Metabolic Engineering of Candida tropicalis for the De Novo Synthesis of ß-Ionone.

ß-ionone, a norisoprenoid, is a natural aromatic compound derived from plants, which displays various biological activities including anticancer, antioxidant and deworming properties. Due to its large biomass and strong environmental tolerance, the nonconventional oleaginous yeast Candida tropicalis was selected to efficiently synthesize ß-ionone. We initially investigated the capacity of the cytoplasm and subcellular compartments to synthesize ß-ionone independently. Subsequently, through adaptive screening of enzymes, functional identification of subcellular localization signal peptides and subcellular compartment combination strategies, a titer of 152.4 mg/L of ß-ionone was achieved. Finally, directed evolution of rate-limiting enzyme and overexpression of key enzymes were performed to enhance ß-ionone production. The resulting titer was 400.5 mg/L in shake flasks and 730 mg/L in a bioreactor. This study demonstrates the first de novo synthesis of ß-ionone in C. tropicalis, providing a novel cellular chassis for terpenoid fragrances with considerable industrial potential.

A Food Odorant, α-Ionone, Inhibits Skin Cancer Tumorigenesis by Activation of OR10A6.

SCOPE: This study aims to investigate the anticancer properties of α-ionone in squamous cell carcinoma (SCC). METHODS AND RESULTS: The expression of OR10A6 together with olfactory receptor signaling components is demonstrated in A431 human SCC cells via RT-PCR and qRT-PCR analysis. OR10A6 activation in A431 cells using the ligand α-ionone inhibits proliferation and migration but induces apoptosis which is confirmed by proliferation assay, colony formation, and western blotting. The mechanism involves the core proteins of the Hippo pathway, where the phosphorylation of large tumor suppressor kinase (LATS), yes-associated protein (YAP), and transcriptional coactivator with PDZ-binding motif (TAZ) is confirmed by western blotting. However, the anticancer effects of α-ionone are abrogated in A431 cells with OR10A6 gene knockdown. In A431 xenograft mouse model, the injection of α-ionone suppresses tumor growth, induces apoptosis, and increases phosphorylation of the LATS-YAP-TAZ signaling axis in the Hippo pathway. None of these effects are observed in xenografted tumors with OR10A6 gene knockdown. CONCLUSION: These findings collectively demonstrate that activation of ectopic OR OR10A6 by α-ionone in SCC cells stimulates the Hippo pathway and suppresses tumorigenesis both in vitro and in vivo, suggesting a novel therapeutic candidate for the treatment of SCC.

Ionones from cigar tobacco leaves.

Phytochemical studies on cigar tobacco leaves led to the isolation of 18 ionone-type compounds, including previously undescribed cigatobanes E (1) and F (2). Additionally, compounds vomifoliol acetate (3), dehydrovomifoliol (4), 8,9-dihydromegastigmane-4,6-diene-3-one (5), 7α,8α-epoxyblumenol B (6), 3-oxoactinidol (12), and loliolide acetate (15), 4ß-hydroxy-dihydroactinidiolide (17), were found in tobacco leaves for the first time. The structural elucidation of all compounds was accomplished through rigorous spectral analysis.

Three New Ionone Glycosides from Rhododendron capitatum Maxim.

Six ionone glycosides (1-3 and 5-7), including three new ones, named capitsesqsides A-C (1-3), together with an eudesmane sesquiterpenoid glycoside (4) and three known triterpenoid saponins (8-10) were isolated from Rhododendron capitatum. The structures of these compounds were determined by extensive spectroscopic techniques (MS, UV, 1D-NMR, and 2D-NMR) and comparison with data reported in the literature. The absolute configurations were determined by comparison of the experimental and theoretically calculated ECD curves and LC-MS analyses after acid hydrolysis and derivatization. The anti-inflammatory activities of these compounds were evaluated in the LPS-induced RAW264.7 cells. Molecular docking demonstrated that 2 has a favorable affinity for NLRP3 and iNOS.

Megastigmane glycosides from the traditional Uighur medicine Cydonia oblonga Mill.

Phytochemical investigation on the fruits of Cydonia oblonga Mill., a traditional Uighur medicine, led to the isolation of seven undescribed and nine known megastigmane glycosides. Their structures including absolute configurations were characterized by an extensive analysis of spectroscopic data including HRESIMS and NMR, combined with ECD calculations. Additionally, compounds 1, 2, 4, and 6-16 exhibited anti-inflammatory activity by inhibiting the secretion of cytokines TNF-α and IL-6 in RAW264.7 cells induced by lipopolysaccharides (LPS) with inhibitory rates of 10.79%-44.58% at 20 µM.

Effects of natural Ionones and derived novel analogues with simplified structures on behavioral responses of whiteflies.

BACKGROUND: Whiteflies are major pests in agriculture, causing damage to crops and transmitting plant viruses. Using Volatile Organic Compounds (VOCs) as semiochemicals offers a sustainable approach for combating whiteflies. One such group of compounds, represented by ß-ionone, has been found to possess repellent/attractant properties. To further explore the behavioral effects of these compounds on whiteflies, we selected five natural ionone compounds and synthesized six novel analogues to examine the impact of structural variations on whitefly behavior. RESULTS: Our results demonstrated that ß-ionone and its analogues have a significant impact on the behavior of whiteflies. Among them, 0.01% pseudo ionone solution exhibited an attractant effect on whiteflies. Notably, the application of 1% ß-ionone and 0.1% ß-ionol solution demonstrated a notable repellent effect and oviposition deterrent effect on whitefly. We also found that the novel ionone analogue (±)1A exhibited a strong repellent effect. Both ß-ionol and compound (±)1A possess high logP values and low saturation vapor pressures, which contribute to enhanced lipophilicity, making them more likely to penetrate insect antennae and prolong their presence in the air. CONCLUSION: The newly discovered ionone analogue (±)1A and ß-ionol exhibit improved repellent effects, while pseudo ionone shows an attractant effect. These three compounds hold promising potential for development as novel biological control agents. Our work highlights the efficacy of VOCs as a protection method against whiteflies. These findings indicate that our new technology for a 'push-pull' control method of B. tabaci can offer a novel tool for integrated pest management (IPM). © 2024 Society of Chemical Industry.

Correlation study on microbial communities and volatile flavor compounds in cigar tobacco leaves of diverse origins.

To elucidate the significant influence of microorganisms on geographically dependent flavor formation by analyzing microbial communities and volatile flavor compounds (VFCs) in cigar tobacco leaves (CTLs) obtained from China, Dominica, and Indonesia. Microbiome analysis revealed that the predominant bacteria in CTLs were Staphylococcus, Aerococcus, Pseudomonas, and Lactobacillus, while the predominant fungi were Aspergillus, Wallemia, and Sampaiozyma. The microbial communities of CTLs from different origins differed to some extent, and the diversity and abundance of bacteria were greater than fungi. Metabolomic analysis revealed that 64 VFCs were identified, mainly ketones, of which 23 VFCs could be utilized to identify the geographical origins of CTLs. Sixteen VFCs with OAV greater than 1, including cedrol, phenylacetaldehyde, damascone, beta-damascone, and beta-ionone, play important roles in shaping the flavor profile of CTLs from different origins. Combined with the correlation analysis, bacterial microorganisms were more closely related to key VFCs and favored a positive correlation. Bacillus, Vibrio, and Sphingomonas were the main flavor-related bacteria. The study demonstrated that the predominant microorganisms were essential for the formation of key flavor qualities in CTLs, which provided a theoretical reference for flavor control of CTLs by microbial technology. KEY POINTS: • It is the high OAV VFCs that determine the flavor profile of CTLs. • The methylerythritol phosphate (MEP) pathway and the carotenoid synthesis pathway are key metabolic pathways for the formation of VFCs in CTLs. • Microbial interactions influence tobacco flavor, with bacterial microorganisms contributing more to the flavor formation of CTLs.

Exploration of Raw Pigmented-Fleshed Sweet Potatoes Volatile Organic Compounds and the Precursors.

Sweet potato provides rich nutrients and bioactive substances for the human diet. In this study, the volatile organic compounds of five pigmented-fleshed sweet potato cultivars were determined, the characteristic aroma compounds were screened, and a correlation analysis was carried out with the aroma precursors. In total, 66 volatile organic compounds were identified. Terpenoids and aldehydes were the main volatile compounds, accounting for 59% and 17%, respectively. Fifteen compounds, including seven aldehydes, six terpenes, one furan, and phenol, were identified as key aromatic compounds for sweet potato using relative odor activity values (ROAVs) and contributed to flower, sweet, and fat flavors. The OR sample exhibited a significant presence of trans-ß-Ionone, while the Y sample showed high levels of benzaldehyde. Starch, soluble sugars, 20 amino acids, and 25 fatty acids were detected as volatile compounds precursors. Among them, total starch (57.2%), phenylalanine (126.82 ± 0.02 g/g), and fatty acids (6.45 µg/mg) were all most abundant in Y, and LY contained the most soluble sugar (14.65%). The results of the correlation analysis revealed the significant correlations were identified between seven carotenoids and trans-ß-Ionone, soluble sugar and nerol, two fatty acids and hexanal, phenylalanine and 10 fatty acids with benzaldehyde, respectively. In general, terpenoids and aldehydes were identified as the main key aromatic compounds in sweet potatoes, and carotenoids had more influence on the aroma of OR than other cultivars. Soluble sugars, amino acids, and fatty acids probably serve as important precursors for some key aroma compounds in sweet potatoes. These findings provide valuable insights for the formation of sweet potato aroma.

Dietary supplementation with α-ionone alleviates chronic UVB exposure-induced skin photoaging in mice.

Photoaging is widely regarded as the most significant contributor to skin aging damage. It is triggered by prolonged exposure to ultraviolet (UV) light and typically manifests as dryness and the formation of wrinkles. Nutritional intervention is a viable strategy for preventing and treating skin photoaging. In previous studies, we demonstrated that α-ionone had ameliorating effects on photoaging in both epidermal keratinocytes and dermal fibroblasts. Here, we investigated the potential anti-photoaging effects of dietary α-ionone using a UVB-irradiated male C57BL/6N mouse model. Our findings provided compelling evidence that dietary α-ionone alleviates wrinkle formation, skin dryness, and epidermal thickening in chronic UVB-exposed mice. α-Ionone accumulated in mouse skin after 14 weeks of dietary intake of α-ionone. α-Ionone increased collagen density and boosted the expression of collagen genes, while attenuating the UVB-induced increase of matrix metalloproteinase genes in the skin tissues. Furthermore, α-ionone suppressed the expression of senescence-associated secretory phenotypes and reduced the expression of the senescence marker p21 and DNA damage marker p53 in the skin of UVB-irradiated mice. Transcriptome sequencing results showed that α-ionone modifies gene expression profiles of skin. Multiple pathway enrichment analyses on both the differential genes and the entire genes revealed that α-ionone significantly affects multiple physiological processes and signaling pathways associated with skin health and diseases, of which the p53 signaling pathway may be the key signaling pathway. Taken together, our findings reveal that dietary α-ionone intervention holds promise in reducing the risks of skin photoaging, offering a potential strategy to address skin aging concerns.

Genomic analysis reveals genes that encode the synthesis of volatile compounds by a Bacillus velezensis-based biofungicide used in the treatment of grapes to control Aspergillus carbonarius.

Fungal control strategies based on the use of Bacillus have emerged in agriculture as eco-friendly alternatives to replace/reduce the use of synthetic pesticides. Bacillus sp. P1 was reported as a new promising strain for control of Aspergillus carbonarius, a known producer of ochratoxin A, categorized as possible human carcinogen with high nephrotoxic potential. Grape quality can be influenced by vineyard management practices, including the use of fungal control agents. The aim of this study was to evaluate, for the first time, the quality parameters of Chardonnay grapes exposed to an antifungal Bacillus-based strategy for control of A. carbonarius, supporting findings by genomic investigations. Furthermore, genomic tools were used to confirm that the strain P1 belongs to the non-pathogenic species Bacillus velezensis and also to certify its biosafety. The genome of B. velezensis P1 harbors genes that are putatively involved in the production of volatiles and hydrolytic enzymes, which are responsible for releasing the free form of aroma compounds. In addition to promote biocontrol of phytopathogenic fungi and ochratoxins, the treatment with B. velezensis P1 did not change the texture (hardness and firmness), color and pH of the grapes. Heat map and hierarchical clustering analysis (HCA) of volatiles evaluated by GC/MS revealed that Bacillus-treated grapes showed higher levels of compounds with a pleasant odor descriptions such as 3-hydroxy-2-butanone, 2,3-butanediol, 3-methyl-1-butanol, 3,4-dihydro-ß-ionone, ß-ionone, dihydroactinidiolide, linalool oxide, and ß-terpineol. The results of this study indicate that B. velezensis P1 presents desirable properties to be used as a biocontrol agent.

Molecular mechanisms and evolutionary robustness of a color switch in proteorhodopsins.

Proteorhodopsins are widely distributed photoreceptors from marine bacteria. Their discovery revealed a high degree of evolutionary adaptation to ambient light, resulting in blue- and green-absorbing variants that correlate with a conserved glutamine/leucine at position 105. On the basis of an integrated approach combining sensitivity-enhanced solid-state nuclear magnetic resonance (ssNMR) spectroscopy and linear-scaling quantum mechanics/molecular mechanics (QM/MM) methods, this single residue is shown to be responsible for a variety of synergistically coupled structural and electrostatic changes along the retinal polyene chain, ionone ring, and within the binding pocket. They collectively explain the observed color shift. Furthermore, analysis of the differences in chemical shift between nuclei within the same residues in green and blue proteorhodopsins also reveals a correlation with the respective degree of conservation. Our data show that the highly conserved color change mainly affects other highly conserved residues, illustrating a high degree of robustness of the color phenotype to sequence variation.

Floral Volatile Organic Compounds Change the Composition and Function of the Endophytic Fungal Community in the Flowers of Osmanthus fragrans.

Endophytic fungi in flowers influence plant health and reproduction. However, whether floral volatile organic compounds (VOCs) affect the composition and function of the endophytic fungal community remains unclear. Here, gas chromatography-mass spectrometry (GC-MS) and high-throughput sequencing were used to explore the relationship between floral VOCs and the endophytic fungal community during different flower development stages in Osmanthus fragrans 'Rixiang Gui'. The results showed that the composition of the endophytic fungal community and floral VOCs shifted along with flowering development. The highest and lowest α diversity of the endophytic fungal community occurred in the flower fading stage and full blooming stage, respectively. The dominant fungi, including Dothideomycetes (class), Pleosporales (order), and Neocladophialophora, Alternaria, and Setophoma (genera), were enriched in the flower fading stage and decreased in the full blooming stage, demonstrating the enrichment of the Pathotroph, Saprotroph, and Pathotroph-Saprotroph functions in the flower fading stage and their depletion in the full blooming stage. However, the total VOC and terpene contents were highest in the full blooming stage and lowest in the flower fading stage, which was opposite to the α diversity of the endophytic fungal community and the dominant fungi during flowering development. Linalool, dihydro-ß-ionone, and trans-linalool oxide(furan) were key factors affecting the endophytic fungal community composition. Furthermore, dihydro-ß-ionone played an extremely important role in inhibiting endophytic fungi in the full blooming stage. Based on the above results, it is believed that VOCs, especially terpenes, changed the endophytic fungal community composition in the flowers of O. fragrans 'Rixiang Gui'. These findings improve the understanding of the interaction between endophytic fungi and VOCs in flowers and provide new insight into the mechanism of flower development.

Effects of Monascus purpureus on ripe Pu-erh tea in different fermentation methods and identification of characteristic volatile compounds.

The present study aimed to explore the key volatile compounds (VCs) that lead to the formation of characteristic flavors in ripe Pu-erh tea (RIPT) fermented by Monascus purpureus (M. purpureus). Headspace solid-phase microextraction coupled with gas chromatography/mass spectrometry (HS-SPME-GC-MS), orthogonal partial least square-discriminant analysis (OPLS-DA) were employed for a comprehensive analysis of the VCs present in RIPT fermented via different methods and were further identified by odor activity value (OAV). The VCs 1,2-dimethoxybenzene, 1,2,3-trimethoxybenzene, (E)-linalool oxide (pyranoid), methyl salicylate, linalool, ß-ionone, ß-damascenone were the key characteristic VCs of RIPT fermented by M. purpureus. OAV and Gas chromatography-olfactometry (GC-O) further indicated that ß-damascenone was the highest contribution VCs to the characteristic flavor of RIPT fermented by M. purpureus. This study reveals the specificities and contributions of VCs present in RIPT under different fermentation methods, thus providing new insights into the influence of microorganisms on RIPT flavor.

De Novo Synthesis of Dihydro-ß-ionone through Metabolic Engineering and Bacterium-Yeast Coculture.

Dihydro-ß-ionone is a common type of ionone used in the flavor and fragrance industries because of its characteristic scent. The production of flavors in microbial cell factories offers a sustainable and environmentally friendly approach to accessing them, independent of extraction from natural sources. However, the native pathway of dihydro-ß-ionone remains unclear, hindering heterologous biosynthesis in microbial hosts. Herein, we devised a microbial platform for de novo syntheses of dihydro-ß-ionone from a simple carbon source with glycerol. The complete dihydro-ß-ionone pathway was reconstructed in Escherichia coli with multiple metabolic engineering strategies to generate a strain capable of producing 8 mg/L of dihydro-ß-ionone, although this was accompanied by a surplus precursor ß-ionone in culture. To overcome this issue, Saccharomyces cerevisiae was identified as having a conversion rate for transforming ß-ionone to dihydro-ß-ionone that was higher than that of E. coli via whole-cell catalysis. Consequently, the titer of dihydro-ß-ionone was increased using the E. coli-S. cerevisiae coculture to 27 mg/L. Our study offers an efficient platform for biobased dihydro-ß-ionone production and extends coculture engineering to overproducing target molecules in extended metabolic pathways.

Integration of rice apocarotenoid profile and expression pattern of Carotenoid Cleavage Dioxygenases reveals a positive effect of ß-ionone on mycorrhization.

Carotenoids are susceptible to degrading processes initiated by oxidative cleavage reactions mediated by Carotenoid Cleavage Dioxygenases that break their backbone, leading to products called apocarotenoids. These carotenoid-derived metabolites include the phytohormones abscisic acid and strigolactones, and different signaling molecules and growth regulators, which are utilized by plants to coordinate many aspects of their life. Several apocarotenoids have been recruited for the communication between plants and arbuscular mycorrhizal (AM) fungi and as regulators of the establishment of AM symbiosis. However, our knowledge on their biosynthetic pathways and the regulation of their pattern during AM symbiosis is still limited. In this study, we generated a qualitative and quantitative profile of apocarotenoids in roots and shoots of rice plants exposed to high/low phosphate concentrations, and upon AM symbiosis in a time course experiment covering different stages of growth and AM development. To get deeper insights in the biology of apocarotenoids during this plant-fungal symbiosis, we complemented the metabolic profiles by determining the expression pattern of CCD genes, taking advantage of chemometric tools. This analysis revealed the specific profiles of CCD genes and apocarotenoids across different stages of AM symbiosis and phosphate supply conditions, identifying novel reliable markers at both local and systemic levels and indicating a promoting role of ß-ionone in AM symbiosis establishment.

Undescribed compounds from Clinopodium chinense (Benth.) O. Kuntze and their coagulation activity studies.

Four rare compounds (1-4), including one 1,4-epoxy-benzoxepane derivative and one ringed prenylated naphthoquinoid skeleton, as well as one isopimarane-type diterpenoid and one megastigmane-type glycoside, along with three known megastigmane-type glycosides (5-7) were isolated from the ethanol extracts of C. chinense. Their structures were determined on the basis of 1D, 2D NMR, HR-ESI-MS and DP4+ analysis. Meanwhile, the in vitro evaluation indicated that compound 2 and 6 exhibited excellent procoagulant activities, which can significantly shorten prothrombin time (PT) and activated partial thromboplastin time (APTT), respectively.

The apocarotenoid ß-ionone regulates the transcriptome of Arabidopsis thaliana and increases its resistance against Botrytis cinerea.

Carotenoids are isoprenoid pigments indispensable for photosynthesis. Moreover, they are the precursor of apocarotenoids, which include the phytohormones abscisic acid (ABA) and strigolactones (SLs) as well as retrograde signaling molecules and growth regulators, such as ß-cyclocitral and zaxinone. Here, we show that the application of the volatile apocarotenoid ß-ionone (ß-I) to Arabidopsis plants at micromolar concentrations caused a global reprogramming of gene expression, affecting thousands of transcripts involved in stress tolerance, growth, hormone metabolism, pathogen defense, and photosynthesis. This transcriptional reprogramming changes, along with induced changes in the level of the phytohormones ABA, jasmonic acid, and salicylic acid, led to enhanced Arabidopsis resistance to the widespread necrotrophic fungus Botrytis cinerea (B.c.) that causes the gray mold disease in many crop species and spoilage of harvested fruits. Pre-treatment of tobacco and tomato plants with ß-I followed by inoculation with B.c. confirmed the effect of ß-I in increasing the resistance to this pathogen in crop plants. Moreover, we observed reduced susceptibility to B.c. in fruits of transgenic tomato plants overexpressing LYCOPENE ß-CYCLASE, which contains elevated levels of endogenous ß-I, providing a further evidence for its effect on B.c. infestation. Our work unraveled ß-I as a further carotenoid-derived regulatory metabolite and indicates the possibility of establishing this natural volatile as an environmentally friendly bio-fungicide to control B.c.

Cyanobacterial VOCs ß-ionone and ß-cyclocitral poisoning Lemna turionifera by triggering programmed cell death.

ß-Ionone and ß-cyclocitral are two typical components in cyanobacterial volatiles, which can poison aquatic plants and even cause death. To reveal the toxic mechanisms of the two compounds on aquatic plants through programmed cell death (PCD), the photosynthetic capacities, caspase-3-like activity, DNA fragmentation and ladders, as well as expression of the genes associated with PCD in Lemna turionifera were investigated in exposure to ß-ionone (0.2 mM) and ß-cyclocitral (0.4 mM) at lethal concentration. With prolonging the treatment time, L. turionifera fronds gradually died, and photosynthetic capacities gradually reduced and even disappeared at the 96th h. This demonstrated that the death process might be a PCD rather than a necrosis, due to the gradual loss of physiological activities. When L. turionifera underwent the death, caspase-3-like was activated after 3 h, and reached to the strongest activity at the 24th h. TUNEL-positive nuclei were detected after 12 h, and appeared in large numbers at the 48th h. The DNA was cleaved by Ca2+-dependent endonucleases and showed obviously ladders. In addition, the expression of 5 genes (TSPO, ERN1, CTSB, CYC, and ATR) positively related with PCD initiation was up-regulated, while the expression of 2 genes (RRM2 and TUBA) negatively related with PCD initiation was down-regulated. Therefore, ß-ionone and ß-cyclocitral can poison L. turionifera by adjusting related gene expression to trigger PCD.

Organ-specific volatiles from Sonoran desert Krameria flowers as potential signals for oil-collecting bees.

The evolution of flowers that offer oils as rewards and are pollinated by specialized bees represents a distinctive theme in plant-pollinator co-diversification. Some plants that offer acetylated glycerols as floral oils emit diacetin, a volatile by-product of oil metabolism, which is utilized by oil-collecting bees as an index signal for the presence of floral oil. However, floral oils in the genus Krameria (Krameriaceae) contain ß-acetoxy-substituted fatty acids instead of acetylated glycerols, making them unlikely to emit diacetin as an oil-bee attractant. We analyzed floral headspace composition from K. bicolor and K. erecta, native to the Sonoran Desert of southwestern North America, in search of alternative candidates for volatile index signals. Using solid-phase microextraction, combined with gas chromatography-mass spectrometry, we identified 26 and 45 floral volatiles, respectively, from whole flowers and dissected flower parts of these two Krameria species. As expected, diacetin was not detected. Instead, ß-ionone emerged as a strong candidate for an index signal, as it was uniquely present in dissected oil-producing floral tissues (elaiophores) of K. bicolor, as well as the larval cells and provisions from its oil-bee pollinator, Centris cockerelli. This finding suggests that the floral oil of K. bicolor is perfused with ß-ionone in its tissue of origin and retains the distinctive raspberry-like scent of this volatile after being harvested by C. cockerelli bees. In contrast, the elaiophores of K. erecta, which are not thought to be pollinated by C. cockerelli, produced a blend of anise-related oxygenated aromatics not found in the elaiophores of K. bicolor. Our findings suggest that ß-ionone has the potential to impact oil-foraging by C. cockerelli bees through several potential mechanisms, including larval imprinting on scented provisions or innate or learned preferences by foraging adults.

Study of Glycosidically Bound Volatile Precursors as Variety Markers to Reveal Not-Allowed Practices in White Wines Winemaking.

Liquid chromatography/high-resolution mass spectrometry (LC/HRMS) can provide identification of grape metabolites which are variety markers. White grapes are poorer in polyphenolics, and the main secondary metabolites which contribute the sensorial characteristics of wines are the glycosidically bound volatile precursors and their aglycones. The profiles of three white grape juices (Pinot grigio, Garganega, and Trebbiano) were characterized by LC/HRMS, and 70 signals of putative glycosidic terpenols, norisoprenoids, and benzenoids were identified. Four signals found only in Pinot grigio corresponded to a norisoprenoid hexose-hexose, 3-oxo-α-ionol (or 3-hydroxy-ß-damascone) rhamnosyl-hexoside, monoterpene-diol hexosyl-pentosyl-hexoside, and hexose-norisoprenoid; three signals were found only in Garganega (putative isopropyl alcohol pentosyl-hexoside, phenylethanol rhamnosyl-hexoside, and norisoprenoid hexose-hexose isomers), and a monoterpenol pentosyl-hexoside isomer only in Trebbiano. These variety markers were then investigated in juice blends of the three varieties. This approach can be used to develop control methods to reveal not-allowed grape varieties and practices in white wines winemaking.