Bitter Compounds Research Articles

Immobilization of Rhodococcus by encapsulation and entrapment: a green solution to bitter citrus by-products.

Debittering of citrus by-products is required to obtain value-added compounds for application in the food industry (e.g., dietary fiber, bioactive compounds). In this work, the immobilization of Rhodococcus fascians cells by encapsulation in Ca-alginate hollow beads and entrapment in poly(vinyl alcohol)/polyethylene glycol (PVA/PEG) cryogels was studied as an alternative to chemical treatments for degrading the bitter compound limonin. Previously, the Rhodococcus strain was adapted using orange peel extract to increase its tolerance to limonoids. The optimal conditions for the encapsulation of microbial cells were 2% Na-alginate, 4% CaCl2, 4% carboxymethylcellulose (CMC), and a microbial load of 0.6 OD600 (optical density at 600nm). For immobilization by entrapment, the optimal conditions were 8% PVA, 8% PEG, and 0.6 OD600 microbial load. Immobilization by entrapment protected microbial cells better than encapsulation against the citrus medium stress conditions (acid pH and composition). Thus, under optimal immobilization conditions, limonin degradation was 32 and 28% for immobilization in PVA/PEG gels and in hollow beads, respectively, in synthetic juice (pH 3) after 72h at 25°C. Finally, the microbial cells entrapped in the cryogels showed a higher operational stability in orange juice than the encapsulated cells, with four consecutive cycles of reuse (runs of 24h at 25°C). KEY POINTS: • Increased tolerance to limonoids by adapting R. fascians with citrus by-products. • Entrapment provided cells with favorable microenvironment for debittering at acid pH. • Cryogel-immobilized cells showed the highest limonin degradation in citrus products.

Faba Bean (Vicia faba L. minor) Bitterness: An Untargeted Metabolomic Approach to Highlight the Impact of the Non-Volatile Fraction.

In the context of climate change, faba beans are an interesting alternative to animal proteins but are characterised by off-notes and bitterness that decrease consumer acceptability. However, research on pulse bitterness is often limited to soybeans and peas. This study aimed to highlight potential bitter non-volatile compounds in faba beans. First, the bitterness of flours and air-classified fractions (starch and protein) of three faba bean cultivars was evaluated by a trained panel. The fractions from the high-alkaloid cultivars and the protein fractions exhibited higher bitter intensity. Second, an untargeted metabolomic approach using ultra-high-performance liquid chromatography-diode array detector-tandem-high resolution mass spectrometry (UHPLC-DAD-HRMS) was correlated with the bitter perception of the fractions. Third, 42 tentatively identified non-volatile compounds were associated with faba bean bitterness by correlated sensory and metabolomic data. These compounds mainly belonged to different chemical classes such as alkaloids, amino acids, phenolic compounds, organic acids, and terpenoids. This research provided a better understanding of the molecules responsible for bitterness in faba beans and the impact of cultivar and air-classification on the bitter content. The bitter character of these highlighted compounds needs to be confirmed by sensory and/or cellular analyses to identify removal or masking strategies.

Current and emerging applications in detection and removal of bitter compounds in citrus fruit juice: A critical review

For decades, producers, researchers, and food safety agencies have faced the challenge of processing bitter citrus fruit and making it more appealing to consumers. The primary compounds responsible for the bitterness in citrus fruit juices are naringin and limonin. In order to extend the shelf life and enhance the acceptability of citrus fruit juices, similar to other tropical fruits, scientists and the food industry have been continuously working to reduce their bitterness. This has led to the development of various physical, chemical, and biological methods, which are commonly used scientific approaches for reducing bitterness in citrus fruit juices. This review emphasizes the significance of citrus fruit in the economy and discusses the different bittering agents present in citrus fruits, as well as the various techniques used to detect their presence. Additionally, it highlights different debittering techniques and the advancement of technologies aimed at achieving fast, reliable, and effective debittering of bitter citrus fruits. The review also examines the effects of different debittering techniques on the chemical and phytochemical properties of citrus juice and briefly discusses future prospects in this field. Furthermore, this review recognizes the potential of advanced techniques such as sensors to replace traditional methods. Traditional approaches often require a significant amount of time and technical expertise and can be expensive. The utilization of sensors and other advanced technologies can address these limitations and provide more efficient and cost-effective solutions in the debittering process.

Extension of immature stage of Hemicentrotus pulcherrimus ovary for reduction of the bitter compound (pulcherrimine) by water temperature manipulation

Extension of immature stage of Hemicentrotus pulcherrimus ovary for reduction of the bitter compound (pulcherrimine) by water temperature manipulation

Sensory-directed isolation and identification of an intense salicin-like bitter compound in infected teas with bird’s eye spot disease

Teas infected with bird’s eye spot disease generally exhibited a lingering and long-lasting, salicin-like bitter taste, which was unpalatable to consumers. Sensory-directed isolation processes have been performed in this study to investigate the salicin-like bitter compounds in infected teas. Results showed that infected teas were extracted using a 70% methanol aqueous solution to produce methanol extract, which was then further separated by sequential solvent extraction (SSE) to obtain dichloromethane extract, which contained the salicin-like bitter compounds. The dichloromethane extract was then isolated by flash chromatography to produce two salicin-like bitter fractions, eluted using 60% and 65% methanol aqueous solution. Finally, these two salicin-like bitter fractions were analyzed by RP-HPLC using 60–68% and 70–75% methanol aqueous solution, respectively, affording the location of the salicin-like bitter compounds in RP-HPLC chromatograms. Moreover, a new ursane-type triterpenoid, camellisin A methyl ester, was identified from infected teas. This study has provided preliminary isolation methods of salicin-like bitter compounds from the infected teas, which were essential to designing targeted debittering strategies for infected teas and improving the quality of the finished tea and the effective utilization of fresh tea leaves.

Hop-derived Humulinones Reveal Protective Effects in in vitro Models of Hepatic Steatosis, Inflammation and Fibrosis.

Nonalcoholic fatty liver disease (NAFLD) is emerging as leading cause of liver disease worldwide. Specific pharmacologic therapy for NAFLD is a major unmet medical need. Recently, iso-alpha acids, hop-derived bitter compounds in beer, have been shown to beneficially affect NAFLD pathology. Humulinones are further hop derived bitter acids particularly found in modern styles of beer. So far, biological effects of humulinones have been unknown. Here, we investigated the effect of humulinones in in vitro models for hepatic steatosis, inflammation and fibrosis. Humulinones dose-dependently inhibited fatty acid induced lipid accumulation in primary human hepatocytes. Humulinones reduced the expression of fatty acid uptake transporter CD36 and key enzymes of (de novo) lipid synthesis. Conversely, humulinones increased the expression of FABP1, CPT1 and ACOX1, indicative for increased lipid combustion. Furthermore, humulinones ameliorated steatosis induced pro-inflammatory gene expression. Furthermore, humulinones significantly reduced the expression of pro-inflammatory and pro-fibrogenic factors in control as well as lipopolysaccharide treated activated hepatic stellate cells, which play a key role in hepatic fibrosis. In conclusion, humulinones beneficially affect different pathophysiological steps of NAFLD. Our data suggest humulinones as promising therapeutic agents for the prevention and treatment of NAFLD.

Effect of Dissolved Oxygen and Chemical Scarification on Andrographis paniculata Seed Germination in Macrobubble Conditions

Andrographis paniculata is used in Thai traditional medicine. This plant contains a bitter compound called andrographolide, which is highly effective in the prevention of many diseases. It is an effective treatment for infectious diseases and has a prophylactic effect owing to its powerful immunity-boosting benefits. Recently, it has been widely used to treat COVID-19. However, commercial planting of A. paniculata is performed by seeding, which leads to seed germination problems. The seed germination is relatively low and not efficient under normal conditions for various reasons, such as a combined dormancy of physical and innate nature, the diversity of the seeds in different lots, and the fact that the germination duration was not uniform in the same lot. An easily applied and inexpensive method for farmers to develop mass plantings to stimulate germination is by using macrobubble conditions by aerating seeds in sterile water in collaboration with chemical scarification, which is the idea of creating a hard seed coat that causes seed dormancy to break while root germination occurs at 25°C. Germination was completed after 16 days. The dissolved oxygen (DO) concentrations in this environment were 5, 6, 7, 8, and 9 mg·L−1. The oxygen intensity of 9 mg·L−1 showed the highest germination percentage (26.33%). It was found to be optimal for macrobubble conditions. Seedlings were treated with chemicals (PEG, NaCl, H2SO4, KCl, KNO3, NaHClO3, and GA3) after soaking in macrobubbles with optimum DO. The results showed that NaHClO3 conc. (30 min) showed a generation percentage reaching 92%, which could greatly promote up to 3.63 folds compared with the control in the macrobubble aeration system.

Faba Bean Flavor Effects from Processing to Consumer Acceptability.

Faba beans as an alternative source of protein have received significant attention from consumers and the food industry. Flavor represents a major driving force that hinders the utilization faba beans in various products due to off-flavor. Off-flavors are produced from degradation of amino acids and unsaturated fatty acids during seed development and post-harvest processing stages (storage, dehulling, thermal treatment, and protein extraction). In this review, we discuss the current state of knowledge on the aroma of faba bean ingredients and various aspects, such as cultivar, processing, and product formulation that influence flavour. Germination, fermentation, and pH modulation were identified as promising methods to improve overall flavor and bitter compounds. The probable pathway in controlling off-flavor evolution during processing has also been discussed to provide efficient strategies to limit their impact and to encourage the use of faba bean ingredients in healthy food design.

Bitter Peptides in Fermented Soybean Foods - A Review.

Fermented soybean foods with a long history are popular worldwide because of rich nutrition. However, many traditional fermented soybean foods have unacceptable bitterness, which mostly comes from the bitter peptides produced from the hydrolysis of soybean proteins. In this review, the bitter peptides in fermented soybean foods is briefly reviewed. The structural properties of bitter receptors and bitter peptides were reviewed. Bitterness is perceived through the binding between bitter compounds and specific sites of bitter receptors (25 hTAS2Rs), which further activate the downstream signal pathway mediated by G-protein. And it converts chemical signals into electrical signals, and transmit them to the brain. In addition, the influencing factors of bitter peptides in fermented soybean foods were summarized. The bitterness of fermented soybean foods primarily results from the raw materials, microbial metabolism during fermentation, unique techniques, and interactions of various flavor compounds. Moreover, the structure-bitterness relationship of bitter peptides was also discussed in this review. The bitterness degree of the bitter peptide is related to the polypeptide hydrophobicity, amino acids in the peptide, peptide molecular weight and polypeptide spatial structure. Studying the bitter peptides and their bitter characteristics in fermented soybean foods is beneficial for improving the sensory quality of fermented soybean foods and prompting more consumers accept them.

Association between Genetic Variation in the TAS2R38 Bitter Taste Receptor and Propylthiouracil Bitter Taste Thresholds among Adults Living in Japan Using the Modified 2AFC Procedure with the Quest Method.

Individual taste sensitivity influences food preferences, nutritional control, and health, and differs greatly between individuals. The purpose of this study was to establish a method of measuring and quantifying an individual's taste sensitivity and to evaluate the relationship between taste variation and genetic polymorphisms in humans using agonist specificities of the bitter taste receptor gene, TAS2R38, with the bitter compound 6-n-propylthiouracil (PROP). We precisely detected the threshold of PROP bitter perception by conducting the modified two-alternative forced-choice (2AFC) procedure with the Bayesian staircase procedure of the QUEST method and examined genetic variation in TAS2R38 in a Japanese population. There were significant differences in PROP threshold between the three TAS2R38 genotype pairs for 79 subjects: PAV/PAV vs AVI/AVI, p < 0.001; PAV/AVI vs AVI/AVI, p < 0.001; and PAV/PAV vs PAV/AVI, p < 0.01. Our results quantified individual bitter perception as QUEST threshold values: the PROP bitter perception of individuals with the PAV/PAV or PAV/AVI genotypes was tens to fifty times more sensitive than that of an individual with the AVI/AVI genotype. Our analyses provide a basic model for the accurate estimation of taste thresholds using the modified 2AFC with the QUEST approach.

Molecular Basis of Hexanoic Acid Taste in Drosophila melanogaster.

Animals generally prefer nutrients and avoid toxic and harmful chemicals. Recent behavioral and physiological studies have identified that sweet-sensing gustatory receptor neurons (GRNs) in Drosophila melanogaster mediate appetitive behaviors toward fatty acids. Sweet-sensing GRN activation requires the function of the ionotropic receptors IR25a, IR56d, and IR76b, as well as the gustatory receptor GR64e. However, we reveal that hexanoic acid (HA) is toxic rather than nutritious to D. melanogaster. HA is one of the major components of the fruit Morinda citrifolia (noni). Thus, we analyzed the gustatory responses to one of major noni fatty acids, HA, via electrophysiology and proboscis extension response (PER) assay. Electrophysiological tests show this is reminiscent of arginine-mediated neuronal responses. Here, we determined that a low concentration of HA induced attraction, which was mediated by sweet-sensing GRNs, and a high concentration of HA induced aversion, which was mediated by bitter-sensing GRNs. We also demonstrated that a low concentration of HA elicits attraction mainly mediated by GR64d and IR56d expressed by sweet-sensing GRNs, but a high concentration of HA activates three gustatory receptors (GR32a, GR33a, and GR66a) expressed by bitter-sensing GRNs. The mechanism of sensing HA is biphasic in a dose dependent manner. Furthermore, HA inhibit sugar-mediated activation like other bitter compounds. Taken together, we discovered a binary HA-sensing mechanism that may be evolutionarily meaningful in the foraging niche of insects.

Lipopolysaccharide increases bitter taste sensitivity via epigenetic changes in Tas2r gene clusters

T2R bitter receptors, encoded by Tas2r genes, are not only critical for bitter taste signal transduction but also important for defense against bacteria and parasites. However, little is known about whether and how Tas2r gene expression are regulated. Here, we show that in an inflammation model mimicking bacterial infection using lipopolysaccharide, the expression of many Tas2rs was significantly upregulated and mice displayed markedly increased neural and behavioral responses to bitter compounds. Using single-cell assays for transposase-accessible chromatin with sequencing (scATAC-seq), we found that the chromatin accessibility of Tas2rs was highly celltype specific and lipopolysaccharide increased the accessibility of many Tas2rs. scATAC-seq also revealed substantial chromatin remodeling in immune response genes in taste tissue stem cells, suggesting potential long-lasting effects. Together, our results suggest an epigenetic mechanism connecting inflammation, Tas2r gene regulation, and altered bitter taste, which mayexplain heightened bitter taste that can occur with infections and cancer treatments.

Effect of Sequential Fermentation with Lachancea thermotolerans/S. cerevisiae on Aromatic and Flavonoid Profiles of Plavac Mali Wine.

In this study, the effects of sequential fermentation of Lachancea thermotolerans/S. cerevisiae on the production of Plavac Mali wines were investigated in comparison with the commonly used inoculation of the commercial Saccharomyces cerevisiae strain and spontaneous fermentation. A total of 113 aroma compounds and 35 polyphenolic compounds were analyzed. Sequential inoculation resulted in a decrease in alcohol content and pH (up to 0.3% v/v and 0.12 units, respectively) and an increase in total acidity (0.6 g/L, expressed as tartaric acid). The wines produced by spontaneous fermentation exhibited the greatest diversity of volatile compounds and the highest concentration of C13 norisoprenoids, lactones, and other compounds. These wines exhibited maximum hydroxycinnamic acids, prodelphinidin monomer units, epigallocatechin, B1, B3, and B4 dimers, and total flavan-3-ols. Sequential inoculation decreased the content of the aromas and polyphenols in the wines. The practical significance of this procedure lies in the selective effect on aroma compounds, the decrease in green aromas, undetectable volatile phenols, and the decrease in bitter and astringent compounds such as gallic acid, flavan-3-ol monomers (catechin and epicatechin), and dimers (B1, B2, B3, and B4). This work demonstrates the potential of sequential and spontaneous fermentation to improve the aromatic characteristics and overall quality of Plavac Mali wines.

Identification of a Bitter Peptide Contributing to the Off-Flavor Attributes of Pea Protein Isolates

The aversive bitter taste of pea protein ingredients limits product acceptability. Compounds contributing to the bitter perception of pea protein isolates were investigated. Off-line multi-dimensional sensory-guided preparative liquid chromatography fractionation of a 10% aqueous PPI solution revealed one main bitter compound that was identified by Fourier transform ion cyclotron resonance mass spectrometry and de novo tandem mass spectrometry (MS/MS) sequencing as the 37 amino acid peptide PA1b from pea albumin and further confirmed by synthesis. Quantitative MS/MS analysis reported that the concentration of the bitter peptide was 129.3 mg/L, which was above the determined bitter sensory threshold value of 3.8 mg/L and in agreement with the perceived bitter taste of the sample.

Regulation of Ras p21 and RalA GTPases activity by quinine in mammary epithelial cells.

Quinine, a bitter compound, can act as an agonist to activate the family of bitter taste G protein-coupled receptor family of proteins. Previous work from our laboratory has demonstrated that quinine causes activation of RalA, a Ras p21-related small G protein. Ral proteins can be activated directly or indirectly through an alternative pathway that requires Ras p21 activation resulting in the recruitment of RalGDS, a guanine nucleotide exchange factor for Ral. Using normal mammary epithelial (MCF-10A) and non-invasive mammary epithelial (MCF-7) cell lines, we investigated the effect of quinine in regulating Ras p21 and RalA activity. Results showed that in the presence of quinine, Ras p21 is activated in both MCF-10A and MCF-7 cells; however, RalA was inhibited in MCF-10A cells, and no effect was observed in the case of MCF-7 cells. MAP kinase, a downstream effector for Ras p21, was activated in both MCF-10A and MCF-7 cells. Western blot analysis confirmed the expression of RalGDS in MCF-10A cells and MCF-7 cells. The expression of RalGDS was higher in MCF-10A cells in comparison to the MCF-7 cells. Although RalGDS was detected in MCF-10A and MCF-7 cells, it did not result in RalA activation upon Ras p21 activation with quinine suggesting that the Ras p21-RalGDS-RalA pathway is not active in the MCF-10A cells. The inhibition of RalA activity in MCF-10A cells due to quinine could be as a result of a direct effect of this bitter compound on RalA. Protein modeling and ligand docking analysis demonstrated that quinine can interact with RalA through the R79 amino acid, which is located in the switch II region loop of the RalA protein. It is possible that quinine causes a conformational change that results in the inhibition of RalA activation even though RalGDS is present in the cell. More studies are needed to elucidate the mechanism(s) that regulate Ral activity in mammary epithelial cells.

Oral CD8 +T RMsurveil taste buds where their local reactivation drives papillary inflammation and bitter receptor expression

Abstract Resident memory T cells (T RM) expedite pathogen detection and clearance at typical portals of pathogen entry. While paradigms in T RMbiology were largely established in a few select locations, comparatively little is known about how CD8 +T RMintegrate within the oral cavity, which encompasses a functionally organized constellation of absorptive, secretory, and chemosensory tissues. To fill this knowledge gap, we developed a Viral-Prime, Epitope-Pull (VPEP) model for locally augmenting tractable populations of oral T RMto manipulate and study. Leveraging this approach, we document CD8 +T RMwithin and surrounding taste buds and papillae of the soft palate and tongue. Reactivation of oral T cells induced discernable inflammation within taste structures that was abrogated with prior in vivoCD103 +T RMdepletion. Taste buds represent a collection of ~50–100 individual taste receptor cells (TRCs) which sense nutritive and potentially harmful dietary components. We hypothesized that reactivation of taste bud associated T RMwould provoke TRC transcriptional changes including genes involved in gustation (taste). Bulk RNAseq was performed on T RM-reactivated circumvallate papillae and soft palate; primary taste-sensing locations in mice and enriched sources of TRCs relative to other oral tissues. Here we found the significant induction of several Tas2r(bitter receptor) genes not observed in control mice or T RMreactivated buccal mucosa. Teleologically, a heightened sensitivity to bitter compounds downstream of oral T RMreactivation may represent an evolved mechanism to protect against the accidental ingestion of noxious chemicals during oral or upper respiratory tract infections when other chemosensory modalities faulter. T90 DE 022732 K99DE031014

Profiling and In vivo studies of Bromelain Bitter Gourd (Momodica charantia) seed protein hydrolysate with antidiabetic activity

Diabetes mellitus is a multifactorial chronic disease that affects the human population and it is the third most common cause of death worldwide. Momordica charantia is commonly known as Bitter melon, Bitter guard and used as a food and natural medicine. The scientific name, Momordica means "to bite," in Latin which is refers to the jagged edges of the leaves. Including fruits, all parts of the plant, contains a bitter compound, momordicinso and very bitter in taste. It has long been used as a traditional medicine for some ailments. Bromelain bitter guard seed protein hydrolysate was profiled by ultrafiltration and SDS-PAGE analysis revealed that the lower molecular weight peptide ≤ 25 kDa exerted the high antidiabetic activity. Spontaneously diabetic rats showed a decrease in the blood Glucose level (Glu), Glycated haemoglobin (HbA1c) level, Glycogen level and also shows lower level of Lipid profile parameters (Chol, HDL, LDL, and TG) in the serum of the diabetic rats after 21 days of oral administration of bromelain bitter guard seed protein hydrolysate at dosages of 100 mg/kg, 200 mg/kg, and 400 mg/kg. However, the effect was dose-dependent. As a novel protein hydrolysate source with invivo antidiabetic activity, future research should aim to demonstrate the molecular mechanism of action and validate its bioactivity through human intervention trials.

Taste 2 receptors in GtoPdb v.2023.1

Taste 2 receptors or Bitter taste receptors (TAS2Rs) are G protein-coupled receptors expressed in oral sensory cells and a variety of non-gustatory tissues. The ~25 human TAS2Rs share low amino acid sequence identities with other GPCR families and are classified as broadly tuned "generalist" receptors with numerous, chemically diverse bitter agonists, as narrowly tuned "specialist" receptors with very few activators, as intermediately tuned receptors with an average number of agonists, or receptors specialized to interact with chemically defined activators [32]. The number of functional bitter taste receptor genes varies among species and orthologues might not be functionally conserved. Due to their expression in various tissues, the signal transduction of TAS2Rs is complex. Some TAS2Rs interact with drugs such as analgesic, anti-inflammatory, and antibacterial compounds. The specialist database BitterDB contains additional information on bitter compounds and receptors [14].

Sex Differences in the Bitterness Perception of an Aromatic Myrtle Bitter Liqueur and Bitter Compounds.

We evaluated sex differences in the perception of bitter compounds and an aromatic bitter herbal liqueur (Mirtamaro) obtained by the infusion of myrtle leaves/berries together with a mixture of Mediterranean herbs/plants as flavoring/bittering ingredients. In a healthy population (n = 231 participants), using bivariate correlations and multivariate linear regression analyses, significant sex differences emerged in quinine bitterness perception, with women showing a higher bitter taste intensity rating than men. Among all participants, 40 subjects (subpopulation) were randomly selected for the evaluation of sex differences in Mirtamaro gustatory and olfactory perception using a hedonic Likert-type scale. Women showed higher ratings in Mirtamaro aroma (odor intensity) and bitterness (taste intensity) perception than men, with a superior capacity to perceive/describe its sensory attributes. 1,8-Cineole and methyl chavicol were the main contributors to the bitter liqueur aroma. A significant correlation (r = 0.564, p < 0.01) between Mirtamaro odor pleasantness/taste pleasantness was observed in women, indicating a positive contribution of aromatic herbs to bitter taste acceptability. Moreover, a higher bitter intensity rating of 6-n-propylthiouracil was evidenced in women than men. Our results highlighted sex differences in bitter taste acuity and the role of aromatic herbs/plants in modulating bitter taste acceptance, which is useful information in the field of precision nutrition and medicine.

Effect of thermal processing on debittering process and its impact on the functional properties of palmyrah young shoot flour

Palmyrah young shoot is rich in starch and fiber and provides good glycemic control. Unfortunately, the consumption rate is limited by the presence of bitter and toxic compounds. Reported literatures reveal that thermal treatment especially dry heat processes like milling the flour at more than 80 °C had an impact on toxicity reduction rather than wet heat processes like boiling. As a result, the current study was aimed to investigate the combined effect of wet (boiling) and dry (radio frequency) heat processing on the debittering of young shoot flour in comparison with tray drying. In the study the flour obtained by a combination of boiling and drying was considered as treated flour and without boiling and drying was considered as untreated flour. The combined effect of wet and dry heat processing had an impact on the nutrients but had no effect on bitterness reduction. The study found tray dried untreated flour favored the feasibility of bitterness reduction as compared to other samples. The compounds (FlabelliferinsB, Flabelliferins–II, isomers of flabelliferins, and vitamin C) responsible for bitterness and its intensity were identified by FTMS. Furthermore, the study revealed that debittering process improved the functional properties of young shoot flour compared to raw flour.