possibility of using in vitro haze development
as a predictor of astringency responses in psychophysical tests.
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One question of interest is whether haze development should be positively
or negatively correlated with sensory responses. Both haze development and
astringency responses increase with concentration
(Lawless et al.,
1994) across the same ranges. Comparing these concentrations to
previous psychophysical data (Lee and
Lawless, 1991), responses would span the middle third of an
intensity scale (roughly from 5 to 10 on a 15 point category scale). This
correspondence implies a positive correlation when concentration is the source
of variation. However, a negative correlation might be expected when variation
across individuals in salivary protein content is the source of variation. If
the function of salivary proteins is to lubricate and protect the epithelium
from reactions with astringents, then a person with higher levels of salivary
protein might be expected to have lower astringency ratings (better
protection), but higher levels of haze development. A negative correlation
would be consistent with the observation that less responsive individuals have
higher overall salivary protein content and more responsive individuals have
lower protein content (Imm and Lawless,
1996). If, on the other hand, the binding of tannins to proteins
produces a precipitate that itself causes astringency, then a positive
correlation would be expected. Experiment 2 was conducted to examine the
relationship of haze and astringency ratings across individuals.
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Experiment 2. Correlation of instrumental turbidity and perceived astringency |
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TopAbstractIntroductionExperiment 1. Instrumental...Experiment 2. Correlation of...General discussionReferences |
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The objective of Experiment 2 was to examine the relationship between instrumental turbidity and perceived astringency. To the extent that salivary proteins are bound by tannins in the initial steps of the development of astringent sensations, and assuming that the salivary haze development seen in Experiment 1 is an indicator of the extent of the saliva—protein interactions, a correlation across individuals is predicted, i.e. individuals with higher levels of protein would show greater haze development. If tannin—protein complexes or precipitates are themselves responsible for the astringent sensations a positive correlation would be expected. However, to the extent that salivary proteins provide a protective mechanism, an inverse relationship would be expected. The relationship between an individual's salivary protein content and the magnitude of their astringency responses is not yet clear (Guinard et al., 1998
; Kallikathraka
et al., 2001). Methods
Subjects were 18 volunteers from the Cornell University campus community in Ithaca, NY (10 females). Informed consent was obtained. Subjects were instructed not to eat or drink anything for at least 1 h prior to each experimental session. Subjects were paid a token cash incentive for their participation.
Samples used for evoking saliva were 1.58 g/l tannic acid (lot 93H0269; Sigma Chemical Co.) and 3.16 g/l citric acid (lot 11K0238, FW 210.1; Sigma Chemical Co.). Samples used to measure perceived astringency were 0.5, 0.89 and 1.58 g/l tannic acid and 1.0, 1.78 and 3.16 g/l citric acid. All samples were 15 ml and were presented at room temperature (20 ± 1°C).
All subjects participated in three experimental sessions. In the first session unstimulated saliva was collected by the whole mouth method for 20 min. Subjects chewed a 2 cm2 piece of Parafilm to evoke the saliva. Following mechanical stimulation of saliva subjects rested for 10 min and saliva was collected again for 150 s, evoked by either 1.58 g/l tannic acid or 3.16 g/l citric acid. Lastly, after another 10 min rest period, saliva was again collected for 150 s, evoked by either the citric acid or the tannic acid stimulus, whichever was not used previously.
Subjects were given a practice trial with water to familiarize themselves
with the time—intensity module of Compusense and how to respond with the
computer mouse. Astringency was not defined for them, although subjects had
participated in previous studies of astringency in our laboratory and none
enquired about further definition of the term. In those previous studies,
astringency was generally defined as a complex of drying, roughing and puckery
sensations experienced in the mouth after rinsing with alum or tannins, with 1
g/l alum generally given as an example. In the second and third sessions
subjects rated three concentrations of tannic acid and three concentrations of
citric acid for perceived astringency by continuous time intensity for 2 min
each. Citric acid was included for several reasons. Sour acids are also
astringent (Rubico and McDaniel,
1992; Hartwig and McDaniel,
1995; Lawless et al.,
1996), although it is unclear whether they share the same
mechanisms as polyphenols, and so a different pattern of correlations could
occur. Also, sour acids are potent sialogens
(Dawes et al., 2000),
so a different relationship with stimulated flow rate might be observed. All
three concentrations of each acid were presented in single sessions and the
order of presentation within and between sessions was randomized. There was a
5 min rest interval between each sample during which time subjects were
instructed to rinse their mouths with deionized water and chew on table water
biscuits. For each sample, subjects were instructed to place the entire sample
into their mouths and swish it about for 15 s, being sure to cover all oral
surfaces. At 15 s subjects were verbally signaled by the experimenter to
expectorate the sample and to begin rating. Data were collected using the
Compusense 5 v.4.0 time—intensity module. Ratings were made by moving
the mouse, which marked off height on a vertical line scale labeled only
`astringency'. The time—intensity module scores the data from 0 to 10
with resolution set at 0.01.
In assessing mechanically stimulated salivary flow, whole mouth saliva was collected in pre-weighed 60 ml plastic cups for 20 min. Immediately following collection individual saliva samples were weighed and flow rates (g/min) were calculated. Individual saliva samples were then filtered through Whatman no. 1 paper under vacuum. The filtrate was dispensed into individual 60 ml sterile screw-top plastic vials and the mass was again measured. Finally, haze was measured in individual saliva samples by combining 4 ml of filtered saliva with 4 ml of 1.58 g/l tannic acid. Haze was measured using the same apparatus and protocols as used in Experiment 1, except measurements were made at 0, 5, 10 and 15 min. Following analysis the saliva was discarded.
In measuring chemically evoked salivary flow, subjects were instructed to place the entire sample into their mouths and swish it about for 15 s, being sure to cover all oral surfaces. At 15 s subjects were verbally signaled by the experimenter to expectorate the sample and to begin collecting saliva. Whole mouth saliva was collected for 150 s using the same protocols as those described above. Immediately following collection individual saliva samples were weighed and flow rates (g/min) were calculated. Haze was not measured in these samples due to the presence of residual tastant chemicals.
Subjects were separated into either a high flow or low flow group and into a high haze or low haze group around the medians of mechanically stimulated salivary flow rates (median 1.5 g/min, range 0.53-2.16 g/min) and of measured haze at 15 min (median 15.2 NTU, range 7.7-28.1 NTU). Individual maximum intensities (Imax) were obtained from the time—intensity data for each sample. Correlation matrices were calculated that included Imax of perceived astringency for all samples, measured haze levels at each time interval, mechanically stimulated salivary flow rate, chemically evoked salivary flow rates, flow group and haze group.
Two-way split plot repeated measures ANOVAs were performed that compared Imax for each acid across the concentrations of each acid and among the two different flow groups and the two different haze groups. A two-way split plot repeated measures ANOVA was performed that compared haze levels (NTU) across all time intervals and among the different flow groups. Statistics reported below are significant at P < 0.05.
Results and discussion
Correlations among salivary flow rates, haze measures and intensity scores from tannic acid and citric acid are shown in Table 1. Haze development was positively correlated with mechanically stimulated salivary flow rate. This `resting' (i.e. chemically unstimulated) rate was negatively correlated with astringency ratings of tannic acid but not with astringency ratings of citric acid. Astringency was negatively correlated with haze development for tannic acid but not for citric acid. These results are consistent with a pattern of low responding subjects having more salivary protein and therefore better protection of the oral surfaces against the stimulation by tannins. Conversely, high flow subjects may have higher amounts of salivary protein, more haze development and are less susceptible to the influences of tannins on the oral epithelium. Responses to citric acid did not correlate significantly with haze development, although astringency was negatively correlated with the stimulated flow rate to citric acid itself. This suggests a simple dilution effect for citric acid and a different mechanism than the haze-developing protein interactions seen with tannic acid.
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A similar pattern of results was provided by between groups analyses of variance. Subjects were divided into high and low flow groups based on resting flow rate and into high and low haze groups based on median splits. This is not meant to imply that there was a bimodal distribution in flow or haze, but simply to provide another way of looking at statistical relationships among salivary flow rate, haze development and psychophysical responses. High flow subjects had lower responses to tannic acid astringency [F(1,16) = 10.04], with greater differences at higher concentrations [interaction F(2,32) = 5.42]. A parallel pattern was seen based upon haze group, with higher haze reflecting lower astringency [F(1,16) = 5.38], and the effect was also greater at higher concentrations [interaction F(2,32) = 5.64]. These relationships are shown in Figure 3. There was also greater haze development in the saliva of the high flow group [F(1,16) = 7.23]. No significant group effects were seen for citric acid.
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General discussion |
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TopAbstractIntroductionExperiment 1. Instrumental...Experiment 2. Correlation of...General discussionReferences |
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The studies reported here show a relationship between the production of turbidity in saliva—tannin mixtures and astringency responses. Our initial thought was that haze development might parallel astringency as haze increased across the range of tannin concentrations that were psychophysically active. However, pilot work from one author (J. Hayes) showed a surprising negative correlation between haze development and the initial height of time—intensity functions. That led to consideration of the delubrication hypothesis and the notion that salivary protein would indeed serve as a protectant against the direct attack of tannins on oral tissues. Therefore, the inverse relationship of haze development and the rated intensity of astringency can be explained by increased protein content among low responding individuals, who are `better protected'.
The inverse relationship of flow rate to astringency intensity (high flow, lower intensity) as shown in Figure 3 might suggest a simple dilution effect rather than protection by salivary proteins. However, flow rate (from tannic acid stimulation) and haze were positively correlated, implicating protein interactions as a probable cause of the effect. If there were a negative relationship between haze and flow, simple dilution would be more plausible. However, dilution might still play a role in the relationship to citric acid stimulation due to its ability to evoke copious flow. No significant relationships were observed between haze and citric acid-stimulated flow.
These studies provide no direct evidence of protein—tannin binding.
However, that hypothesis seems reasonable in light of the extensive literature
showing haze development in solutions of PRPs mixed with polyphenols and in
light of the well-known phenomenon of chill haze in beverages, a function of
tannin—protein interactions (Siebert
et al., 1996; Siebert
and Lynn, 1997; Siebert,
1999). Also, recent studies have addressed changes in haze in
saliva with a changes in pH, a variable known to affect tannin—protein
interactions (K.J. Siebert and A. W. Chassy, submitted for publication).
Demonstrating psychophysical parallels to such pH changes may not be
straightforward, because lowering pH in itself increases astringency
(Lawless et al.,
1996).
Kallikathraka et al.
(Kallikathraka et al.,
2001) demonstrated that different salivary protein fractions from
HPLC analysis were correlated either positively or negatively with
time—intensity measurements of astringency. An early eluting fraction
(peak 1 in their study), tentatively characterized as a hydrophilic PRP, was
positively correlated with total perceived astringency (measured by area under
the curve of the time—intensity response). They also noted a positive
correlation with time to maximum, suggesting that individuals with a higher
salivary protein content would take longer to achieve their peak intensity of
sensation. In a somewhat closer parallel to the current result, subsequent
peaks that were tentatively identified as non-PRP and hydrophobic PRP
fractions were negatively correlated with astringent intensity. This would be
consistent with a lubricating or protective role that minimized astringent
sensations among individuals with greater amounts of protein coating on the
oral epithelium. The observation in Experiment 2 of a negative correlation
with astringent intensity suggests that haze development may be due to binding
these hydrophobic PRPs. Are the individual fractions observed by
Kallikathraka's group differentially efficient at producing the salivary haze
effect? If such relations are found, they would help substantiate the
conclusion that the haze effect bears a mechanistic and predictive
relationship to the initial astringency reactions and is not just an
epiphenomenon.
In conclusion, the haze development effect adds to the evidence that
interactions of salivary proteins and polyphenols are a likely first step in
the development of astringency. Whether this interaction is sufficient to
explain the entire mechanism of astringency is open to question. As Thorngate
and Noble (Thorngate and Noble,
1995) pointed out, mechanoreceptors are likely involved in some of
the astringent sensations, which seems reasonable given the movement involved
in appreciating sensations of roughness or dryness. Binding with salivary
proteins might serve to sequester or inactivate tannins and thus protect the
alimentary tract from the deleterious effects of tannins on nutritional
uptake. However, it is not known whether such protein—tannin complexes
would survive the low pH of the stomach. As an alternative, Prinz and Lucas
(Prinz and Lucas, 2000)
proposed that the increased friction is simply a sensory warning that would
help animals avoid ingesting plants that are high in tannins. The interesting
question then arises of why these sensations are preferred by humans in foods
and beverages under some circumstances.
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Acknowledgments |
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The authors thank Dr Karl Seibert for suggestions and assistance. This work was supported in part by NIH DC00902.
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References |
|---|
|
TopAbstractIntroductionExperiment 1. Instrumental...Experiment 2. Correlation of...General discussionReferences |
|---|
Asano, K., Shinagawa, K. and Hashimoto, N. (1982) Characterization of haze forming proteins of beer and their roles in chill haze formation. J. Am. Soc. Brew. Chemists, 40,147 -154.
Bate-Smith, E. (1954) Astringency in foods. Food Proc. Packag., 23,124 -127.
Bate-Smith, E. (1973) Haemanalysis of tannins: the concept of relative astringency.Phytochemistry , 12,907 -912.[ISI]
Breslin, P.A.S., Gilmore, M.M., Beauchamp, G.K. and
Green, B.G. (1993) Psychophysical evidence that oral
astringency is a tactile sensation. Chem. Senses,18
, 405-417.
Critchley, H.D. and Rolls, E.T. (1996)
Responses of primate taste cortex neurons to the astringent tastant tannic
acid. Chem. Senses, 21,135
-145.
Corrigan Thomas, C.J. and Lawless, H.T.
(1996) Astringent subqualities in acids. Chem.
Senses, 20,593
-600.
Dawes, C., O'Connor, A.M. and Aspen, J.M. (2000) The effect on human salivary flow rate of the temperature of a gustatory stimulus. Arch. Oral Biol.,45 , 957-961.[ISI][Medline]
Fischer, U., Boulton, R.B. and Noble, A.C. (1994) Physiological factors contributing to the variability of sensory assessments: relationships between salivary flow rate and temporal perception of gustatory stimuli. Food Qual. Pref.,5 , 55-64.
Gawel, R. (1998) Red wine astringency: a review. Aust. J. Grape Wine Res.,4 , 74-95.
Gawel, R., Oberholster, A. and Francis, I.L. (2000) A `Mouth-feel Wheel': terminology for communicating the mouthfeel characteristics of red wine. Aust. J. Grape Wine Res., 6,203 -207.
Glendinning, J.I. (1992) Effect of salivary
proline-rich proteins on ingestive responses to tannic acid in mice.Chem. Senses
, 17,1
-12.
Guinard, J., Pangborn, R.M. and Lewis, M.J.
(1986) The time course of astringency in wine upon repeated
ingestions. Am. J. Enol. Viticult.,37
, 184-189.
Guinard, J. X., Zoumas-Morse, C. and Walchak, C. (1998) Relation between parotid saliva flow and composition and the perception of gustatory and trigeminal stimuli in foods.Physiol. Behav. , 63,109 -118.
Hartwig, P. and McDaniel, M.R. (1995) Flavor characteristics of lactic, malic, citric, and acetic acids at various pH levels. J. Food Sci.,60 , 384-388.
Haslam, E. and Lilley, T.H. (1988) Natural astringency in foodstuffs—a molecular interpretation.CRC Crit. Rev. Food Sci. Nutr. , 27,1 -40.
Hay, D.I. and Oppenheim, F.G. (1974) The isolation from human parotid saliva of a further group of proline-rich proteins. Arch. Oral Biol., 19,627 -632.[ISI][Medline]
Imm, B.Y. and Lawless, H.T. (1996) Relationships between salivary responses and astringency, bitterness and sourness responses to aluminum ammonium sulfate. Chem. Senses, 21,618 .
Ishikawa, T. and Noble, A.C. (1995) Temporal perception of astringency and sweetness in red wine.Food Qual. Pref. , 6,27 -33.
Kaufman, D.L. and Keller, P.J. (1979) The basic proline-rich proteins in human parotid saliva from a single subject. Arch. Oral Biol., 24,249 -256.[ISI][Medline]
Kallikathraka, S., Bakker, J. and Clifford, M.N. (1998) Evidence that salivary proteins are involved in astringency. J. Sensory Stud., 13,29 -43.
Kallikathraka, S., Bakker, J., Clifford, M.N. and Vallis, L. (2001) Correlations between salivary protein composition and some TI parameters of astringency. Food Qual. Pref., 12,145 -152.
Kim, H.S. (1994) Tea and iron availability: role of dietary and salivary proteins on iron absorption. Inhibitory effects of tea tannins. PhD thesis, Cornell University, Ithaca, NY.
Lawless, H.T., Corrigan, C.L. and Lee, C.B.
(1994) Interactions of astringent substances.Chem. Senses
, 19,141
-154.
Lawless, H.T., Horne, J. and Giasi, P.
(1996) Astringency of organic acids is related to pH.Chem. Senses
, 21,397
-403.
Lawless, H.T., Hartono, C. Horne, J. and Siebert, K.J. (1999) Interactions of tannins and human salivary proteins assessed by turbidimetry. Chem. Senses,24 , 543-544.
Lee, C.B. and Lawless, H.T. (1991)
Time-course of astringent materials. Chem. Senses,16
, 225-238.
McManus, J.P., Davis, K.G., Lilley, T.H. and Haslam, E. (1981) The association of proteins with polyphenols. J. Chem. Soc. Chem. Commun.,7 , 309-311.
McMurrough, I., Kelly, R. and Byrne, J. (1992) Effect of the removal of sensitive proteins and proanthocyanidins on the colloidal stability of lager beer. J. Am. Soc. Brew. Chemists, 50,67 -76.
Mehansho, H., Hagerman, A.E., Clements, S., Butler, L.G.,
Rogler, J. and Carlson, D.M. (1983) Modulation of
proline-rich protein biosynthesis in rat parotid glands by sorghum with high
tannin levels. Proc. Natl Acad. Sci. USA,80
, 3948-3952.
Murray, N.J., Williamson, P., Lilley, T.H. and Haslam, E. (1994) Study of the interaction between salivary proline-rich proteins and a polyphenol by 1H-NMR spectroscopy.Eur. J. Biochem. , 219,923 -935.[ISI][Medline]
Prinz, J.F. and Lucas, P.W. (2000) Saliva tannin interactions. J. Oral Rehab.,27 , 991-994.[ISI][Medline]
Rubico, S.M. and McDaniel, M.R. (1992)
Sensory evaluation of acids by free-choice profiling. Chem.
Senses, 17,273
-289.
Schiffman, S.S., Suggs, M.S., Sostman, A.L. and Simon, S.A. (1992) Chorda tympani and lingual nerve responses to astringent compounds in rodents. Physiol. Behav.,51 , 55-63.[Medline]
Siebert, K.J. (1999) Effects of protein-polyphenol interactions on beverage haze, stabilization and analysis. J. Agric. Food Chem., 47,353 -362.[ISI][Medline]
Siebert, K.J. and Lynn, P.Y. (1997) Haze active protein and polyphenols in apple juice assessed by turbidimetry. J. Food Sci., 62,79 -84.
Siebert, K.J., Carrasco, A. and Lynn, P.Y. (1996) Formation of protein-polyphenol haze in beverages. J. Agric. Food Chem.,44 , 1997-2005.
Thorngate, J.H. and Noble, A.C. (1995) Sensory evaluation of bitterness and astringency of 3R(-)-epichatechin and 3S(+)-catechin. J. Sci. Food. Agric.,67 , 531-535.
Accepted June 14, 2002
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