Biomimetic Sensors for the Senses: Towards Better Understanding of Taste and Odor Sensation.

Taste and smell are very important chemical senses that provide indispensable information on food quality, potential mates and potential danger. In recent decades, much progress has been achieved regarding the underlying molecular and cellular mechanisms of taste and odor senses. Recently, biosensors have been developed for detecting odorants and tastants as well as for studying ligand-receptor interactions. This review summarizes the currently available biosensing approaches, which can be classified into two main categories: in vitro and in vivo approaches. The former is based on utilizing biological components such as taste and olfactory tissues, cells and receptors, as sensitive elements. The latter is dependent on signals recorded from animals' signaling pathways using implanted microelectrodes into living animals. Advantages and disadvantages of these two approaches, as well as differences in terms of sensing principles and applications are highlighted. The main current challenges, future trends and prospects of research in biomimetic taste and odor sensors are discussed.

Membrane-permeable tastants amplify ß2-adrenergic receptor signaling and delay receptor desensitization via intracellular inhibition of GRK2's kinase activity.

BACKGROUND: Amphipathic sweet and bitter tastants inhibit purified forms of the protein kinases GRK2, GRK5 and PKA activities. Here we tested whether membrane-permeable tastants may intracellularly interfere with GPCR desensitization at the whole cell context. METHODS: ß2AR-transfected cells and cells containing endogenous ß2AR were preincubated with membrane-permeable or impermeable tastants and then stimulated with isoproterenol (ISO). cAMP formation, ß2AR phosphorylation and ß2AR internalization were monitored in response to ISO stimulation. IBMX and H89 inhibitors and GRK2 silencing were used to explore possible roles of PDE, PKA, and GRK2 in the tastants-mediated amplification of cAMP formation and the tastant delay of ß2AR phosphorylation and internalization. RESULTS: Membrane-permeable but not impermeable tastants amplified the ISO-stimulated cAMP formation in a concentration- and time-dependent manner. Without ISO stimulation, amphipathic tastants, except caffeine, had no effect on cAMP formation. The amplification of ISO-stimulated cAMP formation by the amphipathic tastants was not affected by PDE and PKA activities, but was completely abolished by GRK2 silencing. Amphipathic tastants delayed the ISO-induced GRK-mediated phosphorylation of ß2ARs and GRK2 silencing abolished it. Further, tastants also delayed the ISO-stimulated ß2AR internalization. CONCLUSION: Amphipathic tastants significantly amplify ß2AR signaling and delay its desensitization via their intracellular inhibition of GRK2. GENERAL SIGNIFICANCE: Commonly used amphipathic tastants may potentially affect similar GPCR pathways whose desensitization depends on GRK2's kinase activity. Because GRK2 also modulates phosphorylation of non-receptor components in multiple cellular pathways, these gut-absorbable tastants may permeate into various cells, and potentially affect GRK2-dependent phosphorylation processes in these cells as well.

Studies on the aroma of five fresh tomato cultivars and the precursors of cis- and trans-4,5-epoxy-(E)-2-decenals and methional.

Three tasty (BR-139, FA-624, and FA-612) and two less tasty (R-144 and R-175) fresh greenhouse tomato cultivars, which significantly differ in their flavor profiles, were screened for potent odorants using aroma extract dilution analysis (AEDA). On the basis of AEDA results, 19 volatiles were selected for quantification in those 5 cultivars using gas chromatography-mass spectrometry (GC-MS). Compounds such as 1-penten-3-one, ( E, E)- and ( E, Z)-2,4-decadienal, and 4-hydroxy-2,5-dimethyl-3(2 H)-furanone (Furaneol) had higher odor units in the more preferred cultivars, whereas methional, phenylacetaldehyde, 2-phenylethanol, or 2-isobutylthiazole had higher odor units in the less preferred cultivars. Simulation of the odor of the selected tomato cultivars by preparation of aroma models and comparison with the corresponding real samples confirmed that all important fresh tomato odorants were identified, that their concentrations were determined correctly in all five cultivars, and that differences in concentration, especially of the compounds mentioned above, make it possible to distinguish between them and are responsible for the differential preference. To help elucidate formation pathways of key odorants, labeled precursors were added to tomatoes. Biogenesis of cis- and trans-4,5-epoxy-( E)-2-decenals from linoleic acid and methional from methionine was confirmed.

Identification and aroma impact of norisoprenoids in orange juice.

Four norisoprenoids, alpha-ionone, beta-ionone, beta-cyclocitral, and beta-damascenone, along with their putative carotenoid precursors, were identified in Valencia orange juice using time-intensity GC-O, GC-MS, and photodiode array HPLC. alpha-Ionone and beta-cyclocitral are reported in orange juice for the first time. GC-O aroma peaks were categorized into seven groups with similar sensory qualities: citrus/minty, metallic/mushroom/geranium, roasted/cooked/meaty/spice, fatty/soapy/green, sulfury/solventy/medicine, floral, and sweet fruity. The four norisoprenoids contributed approximately 8% of the total aroma intensity and 78% of the total floral aroma category. The putative carotenoid norisoprenoid precursors, alpha- and beta-carotene, alpha- and beta-cryptoxanthin, and neoxanthin, were identified in the same orange juice using photodiode array HPLC retention times and spectral characteristics.

Thermal oxidation of 9'-cis-neoxanthin in a model system containing peroxyacetic acid leads to the potent odorant beta-damascenone.

The potent odorant beta-damascenone was formed directly from 9'-cis-neoxanthin in a model system by peroxyacetic acid oxidation and two-phase thermal degradation without the involvement of enzymatic activity. Beta-damascenone formation was heavily dependent on pH (optimum at 5.0) and temperature, occurring over the two sequential phases. The first was incubation with peroxyacetic acid at 60 degrees C for 90 min, and the second was at above 90 degrees C for 20 min. Only traces of beta-damascenone were formed on application of only one of the two phases. Formate and citrate solutions produced a much better environment for beta-damascenone formation than acetate and phosphate. About 7 microg/L beta-damascenone was formed from 5.8 mg/L 9'-cis-neoxanthin under optimal experimental condition. The detailed pathway by which beta-damascenone is formed remains to be elucidated.

Linalool in orange juice: origin and thermal stability.

Linalool concentrations were determined in juice from three groups of 60 Valencia oranges using pentane:ether extraction and high-resolution capillary GC. The outer peel (flavedo) was removed from one group. The other two groups retained their peel intact. Juice was extricated from the halved fruits of the flavedo-less group and from one of the peel-intact groups using a hand reamer. A peel-cutting/macerating juice extractor was used for the other peel-intact group. Linalool concentrations were 0.004 mg/L in peeled fruit juice and 0.020 and 0.106 mg/L for hand-reamed and mechanically extracted peel-intact juice, respectively. Juice from peeled fruit contained significantly (P < 0.05) less linalool than peel-intact juice. Approximately 80% of the total juice linalool content was associated with peel using reamer design, and 96% was associated with peel-cutting/macerating design. Linalool increased with increasing peel oil levels; however, the increases were not proportionate. Since all commercial juices are mechanically extracted, the vast majority of linalool in commercial orange juice originates from the peel and not from the juice vesicle cytoplasm. Juice from peel-macerated, mechanically extracted fruit increased from 0.106 to 0.134 mg/kg after thermal processing, whereas juice from reamer extraction was essentially unchanged.

GC-olfactometric characterization of aroma volatiles from the thermal degradation of thiamin in model orange juice.

Model orange juice solutions containing 0.024 mM thiamin hydrochloride were stored for up to 8 weeks at 35 degrees C in amber glass containers. Volatiles were evaluated, primarily, using gas chromatography (GC) with olfactometry but also with flame ionization detector, pulsed-flame photometer detector (PFPD) (sulfur specific), and MS detection. Both 2-methyl-3-furanthiol (MFT) and its dimer, bis(2-methyl-3-furyl) disulfide (MFT-MFT) were identified thus confirming that thiamin could serve as the precursor to these potent off-flavors in thermally degraded citrus juices. Thirteen aroma active components were observed. MFT and MFT-MFT were observed after only a few days storage, and produced 33% of the total aroma activity after 7 d storage. Both compounds were observed olfactometrically earlier than they could be detected using PFPD. Other aroma-active compounds included 4,5-dimethylthiazole (skunky, earthy), 3-thiophenethiol (meaty, cooked), 2-methyl-4,5-dihydro-3(2H)-thiophenone (sour-fruity, musty, green), 2-acethylthiophene (burnt), 2-formyl-5-methylthiophene (meaty), and 2-methyl-3-(methyldithio) furan (meaty).

Aroma composition changes in early season grapefruit juice produced from thermal concentration.

Differences in aroma components and total volatiles between a single unpasteurized Marsh grapefruit juice and its 65 Brix concentrate reconstituted to 10 Brix were examined using GC-olfactometry (GC-O) and GC-FID. Total volatiles (FID) in the reconstituted concentrate were reduced to less than 5% of initial values, but 57% of total aroma (GC-O) remained. Forty-one aroma-active compounds were observed in unpasteurized single strength juice, whereas 27 components were found in the unflavored reconstituted concentrate. Aroma-active compounds were classified into grapefruit/sulfury, sweet/fruity, fresh/citrusy, green/fatty/metallic, and cooked/meaty groups. Five of six components in the sweet/fruity and 14 of 18 green/fatty/metallic components survived thermal concentration. However, only 4-mercapto-4-methyl-2-pentanone in the grapefruit/sulfury group, and linalool and nootkatone from the fresh/citrusy group, were found in the reconstituted concentrate. Methional was the only aroma compound in the cooked/meaty category found in both juice types. beta-Damascenone and 1-p-menthen-8-thiol were found only in the reconstituted concentrate. 4-Mercapto-4-methyl-2-pentanol was found for the first time in grapefruit juice.

Differential effects of tomato (Lycopersicon esculentum mill) matrix on the volatility of important aroma compounds.

Significant tomato matrix effects on the volatility of certain fresh tomato odorants were found. The concentrations of odorants such as (E,E)-2,4-decadienal, beta-damascenone, and beta-ionone, in crushed fresh tomato fruit obtained by solid-phase microextraction (SPME), resulting from a tomato matrix calibration curve were 5.5-, 2-, and 12-fold higher, respectively, than those calculated by calibration based on buffer solutions. Static headspace analyses indicated that, in most cases, the tomato matrix significantly retains the odorants relative to the buffer solution. Thus, the concentration of odorants in the headspace of tomato is lower than expected compared to a simple matrix such as buffer. CaCl(2), although needed in crushed fruit tissue to block enzymatic activity, was found to interact specifically with 2-isobutylthiazole, reducing its content in the headspace by at least 6-fold. If a matrix effect is found, analysis of the odorant molecule contents in the headspace rather than in the food is recommended in order to better evaluate their access to the olfactory receptors.

Inhibition of signal termination-related kinases by membrane-permeant bitter and sweet tastants: potential role in taste signal termination.

Sweet and bitter taste sensations are believed to be initiated by the tastant-stimulated T1R and T2R G protein-coupled receptor (GPCR) subfamilies, respectively, which occur in taste cells. Although such tastants, with their significantly diverse chemical structures (e.g., sugar and nonsugar sweeteners), may share the same or similar T1Rs, some nonsugar sweeteners and many bitter tastants are amphipathic and produce a significant delay in taste termination (lingering aftertaste). We report that such tastants may permeate rat taste bud cells rapidly in vivo and inhibit known signal termination-related kinases in vitro, such as GPCR kinase (GRK)2, GRK5, and PKA. GRK5 and perhaps GRK2 and GRK6 are present in taste cells. A new hypothesis is proposed in which membrane-permeant tastants not only interact with taste GPCRs but also interact intracellularly with the receptors' downstream shutoff components to inhibit signal termination.

Some sweet and bitter tastants stimulate inhibitory pathway of adenylyl cyclase via melatonin and alpha 2-adrenergic receptors in Xenopus laevis melanophores.

The sweeteners saccharin, D-tryptophan, and neohesperidin dihydrochalcone (NHD) and the bitter tastant cyclo(Leu-Trp) stimulated concentration-dependent pigment aggregation in a Xenopus laevis melanophore cell line similar to melatonin. Like melatonin, these tastants inhibited (by 45-92%) cAMP formation in melanophores; pertussis toxin pretreatment almost completely abolished the tastant-induced cAMP inhibition, suggesting the involvement of the inhibitory pathway (Gi) of adenylyl cyclase. The presence of luzindole (melatonin receptor antagonist) almost completely abolished the inhibition of cAMP formation induced by saccharin, D-tryptophan, and cyclo(Leu-Trp) but only slightly affected the inhibitory effect of NHD. In contrast, the presence of an alpha2-adrenergic receptor antagonist, yohimbine, almost completely abolished the inhibition of cAMP formation induced by NHD but had only a minor effect on that induced by the other tastants. Thus saccharin, D-tryptophan, and cyclo(Leu-Trp) are melatonin receptor agonists whereas NHD is an alpha2-adrenergic receptor agonist, but both pathways lead to the same transduction output and cellular response. Formation of D-myo-inositol 1,4,5-trisphosphate (IP3) in melanophores was reduced (15-58%, no concentration dependence) by saccharin, D-tryptophan, and cyclo(Leu-Trp) stimulation but increased by NHD stimulation. Tastant stimulation did not affect cGMP. Although some of the above tastants were found to be membrane permeant, their direct activation of downstream transduction components in this experimental system is questionable. MT1 and MT2 melatonin receptor mRNAs were identified in rat circumvallate papilla taste buds and nonsensory epithelium, suggesting the occurrence of MT1 and MT2 receptors in these tissues. Melatonin stimulation reduced the cellular content of cAMP in taste cells, which may or may not be related to taste sensation.