Computational Exploration of Limonin as a Potential Inhibitor of DapB in Klebsiella pneumoniae.

Klebsiella pneumoniae has emerged as a significant multidrug-resistant pathogen, classified as a critical priority by the World Health Organization. The rising rates of antibiotic resistance have led to increased therapeutic failures, diminishing the effectiveness of existing antibiotics. Consequently, there is an urgent need for alternative treatments to effectively inhibit the growth of K. pneumoniae and mitigate associated diseases. Phytochemicals have demonstrated potential advantages over traditional antibiotics, prompting their exploration as innovative therapeutic agents. This study aimed to identify phytochemicals that can inhibit dapB, a vital enzyme in the lysine biosynthesis pathway of K. pneumoniae, which is essential for protein synthesis and the cross-linking of the bacterial peptidoglycan cell wall. We screened 17,934 phytochemicals based on Lipinski's Rule of Five, along with their Absorption, Distribution, Metabolism, Excretion properties and toxicological parameters. Next, we conducted triplicate docking studies against dapB to evaluate the library further. The most promising molecules then underwent 100 ns Molecular Dynamics simulations in triplicate, followed by binding free energy calculations to identify potential dapB inhibitors. This in silico analysis highlighted limonin as a promising inhibitor of dapB in K. pneumoniae. Further experimental validation is crucial to enhance limonin's potential as a novel therapeutic agent against K. pneumoniae-associated diseases.

Oral Ingestion of Yuzu Seed Oil Suppresses the Development of Atopic Dermatitis-like Skin Lesions in NC/Nga Mice.

Long-term oral ingestion of unheated yuzu seed oil in humans reduces lipid peroxides in the blood. Moreover, yuzu seed oil contains limonin, which can induce antioxidant and anti-inflammatory effects by activating the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2). Previously, Nrf2 has been shown to reduce atopic dermatitis (AD). Therefore, we hypothesized that ingesting unheated yuzu seed oil can regulate AD through Nrf2. An AD model was established using NC/Nga mice through repeated local exposure to mite antigens. Unheated and purified yuzu seed oil (100 µL/mice) or water (control, 100 µL/mice) was administered orally once a day using a gastric cannula for rodents for 28 days. On day 28, mice in the unheated yuzu seed oil group exhibited significantly lower clinical skin severity scores and ear thickness than those in the purified yuzu seed oil and water groups. Serum histamine levels remained unaltered among the three AD-induced groups. Serum Dermatophagoides farina body (Dfb)-specific immunoglobulin E (IgE) levels were significantly lower in the unheated yuzu seed oil group. Oral ingestion of yuzu seed oil in NC/Nga AD model mice significantly suppressed dermatitis deterioration and decreased serum IgE levels. Clinical trials (n = 41) have already confirmed that unheated yuzu oil is safe for long-term intake, further suggesting its potential use in improving AD symptoms.

Purification and characterization of limonin D-ring lactone hydrolase from sweet orange (Citrus sinensis (L.) Osbeck) seeds.

BACKGROUND: Citrus products often suffer from delayed bitterness, which is generated from the conversion of non-bitter precursors (limonoate A-ring lactone, LARL) to limonin under the catalysis of limonin D-ring lactone hydrolase (LDLH). In this study, LDLH was isolated and purified from sweet orange seeds, and a rapid and accurate high-performance liquid chromatography method to quantify LARL was developed and applied to analyze the activity and enzymatic properties of purified LDLH. RESULTS: Purified LDLH (25.22 U mg-1) showed bands of 245 kDa and 17.5 kDa molecular weights in native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate PAGE analysis respectively. After a 24 h incubation under strongly acidic (pH 3) or strongly alkaline (pH 9) conditions, LDLH still retained approximately 100% activity. Moreover, LDLH activity was not impaired by thermal treatment at 50 °C for 120 min. Enzyme inhibition assays showed that LDLH was inactivated only after ethylenediaminetetraacetic acid treatment, and other enzyme inhibitors showed no significant effect on its activity. In addition, the LDLH activity of calcium ion (Ca2+) intervention was 108% of that in the blank group, and that of zinc ion (Zn2+) intervention was 71%. CONCLUSION: LDLH purified in this study was a multimer containing 17.5 kDa monomer with a wide pH tolerance range (pH 3-9) and excellent thermal stability. Moreover, LDLH might be a metallopeptidase, and its activity was stimulated by Ca2+ and significantly inhibited by Zn2+. These findings improve our understanding of LDLH and provide some important implications for reducing the bitterness in citrus products in the future. © 2024 Society of Chemical Industry.

Optimization of limonin invertase production by scaling up Aspergillus tubingensis UA13 fermentation to a 5-l scale.

The enzymatic approach is a highly effective and the major scientific method to eliminating bitter components in citrus-derived products nowadays. Microbial production of limonin invertase stands out due to its pivotal role in the removal of the bitter substance, limonin. The optimization of fermentation parameters and the study of scale-up fermentation are imperative for product commercialization. In this study, we focused on optimizing stirring speed, fermentation temperature, and initial pH to enhance the growth and limonin invertase production by the Aspergillus tabin strain UA13 in a 5-l stirred-tank bioreactor. Our results revealed the following optimal parameters are: a stirring speed of 300 rpm, a fermentation temperature of 35°C and a pH 5.0. Under these optimized conditions, the limonin invertase activity reached its peak at 63.38 U ml-1, representing a 1.67-fold increase compared to the unoptimized conditions (38.10 U ml-1), while also reducing the fermentation duration by 12 h. Furthermore, our research demonstrated that limonin invertase effectively hydrolyze limonin in grapefruit juice, reducing its content from 13.28 to 2.14 µg ml-1, as determined by HPLC, resulting in a 6.21-fold reduction of the bitter substance.

Grapefruit Debittering by Simultaneous Naringin Hydrolysis and Limonin Adsorption Using Naringinase Immobilized in Agarose Supports.

Naringin and limonin are the two main bitter compounds of citrus products such as grapefruit juice. The aim of this investigation was to evaluate the reduction in both bitter components simultaneously using a combined biochemical and physical approach. The proposed strategy was based on the use of heterofunctional supports with glyoxyl groups that allow for the covalent immobilization of naringinase, which hydrolyses naringin and alkyl groups that allow for the adsorption of limonin. The supports were butyl-glyoxyl agarose (BGA) and octyl-glyoxyl agarose (OGA), which were characterized in terms of aldehyde group quantification and FTIR analysis. The optimal pH and temperature of free and immobilized enzymes were assessed. The maximum enzyme loading capacity of supports was analyzed. Debittering of grapefruit juice was evaluated using soluble enzyme, enzyme-free supports, and immobilized catalysts. Enzyme immobilized in BGA reduced naringin and limonin concentrations by 54 and 100%, respectively, while the use of catalyst immobilized in OGA allowed a reduction of 74 and 76%, respectively, obtaining a final concentration of both bitter components under their detection threshold. The use of OGA biocatalyst presented better results than when soluble enzyme or enzyme-free support was utilized. Biocatalyst was successfully applied in juice debittering in five repeated batches.

A Green Extraction Method to Achieve the Highest Yield of Limonin and Hesperidin from Lime Peel Powder (Citrus aurantifolia).

Green extraction is aimed at reducing energy consumption by using renewable plant sources and environmentally friendly bio-solvents. Lime (Citrus aurantifolia) is a rich source of flavonoids (e.g., hesperidin) and limonoids (e.g., limonin). Manufacturing of lime products (e.g., lime juice) yields a considerable amount of lime peel as food waste that should be comprehensively exploited. The aim of this study was to develop a green and simple extraction method to acquire the highest yield of both limonin and hesperidin from the lime peel. The study method included ethanolic-aqueous extraction and variable factors, i.e., ethanol concentrations, pH values of solvent, and extraction temperature. The response surface methodology was used to optimize extraction conditions. The concentrations of limonin and hesperidin were determined by using UHPLC-MS/MS. Results showed that the yields of limonin and hesperidin significantly depended on ethanol concentrations and extraction temperature, while pH value had the least effect. The optimal extraction condition with the highest amounts of limonin and hesperidin was 80% ethanol at pH 7, 50 °C, which yields 2.072 and 3.353 mg/g of limonin and hesperidin, respectively. This study illustrates a green extraction process using food waste, e.g., lime peel, as an energy-saving source and ethanol as a bio-solvent to achieve the highest amount of double bioactive compounds.

Enhanced functionality and shelf stability of burfi by incorporating kinnow (mandarin) fruit juice.

Burfi, an Indian traditional dairy dessert is highly popular, however, its low nutritive value and poor shelf life limits commercial viability. Kinnow juice was utilized to improve the phytonutritional profile and shelf stability of burfi. To further improve the quality, pectin was added at different concentrations (1-4%). Compared to control, inclusion of kinnow juice significantly (p < 0.05) improved minerals and phytochemical constituents as confirmed using FTIR analysis. Developed burfi presented a softer texture which was consistent with SEM results. Increasing the pectin levels, increased textural properties of kinnow burfi while decreased moisture content and water activity, without affecting color and nutritional properties. Based on desirable sensory and textural properties, kinnow burfi prepared with pectin (4% of added sugar) was selected for product development and evaluation of shelf life under room (25 ± 2 °C) and refrigerated (4 ± 2 °C) conditions. Irrespective of storage temperatures, moisture content and bioactive constituents decreased while titratable acidity, total and reducing sugars, free fatty acids, hardness of texture, and yeast and mould count increased significantly. Except moisture, the extent of quality changes was rapid at room temperature. The product showed high sensory acceptability as well as microbial safety up to 21 days at room temperature and 28 days under refrigeration.

Limonin ameliorates acute pancreatitis by suppressing JAK2/STAT3 signaling pathway.

Acute pancreatitis (AP) is one of the most common acute abdomen of digestive system and has the characteristics of dangerous condition and rapid development. Limonin has been confirmed to hold anti-inflammatory and antioxidant effects in various diseases. However, its potential beneficial effect on AP and the concrete mechanisms have never been revealed. Here, two mouse models were used to investigate the protective effects of limonin on AP, the caerulein-induced mild acute pancreatitis (MAP) model and L-arginine-induced severe AP (SAP) model. Firstly, it was found that limonin administration attenuated lipase and serum amylase levels and ameliorated the histopathological manifestations of pancreatic tissue in a dose-dependent manner. Additionally, the amelioration of AP by limonin was associated with reduced levels of inflammation initiators (IL-6, IL-1ß, CCL2, and TNF-α). Mechanistically, we found that limonin suppressed the Janus Activating Kinase 2 (JAK2)/Signal Transducer and Activator of Transcription 3 (STAT3) signaling pathway, as evident by the decreased levels of JAK2 and p-STAT3. And activation of JAK2 using JAK2 activator rescued the protective effects of limonin on AP. Thus, our results demonstrate that limonin can ameliorate AP in two mice models via suppressing JAK2/STAT3 signaling pathway.

[Anti-inflammatory effect, plasma effective components and therapeutic targets of Huanglian Jiedu Decoction on ulcerative colitis mice].

This paper was to investigate the effect of Huanglian Jiedu Decoction(HLJD) on ulcerative colitis(UC) in mice, and determine the effective components in plasma, and virtually screen its therapeutic target, and predict its mechanism. Sixty Balb/c mice were randomly divided into blank group, model group, mesalazine treatment group(0.3 g·kg~(-1)), and HLJD treatment groups(24.66, 12.33, 6.17 g·kg~(-1)). Excepted for the blank group, all the mice in HLJD and mesalazine treatment groups were gavage administration. All mice freely drank 2.5% DSS solution for seven days to induce UC. The disease activity index(DAI) was detected each day. At the end of the experiment, HE staining was used to observe the pathological changes in colon. The content of IL-1ß, IL-6 and TNF-α in colon were determined by ELISA. The effective components in plasma were determined by UPLC-Q-TOF-MS. The reverse docking in PharmMapper was used to screen the component targets. The disease targets of UC were collected by searching TTD, OMIM and GeneCards databases. The intersection of the component targets and disease targets was selected as the therapeutic targets. Then the therapeutic targets were imported into the STRING for GO and KEGG enrichment analysis. Discovery Studio was used to simulate the docking between the components and the targets. RESULTS:: showed that the DAI in the model group increased significantly(P<0.05), and the number of inflammatory cells and infiltration degree increased significantly compared with the blank group. The DAI in HLJD treatment group was significantly reduced(P<0.05), and the number and infiltration degree of inflammatory cells were reduced compared with the model group. The ELISA results showed that the levels of IL-1ß, IL-6 and TNF-α were increased significantly in the model group(P<0.01) compared with the blank group, and significantly down regulated in the HLJD treatment group(P<0.05) compared with the model group. After UPLC-Q-TOF-MS analyse, ten components were identified. The network pharmacology analysis showed that the action targets were significantly enriched in 129 of biological processes, such as response to organic substance, chemical and oxygen-containing compound, etc., as well as 16 of signal pathways, such as IL-17, TNF and hepatitis B signal pathways, were enriched too. The results of molecular docking showed that limonin, palmatine and berberine could bind to CASP3 and MMP9 by hydrogen bond. In conclusion, HLJD could alleviate the colonic mucosal inflammatory infiltration and mucosal damage in UC mice. The mechanism may be related to the anti-inflammatory effect on UC mice by reducing the levels of IL-1ß, IL-6 and TNF-α in colon through limonin, palmatine and berberine regulating IL-17 signal pathway and TNF signal pathway via CASP3 and MMP9 meditated.

Limonin as a Starting Point for the Construction of Compounds with High Scaffold Diversity.

Structurally complex natural products have been a fruitful source for the discovery and development of new drugs. In an effort to construct a compound collection populated by architecturally complex members with unique scaffolds, we have used the natural product limonin as a starting point. Limonin is an abundant triterpenoid natural product and, through alteration of its heptacyclic core ring system using short synthetic sequences, a collection of 98 compounds was created, including multiple members with novel ring systems. The reactions leveraged in the construction of these compounds include novel ring cleavage, rearrangements, and cyclizations, and this work is highlighted by the discovery of a novel B-ring cleavage reaction, a unique B/C-ring rearrangement, an atypical D-ring cyclization, among others. Computational analysis shows that 52 different scaffolds/ring systems were produced during the course of this work, of which 36 are unprecedented. Phenotypic screening and structure-activity relationships identified compounds with activity against a panel of cancer cell lines.

Discovery of deoxylimonin δ-lactam derivative with favorable anti-inflammation and antinociception efficacy from chemical modified limonin/deoxylimonin analogs.

Chemical modifications on the A ring of limonin (1) and deoxylimonin (2) afforded 28 structural characterized derivatives, which were firstly subjected to preliminary in vivo analgesic and anti-inflammatory screen by mice model. The most promising candidate, deoxylimonin analog II-B-2 (70 mg/kg) with 3,4-dimethoxyphenylethyl moiety substitued δ-lactam in the A ring, exhibited better analgesic activity than aspirin (200 mg/kg) and stronger anti-inflammatory efficacy than naproxen (150 mg/kg). Further in vivo evaluation confirmed its advantage over limonin and showed dose-response dependent manner, and follow-up research suggested that the anti-inflammatory effect of compound II-B-2 may be attributed to the downregulation of cyclooxygenase 2 expression and the suppression of prostaglandin E2 formation.

Functional characterization and reclassification of an enzyme previously proposed to be a limonoid UDP-glucosyltransferase.

BACKGROUND: A major problem in the orange industry is 'delayed' bitterness, which is caused by limonin, a bitter compound developing from its non-bitter precursor limonoate A-ring lactone (LARL) during and after extraction of orange juice. The glucosidation of LARL by limonoid UDP-glucosyltransferase (LGT) to form non-bitter glycosyl-limonin during orange maturation has been demonstrated as a natural way to debitter by preventing the formation of limonin. RESULT: Here, the debittering potential of heterogeneously expressed glucosyltransferase, maltose-binding protein (MBP) fused to cuGT from Citrus unishiu Marc (MBP-cuGT), which was previously regarded as LGT, was evaluated. A liquid chromatography - mass spectrometry (LC-MS) method was established to determine the concentration of limonin and its derivatives. The protocols to obtain its potential substrates, LARL and limonoate (limonin with both A and D ring open), were also developed. Surprisingly, MBP-cuGT did not exhibit any detectable effect on limonin degradation when Navel orange juice was used as the substrate; MBP-cuGT was unable to biotransform either LARL or limonoate as purified substrates. However, it was found that MBP-cuGT displayed a broad activity spectrum towards flavonoids, confirming that the enzyme produced was active under the conditions evaluated in vitro. CONCLUSION: Our results based on LC-MS demonstrated that cuGT functionality was incorrectly identified. Its active substrates, including various flavonoids but not limonoids, highlight the need for further efforts to identify the enzyme responsible for LGT activity to develop biotechnology-based approaches for producing orange juice from varietals that traditionally have a delayed bitterness. © 2020 Society of Chemical Industry.

[UPLC-MS/MS determination of active components from Wuzhuyu Decoction in plasma and brain tissues of nitroglycerin-induced migraine rats].

A sensitive and specific ultra-performance liquid chromatography-mass spectrometry(UPLC-MS/MS) method was deve-loped for analysis of rutaecarpine(Ru), evodiamine(Ev), rutaevine(Rv), limonin(Li), ginsendside Rb_1(Rb_1), ginsendside Re(Re) in rat plasma and brain tissues of nitroglycerin-induced migraine rats. Male healthy Sprague-Dawley(SD) rats were orally given multiple dose of optimized(OS) and un-optimized Wuzhuyu Decoction(UNOS), and their blood samples and brainstem were collected at different time points after injection of nitroglycerin(10 mg·kg~(-1)) into the frontal region. The drug concentrations of the 6 analytes in plasma and brainstem were determined by UPLC-MS/MS method. Subsequently, the main pharmacokinetics parameters of plasma were calculated by using Phoenix WinNolin 5.2.1 software. The methodological test showed that all of analytes in both plasma and brainstem homogenate exhibited a good linearity within the concentration range(r>0.994 7). The intra-day and inter-day accuracy, precision, matrix effect, stability of the investigated components meet the requirements for biopharmaceutical analysis. The developed method was successfully applied in pharmacokinetic studies on abovementioned ingredients in rat plasma and brain stem. The plasma pharmacokinetic parameters of active ingredients in two different Wuzhuyu Decoction group were compared, it was found that Rb_1 had higher t_(1/2), T_(max), C_(max), AUC_(0-24 h) and AUC_(0-∞ )in OS group. Meanwhile, Ev had higher t_(1/2) and T_(max) but lower C_(max), AUC_(0-24 h) and AUC_(0-∞), Ru has higher t_(1/2 )but lower C_(max), AUC_(0-24 h) and AUC_(0-∞ )in OS group. The brain tissue distribution of each component were compared between the two groups, the component with higher content in OS, such as Ru at 30 min and 2 h after administration, Ev at 30 min, Rb_1 at 30 min and Rb_1 at 2 h after administration have lower brain tissue distribution than those in UNOS group, while the component with higher content in UNOS, such as Rv at 30 min, 2 h and 12 h after administration had higher brain tissue distribution than those in OS group.

Anticancer effect of Limonin against benzo(a)pyrene-induced lung carcinogenesis in Swiss albino mice and the inhibition of A549 cell proliferation through apoptotic pathway.

The main purpose of the current study is to reveal the anticancer action of limonin against benzo(a)pyrene [B(a)P]-treated lung carcinogenesis in Swiss albino mice and A549 lung cancer cells. B(a)P was orally supplemented (50 mg/kg body weight) twice a week for four weeks induction of lung cancer in mice. The lung weight, body weight, incidence of tumor, lipid peroxidation, carcinoembryonic antigen (CEA), enzymatic and nonenzymatic antioxidants (superoxide dismutase, GPx, glutathione, glutathione reductase, catalase, and glutathione S-transferase), serum marker enzymes (aryl hydroxylase, lactate dehydrogenase, 5'-nucleotidases, and γ-glutamyl transpeptidase), and inflammatory mediators (interleukin-1ß, interleukin-6, and tumor necrosis factor-α) were estimated. Moreover, a histopathological study of lung tissues was supported by the biochemical analysis. Furthermore, the anticancer activity of limonin on A549 cells was measured by cell viability, production of reactive oxygen species (ROS), apoptotic morphological changes by AO/EtBr staining. Additionally, the status of apoptosis protein (caspase-9 and -3) expressions was analyzed by the colorimetric analysis. B(a)P-induced mice showed increased lipid peroxidation, CEA, serum marker enzymes and inflammatory cytokines levels with simultaneously decreased in the nonenzymatic and enzymatic antioxidants levels. Limonin supplements significantly reverted back to all these changes in this manner, showing the efficiency of anticancer effect. Furthermore, our in vitro study also supported the anticancer effect of the treatment of limonin-enhanced apoptosis by loss of cell viability, improved ROS production, apoptotic morphological changes, and apoptosis protein expression were analyzed. Overall, these results suggest the anticancer potential of limonin against B(a)P-induced lung cancer in Swiss albino mice and A549 lung cancer cells.

Isolation and screening for limonin-producing endophytic bacteria from Citrus maxima (Burm.) Merr. cv. Shatian Yu.

Limonin, a compound of highly oxidized triterpenoids, has potential functions in preventing or slowing the occurrences of many diseases. In this study, five different bacterial strains were isolated and identified from Citrus maxima (Burm.) Merr. cv. Shatian Yu. Morphological characteristics and 16S rRNA gene sequencing identified them as Bacillus spp, in which two limonin-producing endophytes named P and P9 were discovered by high-performance liquid chromatography and mass spectrometry using an inorganic salt medium and two natural media; also the production was greater in natural medium 1 (4.377 and 0.299 mg/L, respectively) than in natural medium 2 (0.159 and 0.025 mg/L, respectively). The growth and fermentation characteristics of strain P were studied, and during the liquid cultivation of Bacillus sp. P, limonin began to accumulate at the eighth hour in the inorganic salt medium, peaked at the 16th hour, and then decreased sharply. Single-factor experiments revealed that the optimum fermentation conditions for limonin production included 14-H-old cells, 15% inoculum, and 3 g/L glucose.

Limonin: A Review of Its Pharmacology, Toxicity, and Pharmacokinetics.

Limonin is a natural tetracyclic triterpenoid compound, which widely exists in Euodia rutaecarpa (Juss.) Benth., Phellodendron chinense Schneid., and Coptis chinensis Franch. Its extensive pharmacological effects have attracted considerable attention in recent years. However, there is no systematic review focusing on the pharmacology, toxicity, and pharmacokinetics of limonin. Therefore, this review aimed to provide the latest information on the pharmacology, toxicity, and pharmacokinetics of limonin, exploring the therapeutic potential of this compound and looking for ways to improve efficacy and bioavailability. Limonin has a wide spectrum of pharmacological effects, including anti-cancer, anti-inflammatory and analgesic, anti-bacterial and anti-virus, anti-oxidation, liver protection properties. However, limonin has also been shown to lead to hepatotoxicity, renal toxicity, and genetic damage. Moreover, limonin also has complex impacts on hepatic metabolic enzyme. Pharmacokinetic studies have demonstrated that limonin has poor bioavailability, and the reduction, hydrolysis, and methylation are the main metabolic pathways of limonin. We also found that the position and group of the substituents of limonin are key in affecting pharmacological activity and bioavailability. However, some issues still exist, such as the mechanism of antioxidant activity of limonin not being clear. In addition, there are few studies on the toxicity mechanism of limonin, and the effects of limonin concentration on pharmacological effects and toxicity are not clear, and no researchers have reported any ways in which to reduce the toxicity of limonin. Therefore, future research directions include the mechanism of antioxidant activity of limonin, how the concentration of limonin affects pharmacological effects and toxicity, finding ways to reduce the toxicity of limonin, and structural modification of limonin-one of the key methods necessary to enhance pharmacological activity and bioavailability.

Effects of Postharvest Time, Heat Treatment, pH and Filtration on the Limonin Content in Newhall Navel Orange (Citrus sinensis Osbeck cv. Newhall) Juice.

Delayed bitterness causes severe economic loss in citrus juice industry worldwide, which is mostly due to the formation of limonoid compounds, especially limonin, in juice. In this study, effects of postharvest time of fruits, heat treatment, pH and filtration of juice on limonin content in Newhall navel orange (Citrus sinensis Osbeck cv. Newhall) juice were investigated. Our research indicated for the first time that: (1) limonin content in juice would gradually increase to a maximal level and then remained almost constant thereafter as storage time going on, whereas the maximum constant value (MCV) of limonin content in juice significantly (p < 0.05) decreased with the increment of postharvest time of fruits being juiced; (2) heat treatment and acidification of juice only speeded up the formation of limonin to the maximal level while without changing the MCV of limonin content; (3) the juice after filtration exhibited much lower MCV of limonin content compared with the unfiltered one. These experimental observations might not only provide useful information for the development of new debitterness method for navel orange juice, but also strongly support the acid-promoted delayed bitterness mechanism, suggesting the formation of delayed bitterness might primary due to the acid-promoted rather than the enzyme-catalyzed lactonization of limonoate A-ring lactone (LARL) to produce limonin in juice of navel orange.

Dose-response relationship in Schistosoma mansoni juvenile and adult stages following limonin treatment in experimentally infected mice.

Preventive chemotherapy with praziquantel is the mainstay of schistosomiasis control. However, drug resistance is an imminent threat, particularly with large-scale administration of praziquantel, in addition to much less efficacy against young schistosomes. Several biological activities of limonin have been explored such as insecticidal, insect antifeedant, and growth-regulating activity on insects as well as antimalarial, antiviral, anticancer, cholesterol-lowering, and antioxidant activities. This study investigates limonin as an alternative antischistosomal compound using two novel, single, oral dose regimens. In the current work, the therapeutic efficacy of different limonin dosing protocols was evaluated in experimentally infected mice harboring Schistosoma mansoni (Egyptian strain) juvenile or adult stages. Oral administration of limonin in a single dose of 50 or 100 mg/kg on day 21 post-infection (p.i.) resulted in a significant worm burden reduction of 70.0 and 83.33 %, respectively. The same dose given on day 56 p.i. reduced total worm burdens by 41.09 and 60.27 %, respectively. In addition, significant reductions of 34.90 and 47.16 % in the hepatic and 46.67 and 56.1 % in the intestinal tissue egg loads, respectively, associated with significant alterations in the oogram pattern with elevated dead egg levels. Limonin produced ameliorations of hepatic pathology with reduction in dimensions and number of granulomas. Limonin also produced a variety of tegumental alterations in treated worms including tubercular disruption, edema, blebbing, and ulcerations. Results obtained by this work elucidated promising limonin bioactivity against S. mansoni juvenile and adult stages and provided a basis for subsequent experimental and clinical trials.

Development of delayed bitterness and effect of harvest date in stored juice from two complex citrus hybrids.

BACKGROUND: Mandarins and mandarin hybrids have excellent flavor and color attributes, making them good candidates for consumption as fresh fruit. When processed into juice, however, they are less palatable, as they develop delayed bitterness when stored for a period of time. In this study the kinetics of delayed bitterness in two citrus mandarin hybrid siblings, 'Ambersweet' and USDA 1-105-106, was explored by sensory and instrumental analyses. In addition to the bitter limonoids, other quality factors (i.e. sugars, acids, pH, soluble solids content (SSC), titratable acidity (TA) and the ratio SSC/TA) were also measured. RESULTS: The two citrus hybrid siblings had different chemical profiles, which were perceived by taste panels. USDA 1-105-106 developed delayed bitterness when the juice was stored for more than 4 h, similar to juice from 'Navel' oranges, but 'Ambersweet' did not. Bitterness in 'Ambersweet' was more affected by harvest maturity, as juice from earlier harvest had lower SSC but higher TA and bitter limonoids. CONCLUSION: Since juice of USDA 1-105-106 shows delayed bitterness when stored for more than 4 h, this cultivar is not suitable for juice processing. Our finding that siblings can differ in chemical and sensory properties emphasize the importance of post-processing storage studies before releasing cultivars for juice.

Limonin, a Component of Dictamni Radicis Cortex, Inhibits Eugenol-Induced Calcium and cAMP Levels and PKA/CREB Signaling Pathway in Non-Neuronal 3T3-L1 Cells.

Limonin, one of the major components in dictamni radicis cortex (DRC), has been shown to play various biological roles in cancer, inflammation, and obesity in many different cell types and tissues. Recently, the odorant-induced signal transduction pathway (OST) has gained attention not only because of its function in the perception of smell but also because of its numerous physiological functions in non-neuronal cells. However, little is known about the effects of limonin and DRC on the OST pathway in non-neuronal cells. We investigated odorant-stimulated increases in Ca(2+) and cAMP, major second messengers in the OST pathway, in non-neuronal 3T3-L1 cells pretreated with limonin and ethanol extracts of DRC. Limonin and the extracts significantly decreased eugenol-induced Ca(2+) and cAMP levels and upregulated phosphorylation of CREB and PKA. Our results demonstrated that limonin and DRC extract inhibit the OST pathway in non-neuronal cells by modulating Ca(2+) and cAMP levels and phosphorylation of CREB.