Limonoid Aglycones Research Articles

Assessment of the in vitro anti-diabetic activity with molecular dynamic simulations of limonoids isolated from Adalia lemon peels

Limonoids are important constituents of citrus that have a significant impact on promoting human health. Therefore, the primary focus of this research was to assess the overall limonoid content and isolate limonoids from Adalia lemon (Citrus limon L.) peels for their potential use as antioxidants and anti-diabetic agents. The levels of limonoid aglycones in the C. limon peel extract were quantified through a colorimetric assay, revealing a concentration of 16.53 ± 0.93 mg/L limonin equivalent. Furthermore, the total concentration of limonoid glucosides was determined to be 54.38 ± 1.02 mg/L. The study successfully identified five isolated limonoids, namely limonin, deacetylnomilin, nomilin, obacunone 17-O-β-D-glucopyranoside, and limonin 17-O-β-D-glucopyranoside, along with their respective yields. The efficacy of the limonoids-rich extract and the five isolated compounds was evaluated at three different concentrations (50, 100, and 200 µg/mL). It was found that both obacunone 17-O-β-D-glucopyranoside and limonin 17-O-β-D-glucopyranoside possessed the highest antioxidant, free radical scavenging, and anti-diabetic activities, followed by deacetylnomilin, and then the limonoids-rich extract. The molecular dynamic simulations were conducted to predict the behavior of the isolated compounds upon binding to the protein's active site, as well as their interaction and stability. The results revealed that limonin 17-O-β-D-glucopyranoside bound to the protein complex system exhibited a relatively more stable conformation than the Apo system. The analysis of Solvent Accessible Surface Area (SASA), in conjunction with the data obtained from Root-Mean-Square Deviation (RMSD), Root-Mean-Square Fluctuation (RMSF), and Radius of Gyration (ROG) computations, provided further evidence that the limonin 17-O-β-D-glucopyranoside complex system remained stable within the catalytic domain binding site of the human pancreatic alpha-amylase (HPA)-receptor. The research findings suggest that the limonoids found in Adalia lemon peels have the potential to be used as effective natural substances in creating innovative therapeutic treatments for conditions related to oxidative stress and disorders in carbohydrate metabolism.

Nomilin from Yuzu Seed Has In Vitro Antioxidant Activity and Downregulates Melanogenesis in B16F10 Melanoma Cells through the PKA/CREB Signaling Pathway.

Yuzu (Citrus junos) is a citrus plant native to Asian countries, including Korea, Japan, and China. Yuzu peel and seed contain abundant vitamin C, citric acid, and polyphenols. Although the antioxidative and antimelanogenic activities of other citrus fruits and yuzu extract have been reported, the tyrosinase inhibitory activity of the limonoid aglycone contained in yuzu seed extract is unknown. We separated yuzu seeds into the husk, shell, and meal and evaluated antioxidant activity of each. The limonoid glucoside fraction of the husk identified nomilin, a novel tyrosinase inhibitor. We performed tyrosinase inhibitory activity and noncompetitive inhibition assays and docking studies to determine nomilin binding sites. Furthermore, we evaluated the antioxidative mechanism and antimelanogenic activity of nomilin in B16F10 melanoma cells. The concentration of nomilin that did not show toxicity was <100 µg/mL. Nomilin suppressed protein expression of TYR, TRP-1, TRP-2, and microphthalmia-associated transcription factor (MITF) in a concentration-dependent manner. Nomilin significantly reduced the levels of p-CREB and p-PKA at the protein level and decreased the levels of skin-whitening-related factors MITF, tyrosinase, TRP-1, and TRP-2 at the mRNA level in a concentration-dependent manner. Thus, nomilin from yuzu seed husk can be used as a skin-whitening agent in cosmetics.

A Targeted Metabolomics Approach to Study Secondary Metabolites and Antioxidant Activity in 'Kinnow Mandarin' during Advanced Fruit Maturity.

In this study, we investigated the impact of harvest maturity stages and contrasting growing climates on secondary metabolites in Kinnow mandarin. Fruit samples were harvested at six harvest maturity stages (M1–M6) from two distinct growing locations falling under subtropical–arid (STA) and subtropical–humid (STH) climates. A high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) technique was employed to identify and quantify secondary metabolites in the fruit juice. A total of 31 polyphenolics and 4 limonoids, with significant differences (p < 0.05) in their concentration, were determined. With advancing maturity, phenolic acids and antioxidant activity were found to increase, whereas flavonoids and limonoids decreased in concentration. There was a transient increase in the concentration of some polyphenolics such as hesperidin, naringin, narirutin, naringenin, neoeriocitrin, rutin, nobiletin and tangeretin, and limonoid aglycones such as limonin and nomilin at mid-maturity stage (M3) which coincided with prevailing low temperature and frost events at growing locations. A higher concentration of limonin and polyphenolics was observed for fruit grown under STH climates in comparison to those grown under STA climates. The data indicate that fruit metabolism during advanced stages of maturation under distinct climatic conditions is fundamental to the flavor, nutrition and processing quality of Kinnow mandarin. This information can help in understanding the optimum maturity stage and preferable climate to source fruits with maximum functional compounds, less bitterness and high consumer acceptability.

The constitutions of terpenoids and hormones of leaves in red-flesh citrus mutants and their wild types

To study the profiles and relationships of terpenoids and phytohormones in leaves of red-flesh citrus, we examined four red-flesh mutants that accumulate lycopene in the fruit and their corresponding wild types. We compared their contents of monoterpenoid and sesquiterpenoid volatiles, triterpenoid limonoid aglycones, and tetraterpenoid carotenoids. The types and concentrations of these terpenoids in the leaves varied among genotypes, and the leaf terpenoid profiles differed between red-flesh mutants and their wild types. However, lycopene was not detected in citrus leaves of all eight varieties, including the four mutants. According to phytohormone accumulation in the leaves, the citrus varieties could be classified into a low-phytohormone group (‘Red Anliu’ sweet orange, ‘Anliu’ sweet orange, and ‘Cara Cara’ navel orange), a high-phytohormone group (‘Seike’ navel orange), and a high-jasmonic acid group (‘Red-flesh Guanxi’ pummelo, ‘Guanxi’ pummelo, ‘Chuhong’ pummelo, and ‘Feicui’ pummelo). The contents of terpenoid volatiles were very low in leaves of ‘Red-flesh Guanxi’ and ‘Guanxi’ pummelo; therefore, they are ideal materials for verifying the functions of genes related to terpenoid volatiles in overexpression analyses. Sesquiterpenoid volatiles were positively correlated with phytohormones of abscisic acid, jasmonic acid, and salicylic acid. Taken together, lycopene was undetected in the leaves of red-flesh citrus mutants with abundant lycopene in their juice sacs, which suggested a tissue-specific accumulation pattern of lycopene in citrus red-flesh mutants. Furthermore, leaf volatile profiles could be suitable to develop fingerprint chromatograms of citrus resources, since leaf volatile profiles were dominated by several compounds that varied among genotypes in their combinations and concentrations.

Lycopene Accumulation in Cara Cara Red-flesh Navel Orange Is Correlated with Weak Abscisic Acid Catabolism.

Lycopene is the main pigment in red-flesh citrus fruits, and its formation is a research hotspot. To explore the basis of lycopene accumulation in red-flesh mutants, we profiled the terpenoid metabolites. Compared with their respective wild types, Cara Cara (Cara) [and Red-Anliu (R-An)] oranges showed increased carotenoid and limonoid aglycone contents and decreased contents of abscisic acid (ABA) catabolites, monoterpenoid volatiles, and sesquiterpenoid volatiles. Cara contained less than half of the amount of ABA glucose ester (ABAGE), the main ABA derivative in oranges. Parallel lower transcript levels of NCED and ABA glucosyltransferase in Cara were detected at the mature green stage. These results document the changes in terpenoid profiles in Cara and show that the red flesh of citrus color mutants is related to weak ABA catabolism, especially ABAGE, and decreased transcript levels of two genes encoding uridine diphosphate (UDP)-glycosyltransferases that catalyze ABAGE biosynthesis.

Identification and quantification of limonoid aglycones content of Citrus seeds

Citrus fruits are recognized as an important source of bioactive molecules such as limonin and nomilin. However, these molecules exhibit low bioavailability, therefore, obtaining these molecules using biotechnological techniques may be an alternative to harvesting them directly from fruits. The aim of this study was to quantify and identify limonoids in the dichloromethane extracts of Citrus seeds of Criolla orange, Oneco tangerine, Tangerine-lemon, Sour orange and Valencia orange from department of Antioquia-Colombia by high performance liquid chromatography with diode array detection, and high-resolution mass spectrometry. Although in all the samples total glycosidic free limonoids were present, Oneco tangerine seeds had the highest concentration, followed by Tangerine-lemon seeds, equivalent to 0.75% and 0.53% per total dry weight, respectively. These results suggest Oneco tangerine seeds may be used as an elite material for biotechnological processes looking for increased production of limonoids to support research and drug development.

The Effect of Processing Methods on Phytochemical Composition in Bergamot Juice.

Experimental and epidemiological studies show a positive relation between consumption of citrus juices and reduction of risk for some chronic disorders, such as diabetes and cardiovascular diseases. In particular, the bergamot juice is characterized by noticeable amounts of phytochemicals such as flavanone glycosides, limonoids, and quaternary ammonium compounds, all health-beneficial biomolecules. In vitro and in vivo studies have shown anti-inflammatory, cholesterol-lowering, and anti-diabetic activities attributed to these compounds depending on their chemical structure. However, nutritional content of bergamot juice may vary as consequence of different processing techniques, thus needing to address this claim. For this reason, the objective of this research was to evaluate the effects of different processing systems on the proximate constituents, the composition, and the antioxidant activity of the correspondent juices. Overall, the results indicate that the process employed may influence the chemical composition and the functional properties of the ended juice. Screw press method produced a juice with greater content of flavanone glycosides (ranged from 37 to 402 mg/L) and limonoid aglycones (ranged from 65 to 67 mg/L) than the other processes (p < 0.001). However, the process used for extraction of bergamot juice did not affect significantly the N,N-dimethyl-L-proline content (p < 0.5). Moreover, the screw press juice showed the highest antioxidant activity with EC50 value of 9.35 µg/mL, thus suggesting that this method maintains for health the nutritional quality of a fresh-pressed juice.

Integrative Analysis of Terpenoid Profiles and Hormones from Fruits of Red-Flesh Citrus Mutants and Their Wild Types.

In citrus color mutants, the levels of carotenoid constituents and other secondary metabolites are different in their corresponding wild types. Terpenoids are closely related to coloration, bitterness, and flavor. In this study, terpenoid profiles and hormones in citrus fruits of two red-flesh mutants—Red Anliu orange and Red-flesh Guanxi pummelo—and their corresponding wild types were investigated using GC/MS, HPLC, and LC-MS/MS. Results showed that Red Anliu orange (high in carotenoids) and Anliu orange (low in carotenoids) accumulated low levels of limonoid aglycones but high levels of monoterpenoids; conversely, Red-flesh Guanxi pummelo (high in carotenoids) and Guanxi pummelo (deficient in carotenoids) accumulated high levels of limonoid aglycones but low levels of monoterpenoids. However, isopentenyl diphosphate was present at similar levels. A correlation analysis indicated that jasmonic and salicylic acids might play important roles in regulating terpenoid biosynthesis. Additionally, the similarities of carotenoid and volatile profiles between each mutant and its corresponding wild type were greater than those between the two mutants or the two wild types. The flux balance of terpenoid metabolism in citrus fruit tends toward stability among various citrus genera that have different terpenoid profiles. Bud mutations could influence metabolite profiles of citrus fruit to a limited extent.

Comprehensive metabolic profiling to decipher the influence of rootstocks on fruit juice metabolome of Kinnow (C. nobilis × C. deliciosa)

The effect of rootstocks on flavor-related compounds, and nutrition and processing quality of Kinnow mandarin (C. nobilis × C. deliciosa) was investigated by targeted metabolomics approach employing LC–MS/MS and GC–MS/MS. Fruit samples were collected from mature trees grafted on six rootstocks, viz. Rough lemon 1 (C. jambhiri Lush), Rough lemon 2 (C. jambhiri), Sour orange (C. aurantium ‘Karun Jamir’), Cleopatra mandarin (C. reshni), Pectinifera (C. × depressa ‘Pectinifera’), and Shekwasha (C. × depressa ‘Shekwasha’). The data revealed significant differences in the concentration of some primary and secondary metabolites. Sour orange rootstock showed a higher concentration of polyphenolics and limonoid aglycones and a lower concentration of sugars, organic acids and aroma volatiles as compared to Shekwasha and Pectinifera. Rough lemon and Cleopatra rootstock presented desired concentration of organic acids and limonoid aglycones with moderate concentration of vitamins (B-complex and C), polyphenolics, and aroma volatiles. Principal component analysis (PCA) segregated Kinnow fruit grafted on different rootstocks into different PCs based on primary and secondary metabolites. Such differences indicate the influence of rootstock on Kinnow metabolome. This study highlights the importance of metabolomics in illuminating metabolic profile of citrus, thereby aiding in the selection of the desired rootstock for intended fruit quality.

Characterization of Limonoids in Citrus Essential Oils by Means of Supercritical Fluid Chromatography Tandem Mass Spectrometry

The dual purpose of this research was to investigate the presence of limonoid aglycones in Citrus essential oils and to develop a rapid and reliable SFC-APCI-QqQ MS method for their characterization. In this study, 12 limonoid aglycones were tentatively identified in 11 different citrus essential oils. The developed method allowed a very fast separation in less then 7 min with a very low amount of organic solvent. Calibration curve of limonin was constructed by using a triple quad MS detector in order to quantify the limonin content in the analyzed samples. The content of limonin ranged from 0.5 (clementine essential oil) to 21.2 mg L−1 (bergamot essential oil) in the cold-pressed Citrus essential oils analyzed. Moreover, product ion scan (PIS) acquisition mode was also used for the structure elucidation of isobaric compounds. To the best of our knowledge, this is the first investigation on limonoid aglycones in Citrus essential oils.

Decoding the Nonvolatile Sensometabolome of Orange Juice ( Citrus sinensis).

Activity-guided fractionation in combination with the taste dilution analysis, followed by liquid chromatography-tandem mass spectrometry and nuclear magnetic resonance experiments, led to the identification of 10 polymethoxylated flavones (PMFs), 6 limonoid glucosides, and 2 limonoid aglycones as the key bitterns of orange juice. Quantitative studies and calculation of dose-over-threshold factors, followed by taste re-engineering, demonstrated for the first time 25 sensometabolites to be sufficient to reconstruct the typical taste profile of orange juices and indicated that not a single compound can be considered a suitable marker for juice bitterness. Intriguingly, the taste percept of orange juice seems to be created by a rather complex interplay of limonin, limonoid glucosides, PMFs, organic acids, and sugars. For the first time, sub-threshold concentrations of PMFs were shown to enhance the perceived bitterness of limonoids. Moreover, the influence of sugars on the perceived bitterness of limonoids and PMFs in orange juice relevant concentration ranges was quantitatively elucidated.

Citrus Limonoid Glucosyltransferase: AKey Player For Natural Debittering And Anticancerous Potential

Citrus fruits and juices are rich source of health benefitting phytochemicals which play a vital role in balanced diet and disease prevention. Citrus limonoids and flavonoids are the major phytochemicals which are of great interest in pharmaceutical industries because of their demonstrated anticancerous, antioxidant, anti-inflammatory, hormonal stimulation, antibacterial and antiviral actions. Citrus limonoid biosynthetic pathway contains an important regulatory limonoid glucosyltransferase enzyme (LGT). LGT is the natural debittering enzyme encoded by a single copy gene which has been isolated from different Citrus spp. This enzyme is mainly responsible for conversion of all limonoid aglycones (mostly bitter) to their corresponding glucosides (mostly nonbitter) but only during late fruit developmental stage of citrus. Citrus LGT belongs to glycosyltransferase super family whose members are the wide managers to catalyze the transfer of sugar molecules to their acceptor molecules to play several key modifications in plant secondary metabolites. These reveal great significance value in plant cell metabolism especially in detoxification of xenobiotics, production and storage of natural products. Despite to the fact that over expression of LGT in citrus will lead to reduce the delayed bitterness caused by limonin (an aglycone) but in addition will enhance the accumulation of limonoid glucosides in fruits. Further, recent studies suggest that citrus limonoids especially glucosides have shown importance against brain, pancreas, colon, and breast cancers. Thus, future studies should be focused on utilizing the potential of LGT present in citrus plants in terms of anticancerous properties as well as reducing the delayed bitterness problem important for citrus juice industry

Functional evaluation of yuzu (Citrus junos) extracts containing limonoids and polyamine for life extension

We determined high concentrations of arginine and polyamines (putrescine, spermidine, and spermine) in yuzu (Citrus junos) seeds. Yuzu produces large quantities of limonoids. We fed limonoid aglycones extracted from yuzu seeds and commercial spermine asa polyamine to Sandhoff disease (SD) mice with abnormal glycolipid metabolism and autosomal recessive inheritance, as found in humans. Ingestion of limonoid aglycones and spermine by SD mice extended their longevity by about 10–12%, where it alleviated inflammation of the central nervous system. After administering limonoid aglycones or limonoid aglycones/spermine, analyses of the bacterial flora in the feces based on 16S rDNA indicated changes in the relative ratio between Bacteroidetes (Bacteroidales and Lactobacillus) and Firmicutes (Clostridiales and Erysipelotrichaceae). In both SD and wild-type mice, the amount of short-chain fatty acids produced by the bacterial flora increased according to the intake of limonoid aglycones and spermine.

Largely different contents of terpenoids in beef red-flesh tangerine and its wild type

BackgroundNiurouhong (Citrus reticulata Blanco. Niurouhong) (NRH) is a spontaneous beef-red flesh mutant with distinctive flavor compared with its wild type orange-red flesh Zhuhongju (ZHJ). To illustrate the biochemical mechanism of its special flesh color and flavor, fruits at commercial mature stage were used to profile the volatiles in the flavedo and determine the levels of carotenoids, limonoid aglycones and phytohormones in the juice sacs in two seasons.ResultsOur results showed the content of total volatile terpenoids in NRH was 1.27-fold that in ZHJ. The components of volatiles were found to be common between the two tangerines. This result indicates that the distinctive flavor of NRH might not be derived from the presence/absence of specific volatiles; instead, it was derived from the altered concentrations or balance of α-citral, β-citral, 2-cyclohexen-1-one, (S)-3-methyl-6-(1-methylethenyl) and n-hexadecanoic acid. Analyses of the contents of total and specific carotenoids indicated that the beef-red color of NRH flesh might be largely attributed to the over accumulation of β-cryptoxanthin and β-carotene. However, lower ABA level was found in NRH than in ZHJ, reflecting a possible feedback regulation of ABA biosynthesis on carotenogenesis and the balance in the metabolism among terpenoids.ConclusionsCollectively, our study suggested that the MEP pathway was enhanced in NRH tangerine. However, a certain unknown co-regulatory mechanism might be present in the metabolism pathway of secondary metabolites (especially terpenoids) in beef-red flesh mutant. Our study provides new insights into the regulatory network of terpenoid metabolism and mutation mechanism of red-fleshed citrus.

Cloning and Characterization of Limonoid Glucosyltransferase from Kinnow Mandarin (Citrus reticulata Blanco).

Kinnow mandarin (Citrus reticulata Blanco) is a popular citrus crop of northwestern India and it occupies maximum fruit area in Punjab. However, citrus juice processing industry is still suffering from delayed bitterness problem caused mainly by limonoid aglycones such as limonin. In order to study citrus limonoid metabolism, limonoid glucosyltransferase (LGT) gene, which encodes a natural debittering enzyme, was isolated from the fruit tissues of Kinnow mandarin. After confirmation and characterization, its full-length gene sequence (1533 bp) was submitted to National Centre for Biotechnology Information. Citrus reticulata limonoid glucosyltransferase (CrLGT) occupies a position on an independent branch in the largest subgroup and is phylogenetically different from those in other mandarin species like C. unshiu, showing its uniqueness in several features. The transcript expression of CrLGT, evaluated in different tissues such as young leaf, flavedo, albedo, sac covering and seed of Kinnow mandarin during early (90 days after flowering (DAF)), mid (150-210 DAF) and late (240 DAF) fruit developmental stages using semi-quantitative method, showed the highest expression in flavedo. Thus, it was concluded that the isolated LGT gene has an effect on limonoid metabolic engineering in citrus. Overexpression of this gene can reduce the delayed bitterness problem in citrus juice and enhance the accumulation of specific glucosides that have anticancer effects.

Liquid Chromatography-Electrospray Ionization Mass Spectrometry Analysis of Limonoids and Flavonoids in Seeds of Grapefruits, Other Citrus Species, and Dietary Supplements.

A selective UHPLC-DAD-QToF-MS method was developed to screen grapefruit seeds, and the seeds of other Citrus species for limonoid aglycones, acids, glucosides, and flavonoids. These classes of compounds were identified in positive and negative ion modes over a mass-to-charge range from 100-1500. Accurate mass values, elution times, and fragmentation patterns obtained by QToF-mass spectrometry were used to identify or tentatively characterize the compounds detected in the sample of this study. Limonin was the major limonoid in most of the seeds of Citrus species, followed by nomilin. This analytical method was successfully applied for the analysis of commercial extracts and dietary supplements claiming to contain grapefruit seed extract, or extracts made from the seed and other fruit parts such as the peel or pulp. Many commercial products contained large numbers of flavonoids, indicating the use of peel, pulp, or seed coat. This method also permitted detection of synthetic preservatives such as benzethonium chloride, methylparaben, and triclosan in commercial grapefruit seed extract products. Out of the 17 commercial products analyzed, two contained the synthetic antimicrobial agent benzethonium chloride.

Bergamot (Citrus bergamia Risso) as a source of nutraceuticals: Limonoids and flavonoids

Limonoids are human health promoters, and have many pharmacological properties: anticancer, antioxidant, antibacterial and antifungal. To the best of the authors' knowledge, there are no previous reports on the composition of limonoids in bergamot fruits, so the aim of this work was to characterize bergamot peel, pulp, seed and juice. Samples analysed showed the presence of both limonoids aglycones and glucosides. Limonoid aglycones are the most abundant in seeds and peels (70 and 80% of the total, respectively), while limonoid glucosides are the more abundant in juices and pulps (61 and 76% of the total, respectively). Moreover a method to isolate pure limonoids from bergamot seeds and juice by means of a 2D-HPLC/PDA/MS preparative system is proposed. Pure molecules isolated were used to build calibration curves to quantify limonoids in real samples. Isolation of nutraceuticals from seeds and juices can represent an advantageous way to re-evaluate these by-products.

Evaluation of limonoid production in suspension cell culture of Citrus sinensis

The use of cell and plant tissue culture techniques to produce economically important active metabolites has been growing. Among these substances are total limonoid aglycones, which are produced by “pera” orange (Citrus sinensis (L.) Osbeck, Rutaceae) and have received considerable attention because of their anticancer actions. The main objective of the present study was to analyze and compare the levels of limonoid aglycones in seeds, callus cultures (originating from seeds), callus cultures (originating from hypocotyls), cell suspensions from hypocotyls cells, and cell suspensions from cotyledons. The cell cultures or C. sinensis were obtained by inoculating two strains of callus in MS medium supplemented with 2.0μM 2,4-dichlorophenoxyacetic acid, 7.0μM benzyl aminopurine, and 3% (w/v) sucrose in the dark. The highest concentrations of limonoid aglycone that were obtained were observed in cotyledon cell lines (240mg/100g dry weight) that were produced on day 21 of culture and hypocotyl cell lines on day 7 (210mg/100g dry weight). Explants of different origins under the same culture conditions had different limonoid aglycone content. The present results may suggest strategies for enhancing the productivity of biologically important limonoid aglycones and investigating the complex pathways of these secondary metabolites in plant tissue cultures.

Features of citrus terpenoid production as revealed by carotenoid, limonoid and aroma profiles of two pummelos (Citrus maxima) with different flesh color.

Terpenoids are major components of carotenoids, limonoids and aromas in citrus fruits, resulting in fruit coloration, bitterness and aroma. In this study the carotenoid, limonoid and volatile profiles of red-flesh Chuhong pummelo (CH) and pale green-flesh Feicui pummelo (FC) were investigated by HPLC and GC/MS. Large differences were found in constituents of carotenoids and limonoids in juice sacs and flavedo and of aromas in flavedo of the two pummelos. For carotenoids in juice sacs, CH contained 57 times the amount in FC, mainly all-trans-lycopene and phytoene, whereas in flavedo it contained only 25% of that in FC, the latter showing a high proportion of β-carotene and other chloroplastic carotenoids. In comparison with FC, limonin and nomilin aglycone production was boosted in juice sacs of CH while being almost absent in flavedo. For volatiles in flavedo, the total amount was significantly higher in CH. PCA suggested that germacrene-type sesquiterpenoids, etc. were principal in distinguishing volatile profiles of the two pummelos. The data showed a different tissue-biased pattern of carotenoid and limonoid aglycone synthesis in pummelos with different flesh color, and the possible independently regulated synthesis of those metabolites in different fruit tissues. Furthermore, decreased carotenoid and limonoid aglycone production accompanied by increased accumulation of volatile terpenoids in flavedo of red-flesh CH was identified, indicating that a total capacity or a balance of production of various terpenoids might exist in pummelo fruit tissues. It was also suggested that substrate concentration is not the key factor affecting product concentrations during the synthesis of monoterpene derivatives.

The functional evaluation of waste yuzu (Citrus junos) seeds

We have succeeded in extracting a large amount of expensive limonoids and the high total antioxidant capability yuzu seed oil from waste yuzu seed by simple methods. Yuzu seeds contain higher amounts of fat-soluble limonoid aglycone (330.6 mg g(-1) of dry seed), water-soluble limonoid glycoside (452.0 mg g(-1) of dry seed), and oil (40 mg g(-1) of green seed) than other citrus fruits. The antioxidant activities of yuzu seed aglycone, glycoside, and seed oil were evaluated in vitro. The potential antioxidant activity in oil solution, diphenylpicrylhydrazyl radical-scavenging activity, and hydrogen peroxide-scavenging activity effects of the seed extracts were also investigated. The antioxidant activity of yuzu seed oil was two times that of grapefruit seed oil, which has high activity. Yuzu glycoside produced the same high antioxidant activity as Luo Han Guo glycoside.