ON THE GUSTATORY EFFECTS OF MIRACULIN AND GYMNEMIC ACID IN THE MONKEY

The summated response from the chorda tympani proper nerve of9 monkeys was recorded during stimulation with solutions ofacetic and citric acids, sodium chloride, quinine sulfate, sucrose,glucose and fructose before and after application of extractsof Synsepalum dulcificum-miraculin- and Gymnema sylvestre-gymnemicacid-on the tongue. It was observed that (a) miraculin enhancedthe response to all acids used (b) miraculin had no significanteffect on the response of the other taste stimuli (c) its effectlasts for more than h and was not removed by rubbing of thetongue (d) gymnemic acid had no significant effect on the responseto any of the stimuli used if miraculin had not been appliedbeforehand (e) gymnemic acid applied after miraculin diminishedthe response to acid, then miraculin enhanced the response toacid again. It was concluded that these electrophysiologicalfindings in monkey parallel the psychophysical observationsin man with regard to the effect of miraculin and gymnemic acidon the response to acids, but that they differ with regard tothe effect of gymnemic acid on the response to sugars. ^* On leave from Dept. of Psychology, University of New Hampshire,U.S.A. ^** On leave from Dept. of Oral Physiology, Osaka University,Japan.     

On the sense of taste in two Malagasy Primates (Microcebus murinus and Eulemur mongoz)

The relationship between phylogeny and taste is of growing interest.In this study we present recordings from the chorda tympaniproper (CT) nerve of two lemuriforme primates, the lesser mouselemur (Microcebus murinus) and the mongoose lemur (Eulemur mongoz),to an array of taste stimuli which included the sweeteners acesulfame-K,alitame, aspartame, D-glucose, dulcin, monellin, neohesperidindihydrochalcone (NHDHC), saccharin, sodium superaspartame, stevioside,sucralose (TGS), sucrose, suosan, thaumatin and xylitol, aswell as the non–sweet stimuli NaC1, citric acid, tanninand quinine hydrochloride. In M.murinus the effects of the tastemodifiers gymnemic acid and miraculin on the CT response wererecorded. Conditioned taste aversion (CTA) experiments in M.murinusand two-bottle preference (TBP) tests in E.mongoz were alsoconducted. We found that all of the above tastants except thaumatinelicited a CT response in both species. The CTA technique showedthat M.murinus generalized from sucrose to monellin but notto thaumatin. The intake of aspartame, ranging in concentrationfrom 0.1 to 30 mM was measured in E.mongoz with TBP tests. Atno concentration did we see a preference, but there was a significantrejection of 10 and 30 mM aspartame (P     

The taste of polycose in hamsters

Hamsters show a preference for Polycose, a mixture of starch-derived glucose polymers, that is as strong as their preference for sucrose. However, in the hamster, taste aversions to Polycose may be less easily acquired than taste aversions to sucrose and the qualitative aspects of Polycose are unknown in this species. In order to examine the taste of Polycose in the hamster, we utilized a taste-aversion protocol with two conditioning trials. Animals were trained to avoid one of three different conditioning stimuli: 50 mM sucrose, 100 mM Polycose and a mixture of 50 mM sucrose with 100 mM Polycose. Control animals were conditioned with deionized water. After the second conditioning trial, generalization testing began for the three conditioning stimuli plus 3 mM citric acid, 300 mM KCI and 30 mM NaCl. The results showed that aversions to Polycose, sucrose or the Polycose/sucrose mixture cross- generalized, demonstrating that Polycose and sucrose share a common taste percept in the hamster. None of the aversions generalized to NaCl, citric acid or KCI. In addition, comparisons among the patterns of taste generalizations indicated that the tastes of Polycose and sucrose also had distinct qualitative components. Finally, although the taste of 100 mM Polycose was more salient than the taste of 50 mM sucrose, the taste of sucrose could still be detected in a mixture with Polycose.      

Qualitative psychophysical studies on the gustatory effects of the sweet tasting proteins thaumatin and monellin

The gustatory effects of the sweet tasting proteins thaumatinand monellin were studied aftei application to small areas onthe anterior third of the tongue or to single fungiform papillae.The sweet sensation caused by thaumatin and monellin developedmore slowly, but reached a higher intensity and had a longerduration than that given by sucrose. Also, the response evokedby these sweet tasting proteins was more pronounced at the lateraledges, whereas that evoked by sucrose was stronger at the tipof the tongue. The taste modifier, miraculin, had no noticeableeffect on the sweet taste elicited by thaumatin, monellin andsucrose. Gymnemic acid abolished the sweet taste of all threecompounds. Experiments with time intervals of less than one minute betweenstimuli showed strong crossadaptation between thaumatin andmonellin, between the two proteins and sucrose, and betweenthe two proteins and miraculin-induced sweet taste of citricacid. While the differences in response to the sweet tasting proteinsand sucrose may be taken as evidence in favor of the existenceof more than one kind of sweet receptor, the cross-adaptationnoted between the various substances tested, would seem to indicatethat, at some point, they engage a common neural mechanism. ¹On leave from Dept. of Prosthetics, Faculty of Odontology,Karolinska Institutet. Present address: Dept. of Histology,Karolinska Institutet, S-104 01 STOCKHOLM, Sweden.     

Comparative studies on citric acid production by Aspergillus niger and Candida lipolytica using molasses and glucose

Citric acid production by Aspergillus niger NCIM 548 and Candida lipolytica NCIM 3472 has been studied in shake culture using glucose and molasses as carbon sources. Methanol addition (3% v/v) at 40 h of fermentation enhanced the production of citric acid by Aspergillus niger whereas a reduction in citric acid production by Candida lipolytica was observed with addition of methanol. Maximum citric acid concentration of 12 kg/m³ was obtained with Aspergillus niger using molasses in the presence of methanol, while maximum citric acid concentration of 8.4 kg/m³ was obtained with Candida lipolytica using glucose without methanol. It appears that product formation by Aspergillus niger is either non-growth associated or partially growth associated depending on the substrate. Methanol addition changes the nature of product formation in case of Candida lipolytica.     

Chorda tympani proper nerve responses to intra-arterial and surface stimulation of the tongue in rhesus monkey and rat

The effect of intra-arterial injection of the proteins monellin,thaumatin and miraculin on the activity of the chorda tympaniproper nerve were recorded in the rhesus monkey (Macaca mulatto)and the rat (Sprague–Dawley). The substances were injectedinto the blood stream to the lingual artery. It was found thatmonellin and thaumatin elicited a response only in the monkeyand not in the rat. Acetylated thaumatin, a tasteless substance,gave no increase of the nerve activity. Miraculin had no effectin either species. NaCl, sucrose and citric acid injected intra-arteriallygave a response in both species. It is concluded that the responsesto intra-arterial injections were caused by stimulation of thetaste buds and not nerve fibers. The results suggest there aretaste receptors also on the parts of the taste cells not facingthe oral cavity. The finding that there was no cross-adaptationbetween intra-arterial and oral application supports this conclusion.     

Influence of physical exercise on human preferences for various taste solutions

The effects of physical exercise on preference for various sapid solutions was studied in 58 healthy university students. After 30 min of exercise using a bicycle ergometer at 50% VO2max (maximal oxygen uptake) intensity, a rating scale test on taste hedonic tone and the triangle test for taste absolute threshold were done. The test solutions were sucrose, NaCl, citric acid, caffeine and monosodium glutamate (MSG). Preference scale values for sucrose and citric acid increased after exercise, whereas the values for NaCl, caffeine and MSG were not changed. The absolute thresholds for all the sapid solutions did not differ for pre- and post-exercise. These findings indicate that in humans preference for sucrose and citric acid increase after physical exercise.      

Effects of amiloride on salt taste in humans

Amiloride reduces several responses of the taste system to NaCl.These effects are crucial support for the ion transport theoryof salt taste. Here, three experiments tested for the inhibitoryeffect of amiloride on salt taste in normal, whole-mouth tastingin humans. There was no evidence that amiloride reduces thesaltiness of NaCl when used as a pretreatment, mixed into solutionwith NaCl, or both. In a fourth experiment, pretreating thetongue with amiloride-soaked filter paper and presenting NaClon filter paper within the treated area also had no effect onsaltiness. In addition, amiloride, a bitter compound, did notsuppress the taste of sucrose, and caffeine did not enhancethe taste of NaCl. These results suggest the ion transport theoryof NaCl taste needs to be modified or extended to account fornormal whole-mouth tasting of NaCl in humans. ¹ Present address: 1605 Harbor Court Tower, 10 East Lee St.,Baltimore, MD 21202, USA     

Taste modifiers: parotid reflexes can differ from taste judgements

Sweetness-depressing gymnemic acid (G) and sweetness-inducingmiraculin (M) helped determine the extent to which parotid salivaryresponses match behavioral and neural gustatory responses. Parotidflow rates and tastes intensities were obtained from four subjectsfor four sweeteners (before and after G) and for citric acid(CA) before and after M and G. A mixture of 20 mM CA and 10%sucrose was also tested. Although G depressed sweetness forglucose, sucrose, fructose and aspartame, G generally failedto alter parotid responses or depress post-G ratings for bitterintensity. In fact, G markedly elevated parotid responses forthe acid–sucrose mixture. Residual sweetness after G,detected mainly from the posterior tongue, probably contributedonly partially to sustaining post–G parotid responses.We speculate that side tastes from the sweeteners and oral irritationfrom CA in the mixture contributed to elevated flow rates afterG. Sucrose- and M-induced sweetness generally elevated parotidresponses for 20 mM CA. This result agrees with human chordatympani responses after M but differs for sucrose–acidmixtures in that parotid responses approached the calculatedsum of the components. We speculate that non-gustatory inputsmay also affect sweet–sour responses and advise cautionin relating parotid reflexes only to taste judgements. ¹Present address: Department of Biological Sciences, San JoseState University, San Jose, CA 95192, USA     

Taste responses of human chorda tympani nerve

Records from humans of summated action potential dischargesof the chorda tympani nerve were examined. The magnitudes ofneural and psychophysical responses were well related only whenthe comparison was made within a given taste quality. The responseto a mixture of 0 02 M citric acid and 0.5 M sucrose was lessthan the sum of the separate responses to the mixture components.Citric acid failed to cross-adapt the response to sucrose, implyingthe receptor sites for sucrose are independent of citric acid.The human chorda tympani nerve shows vigorous responses to mechanicalstimulation and cooling of the tongue that are maintained aftertreatment of the tongue with a water extract of the herb Gymnemasylvestre. Gymnema extract selectively suppressed the responseto all sweeteners tested (sucrose, fructose, saccharin and cyclamate)and also suppressed by – 50% the water-after-citric-acidresponse which has a predominantly sweet taste. Gymnema suppressedby 0 – 10% the water-rinse response following NaCl. fructoseand sucrose that have a predominantly bitter-sour taste. Water-rinseresponses were present even when mechanical and thermal stimulationof the tongue were minimized. The human chorda tympani nerveappears to have positive water-rinse taste responses. Theseare solute-specific off-responses that are probably mediatedby receptor sites independent of those responsible for the on-responseto the given solute.     

In vitro production of gymnemic acid from cell suspension cultures of Gymnema sylvestre R. Br

Gymnema sylvestre is an important medicinal plant that bears bioactive compound namely gymnemic acids. The present work deals with the optimization of a cell suspension culture system of Gymnema sylvestre for the production of biomass and gymnemic acid, which has anti‐diabetic properties. We investigated the effect of inoculum densities (2.5–20.0 g/L), the strength of the Murashige and Skoog (MS) medium (0.25–2.0), carbon source (sucrose, glucose, fructose, maltose), and the concentration of the sucrose (1–8% w/v) to determine their effects on biomass accumulation and production of gymnemic acid. Overall, 10 g/L of inoculum density, full‐strength MS medium supplemented with 2,4‐dichlorophenoxy acetic acid (2.0 mg/L) and Kinetin (0.1 mg/L), and 3% w/v sucrose was found best for the accumulation of biomass and gymnemic acid content (9.95 mg/g dry weight). The results of the current study will be useful for bioprocess and biochemical engineers for large‐scale production of gymnemic acid in cell culture.     

Hypoxia affects root sodium and chloride concentrations and alters water conductance in salt-treated jack pine (<Emphasis Type="Italic">Pinus banksiana</Emphasis>) seedlings

The effects of NaCl were studied in 6-month-old jack pine (Pinus banksiana Lamb.) seedlings growing in solution culture under hypoxic (approximately 2 mg l^у O₂) and well-aerated (approximately 8 mg l^у O₂) conditions. The results showed that hypoxia led to further reduction of stomatal conductance (g_s) in plants treated with 45 mM NaCl. This effect was likely due to a reduction in root hydraulic conductance by both stresses. When applied individually or together, neither 45 mM NaCl nor hypoxia affected cell membrane integrity of needles as measured by tissue electrolyte leakage. Hypoxia did not alter shoot Na⁺ and Cl^m concentrations in NaCl-treated plants. However, root Na⁺ concentrations were lower in NaCl-treated hypoxic plants, suggesting that hypoxia affected the ability of roots to store Na⁺. Hypoxia also induced root electrolyte leakage from NaCl-treated and control plants. The higher root Cl^m concentrations compared with Na⁺ and the positive correlation between root Cl^m concentrations and electrolyte leakage suggest that Cl^m played a major role in salt injury observed in jack pine seedlings. Roots of well-aerated plants treated for 1 week with NaCl contained almost two-fold higher concentration of total non-structural carbohydrates compared with plants from other experimental treatments and these concentrations decreased in subsequent weeks. We suggest that under prolonged hypoxic conditions, roots lose the ability to prevent Cl^m uptake resulting in the increase in root Cl^m concentration, which has damaging effects on root cell membranes.     

Confusion in the use of the taste adjectives 'sour' and 'bitter'

Subjects from Britain and America were tested on their skillat applying the taste adjectives ‘sweet’, ‘sour’,‘salty’, and ‘bitter’ to clearly distinguishablesolutions of sucrose, citric acid, NaCl and quinine sulphate,respectively. The main error that occurred was calling citricacid ‘bitter’ while the tendency to call quininesulphate ‘sour’ was not so common; this is the wellknown sour-bitter confusion. A sour-salty confusion was alsonoted as well as a tendency to call citric acid ‘sweet’.All these confusions were rectified by mere definition usingstandards. Skill at applying taste adjectives was not alwaysfound to be consistent over time. More errors occurred at lowersolution concentrations, even though stimuli were clearly distinguishable;indistinguishability of stimuli may account for some confusionsin other studies. ^*Present address: Department of Psychology, University of Bristol,Bristol BS8 1HH, England.     

Salt, nutrient uptake and transport, and ABA of Populus euphratica; a hybrid in response to increasing soil NaCl

The uptake and transport of salt ions (Na⁺, Cl^-), macronutrients (K⁺, Ca²⁺, Mg²⁺) and abscisic acid (ABA) response to increasing soil salinity were examined in 2-year-old seedlings of Populus euphratica and a hybrid, P. talassica Kom 2 (P. euphratica + Salix alba L.). Leaf burn symptoms appeared in the hybrid after 8 days of exposure to salinity when soil NaCl concentration increased to 206 mM, whereas P. euphratica exhibited leaf damage after day 21 when soil NaCl exceeded 354 mM. Leaf necrosis was the result of excess salt accumulation since the injury followed an abrupt increase of endogenous salt levels. Compared with the hybrid, P. euphratica exhibited a greater capacity to exclude salt ions from leaves under increasing salinity, especially Cl^-. Salt treatment altered nutrient balance of the hybrid, leaf K⁺, Ca²⁺ and Mg²⁺ concentrations significantly declined and the same trends were observed in roots with the exception of K⁺. Although K⁺ levels decreased in salinised P. euphratica, increasing salinity did not affect the levels of Ca²⁺ and Mg²⁺ in leaves, but did increase the uptake of these nutrients when salt stress was initiated. NaCl-induced increase of ABA concentration in xylem sap [ABA] was observed in the two tested genotypes, however xylem [ABA] increased more rapidly in P. euphratica and a fivefold increase of xylem [ABA] was recorded after the first day of exposure to salt stress. Therefore, we conclude that the increase of Ca²⁺ uptake may be associated with the rise of ABA, and thus contributes to membrane integrity maintenance, which enables P. euphratica to regulate uptake and transport of salt ions under high levels of external salinity in the longer term.     

Chemosensitive responses from the cat epiglottis

Responses were recorded from single and few-fiber preparationsof the cat superior laryngeal nerve during stimulation of theepiglottis with 0.5 M KCl, NH⁴Cl, NaCl, and LiCl; distilledwater, 0.005 M citric acid, 0.01 N HCl; and light touch. Epiglottalreceptive fields were mapped. The order of effective stimuliis KCl > HCl > NH⁴Cl > distilled water > citricacid; NaCl and LiCl are generally not effective. Since (a) similarstimuli elicit responses from the cat chorda tympani nerve duringstimulation of tongue taste buds, and (b) receptive fields ofsuperior laryngeal nerve fibers compare well with a map of epiglottaltaste buds, we conclude that the chemosensitive responses fromthe epiglottis are probably mediated by taste buds.     

Optimized fed-batch fermentation of L-β-hydroxy isobutyric acid by Yarrowia lipolytica

Yarrowia lipolytica KCCM50506, which transforms isobutyric acid to L-#-hydroxy isobutyric acid (L-#-HIBA), was screened. Chemostat cultures were carried out in jar fermentors at dilution rates of 0.02 h^у to 0.12 h^у. L-#-HIBA fermentation-regulating factors were determined to be specific growth rate, and concentrations of glucose and isobutyric acid in fermentor from analysis of steady-state data. The specific productivity of L-#-HIBA increased as the specific growth rate increased, apparently as a growth-associated type of product formation. A fed-batch culture was carried out under optimum conditions where the concentrations of glucose and isobutyric acid in the fermentor were maintained at 23 g l^у and 9 g l^у, respectively. The concentrations of cells and L-#-HIBA obtained at the end of fermentation were 20 g l^у and 49 g l^у, respectively, corresponding to 2.0 and 2.7 times more than concentrations in batch culture.     

Taste function in patients with oral burning

Burning mouth syndrome (BMS) is an oral pain disorder occurring primarily in post-menopausal women and is frequently accompanied by taste complaints. This association of symptoms suggests an interaction between the mechanisms of nociception and gustation, two senses with strong hedonic components. Seventy-three patients of the Taste and Smell Clinic at the University of Connecticut Health Center who reported experiencing 'unexplained oral burning' were evaluated for taste function. Both intensity ratings and quality identifications were measured for a concentration series of sucrose ('sweet'), NaCl ('salty'), citric acid ('sour') and quinine-HCl ('bitter'). The 57 women with BMS gave lower intensity ratings to NaCl and sucrose than comparably aged, same sex controls. Concentrations of NaCl and sucrose >0.10 M were most affected; concentrations of sucrose and NaCl <0.10 M were rated similarly by BMS and control women. No intensity differences were found for citric acid or quinine-HCl at any concentration and no differences were evident between the 16 BMS men and the 14 control men for any stimulus. The BMS women also misidentified the quality of 19% of the stimuli that were detected whereas control women misidentified 8%. Both groups detected a similar proportion of stimuli and found lower stimulus concentrations more difficult to identify than higher concentrations. Identification of NaCl as 'salty' and citric acid as 'sour' was particularly difficult for BMS women. The present findings are consistent with the hypothesis that pain pathway activation may affect neural and behavioral taste function.     

INTERACTION OF ASTRINGENCY AND TASTE CHARACTERISTICS1

The perception of astringency and basic taste in mixtures and their interaction effects were investigated by two procedures. In Experiment 1, focused and nonfocused testing procedures were compared using mixtures of low and high concentrations of alum and basic taste solutions. Both procedures yielded taste and astringency intensities that were modality‐dependent. Nonfocused testing was used in Experiment 2 to investigate the interactions of astringent phenolic (tannic acid) and nonphenolic (alum) compounds with each basic taste. Sweetness of sucrose increased with increased concentration with or without alum or tannin present. Changes in salty, bitter, and sour taste intensities were modality‐dependent. Astringency either remained unchanged or decreased with the addition of sucrose, sodium chloride, citric acid, or caffeine depending upon the taste concentration. Bitterness of tannin and alum at high concentrations was suppressed by the addition of sucrose, sodium chloride, or citric acid; sourness also decreased in the presence of sucrose or sodium chloride as well as a high level of caffeine.     

Amiloride-sensitive and amiloride-insensitive responses to NaCl + acid mixtures in hamster chorda tympani nerve

Component signaling in taste mixtures containing both beneficial and dangerous chemicals depends on peripheral processing. Unidirectional mixture suppression of chorda tympani (CT) nerve responses to sucrose by quinine and acid is documented for golden hamsters (Mesocricetus auratus). To investigate mixtures of NaCl and acids, we recorded multifiber responses to 50 mM NaCl, 1 and 3 mM citric acid and acetic acid, 250 μM citric acid, 20 mM acetic acid, and all binary combinations of each acid with NaCl (with and without 30 μM amiloride added). By blocking epithelial Na(+) channels, amiloride treatment separated amiloride-sensitive NaCl-specific responses from amiloride-insensitive electrolyte-generalist responses, which encompass all of the CT response to the acids as well as responses to NaCl. Like CT sucrose responses, the amiloride-sensitive NaCl responses were suppressed by as much as 50% by citric acid (P = 0.001). The amiloride-insensitive electrolyte-generalist responses to NaCl + acid mixtures approximated the sum of NaCl and acid component responses. Thus, although NaCl-specific responses to NaCl were weakened in NaCl-acid mixtures, electrolyte-generalist responses to acid and NaCl, which tastes KCl-like, were transmitted undiminished in intensity to the central nervous system. The 2 distinct CT pathways are consistent with known rodent behavioral discriminations.     

Temporal Characteristics of Gustatory Responses in Rat Parabrachial Neurons Vary by Stimulus and Chemosensitive Neuron Type

It has been demonstrated that temporal features of spike trains can increase the amount of information available for gustatory processing. However, the nature of these temporal characteristics and their relationship to different taste qualities and neuron types are not well-defined. The present study analyzed the time course of taste responses from parabrachial (PBN) neurons elicited by multiple applications of “sweet” (sucrose), “salty” (NaCl), “sour” (citric acid), and “bitter” (quinine and cycloheximide) stimuli in an acute preparation. Time course varied significantly by taste stimulus and best-stimulus classification. Across neurons, the ensemble code for the three electrolytes was similar initially but quinine diverged from NaCl and acid during the second 500ms of stimulation and all four qualities became distinct just after 1s. This temporal evolution was reflected in significantly broader tuning during the initial response. Metric space analyses of quality discrimination by individual neurons showed that increases in information (H) afforded by temporal factors was usually explained by differences in rate envelope, which had a greater impact during the initial 2s (22.5% increase in H) compared to the later response (9.5%). Moreover, timing had a differential impact according to cell type, with between-quality discrimination in neurons activated maximally by NaCl or citric acid most affected. Timing was also found to dramatically improve within-quality discrimination (80% increase in H) in neurons that responded optimally to bitter stimuli (B-best). Spikes from B-best neurons were also more likely to occur in bursts. These findings suggest that among PBN taste neurons, time-dependent increases in mutual information can arise from stimulus- and neuron-specific differences in response envelope during the initial dynamic period. A stable rate code predominates in later epochs.