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Mercury (Hg2+) as environmental pollutant is a widespread concern due to its cytotoxic effect in humans and animals and needs to be monitored through cost effective methods based on naked-eye detection. Herein, in this study, we extracted and isolated the limonin (LMN) via a facile procedure and then employed it as capping agent for the synthesis of silver nanoparticles (LMN-AgNPs) as a first report. LMN-AgNPs were characterized through Field Emission Scanning Electron Microscopy (FESEM), Atomic Force Microscopy (AFM), Dynamic Light Scattering (DLS), Zeta Potential Analyzer (ZPA) and Fourier Transform Infrared (FTIR) spectroscopy. As-formed LMN-AgNPs were recognized as extremely selective and highly sensitive colorimetric sensor for Hg2+ with potential analytical application. The developed sensor showed an outstanding linear correlation with the concentration of Hg2+ in the range of 0.002 − 55 µM via a color change from deep yellow to transparent showing hypsochromic-hypochromic shift with the limit of detection (LOD) and the limit of quantification (LOQ) as low as 0.21 nM and 0.7 nM respectively. The sensor was further allied with smartphone for immediate and on-site quantification of Hg2+. The LOD and LOQ of 0.42 µM and 1.4 µM was true for smartphone based sensing in the range of 7.5–55 µM Hg2+. The detection of Hg2+ was not disrupted by the presence of other metals in either of mentioned cases. The practical applicability of the proposed Hg2+ sensor was tested using spectrophotometric and smartphone based approaches in human serum and urine as well as in tap water samples with acceptable ranges of recovery. As-developed sensor can work as a potential candidate for monitoring of Hg2+ pollution in diverse fields of studies. [ABSTRACT FROM AUTHOR]

Ultrasound-assisted extraction (UAE) is a promising technique for isolating bioactive compounds from plant materials, as ultrasound disrupts cell walls through mechanical effects, and facilitates the release of cellular constituents. Despite its growing popularity, the efficiency of UAE for extracting limonin from citrus peels, particularly Wenling Gaocheng, remains understudied. In the present work, we systematically explored the influence of various operational parameters on the yield of limonin using UAE. The limonin content was quantified using a Waters 2695 HPLC system coupled with a Waters 2487 UV detector. The effects of six parameters on the UAE of limonin from Wenling Gaocheng peels was examined through single-factor experiments. We observed that the limonin yield first increased and then decreased with decreasing particle size. The yield also increased as the solvent/material ratio increased, but the rate of increase decreased. Among the tested solvent concentrations, 70% ethanol was found to be the most effective. The yield significantly improved with increasing ultrasonic intensity up to 0.2556 W/cm² before decreasing sharply. Between 20 and 70°C, the yield fluctuated slightly, and a gradual improvement was observed with longer extraction times. These findings provided valuable insights for future commercialisation of UAE to extract limonin from citrus peels. [ABSTRACT FROM AUTHOR]

To investigate the effects of limonin (Lim) on oxidative stress and early apoptosis in bovine oocytes during in vitro maturation (IVM), different concentrations of Lim (0, 10, 20, 50 μmol/L) were added to bovine IVM medium. Oocyte maturation rates and development 24 h after in vitro fertilization (IVF) were examined to determine the optimal Lim concentration. The optimal Lim concentration was added to the IVM medium, and 0 μmol/L Lim was used as the control. Immunofluorescence staining was used to detect the abnormal rate of spindle assembly, reactive oxygen species (ROS), glutathione (GSH), mitochondrial membrane potential (MMP) levels, mitochondrial distribution, and the fluorescence intensity of cathepsin B (CB)-active LC3 protein. RT‒qPCR was used to detect the mRNA expression levels of antioxidant-, apoptosis- and autophagy-related genes in oocytes. The total number of blastocysts and the proportion of apoptotic cells among blastocysts were detected. The results showed that the PBI ejection rate, cleavage rate and blastocyst rate of bovine oocytes in the 20 μmol/L Lim group were significantly higher than those in the control group (P < 0.05). Compared with those in the control group, ROS levels, abnormal mitochondrial distribution, the proportion of abnormal spindle assembly, CB activity and LC3 protein fluorescence intensity of oocytes in the 20 μmol/L Lim group were significantly decreased (P < 0.05), and GSH and MMP levels were significantly increased (P < 0.05). The expression of antioxidant genes (Prdx3, Prdx6, Sirt1) and antiapoptotic genes (Bcl-xl, Survivin) were significantly upregulated (P < 0.05), and the expression levels of proapoptotic genes (Caspase-4, BAX) and autophagy-related genes (LC3) were significantly downregulated (P < 0.05). The total number of cells among in vitro fertilized embryos was significantly increased (P < 0.05), and the apoptosis rate of blastocysts was significantly decreased (P < 0.05). Here, we show that Lim exerts positive effects on bovine oocyte IVM by regulating REDOX homeostasis, reducing spindle damage and enhancing mitochondrial function during IVM, thereby inhibiting oocyte apoptosis and autophagy. • Limonin is a natural antioxidant. • Its role in the repair of oxidative damage has attracted more and more attention from researchers, but the specific role of limonin in animal reproduction, especially in female gamete development, is rarely studied. • Lim has a positive effect on the IVM of bovine oocytes by regulating REDOX homeostasis during IVM to reduce spindle damage and enhance mitochondrial function, thereby inhibiting oocyte apoptosis and autophagy. [ABSTRACT FROM AUTHOR]

[Objective] The construction of limonin-albumin nanoparticles provides a theoretical basis for the further application of limonin. [Methods] Limonin-albumin nanoparticles were prepared by desolvation-chemical crosslinking method using bovine serum albumin as nanocarriers. The optimal preparation process was determined by single factor experiment and response surface test with embedding rate as index, and the nanoparticles were characterized. [Results] When the amount of bovine serum albumin was 64 mg, the amount of lemon bitter was 4 mg, the specific volume ratio of alcohol to water was 5∶1, and the amount of crosslinker was 125 μL, the average particle size and polydispersion index of limonin nanoparticles were (152.2±0.5) nm and 0.196±0.026 respectively. The embedding efficiency was (79.21±0.36)%. The limonin-albumin nanoparticles observed under low-pressure transmission electron microscopy showed a spherical shape. Infrared spectroscopy and X-ray diffraction results showed that limonin was successfully encapsulated in albumin in an amorphous or disordered state. The limonin-albumin nanoparticle suspension has good storage stability. [Conclusion] The limonin-albumin nanoparticles prepared by desolvation-chemical crosslinking method had good storage stability.

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. [ABSTRACT FROM AUTHOR]

Objective To investigate the influence of limonin on the malignant biological behavior of non-small cell lung cancer (NSCLC) cells by regulating the protein tyrosine kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling pathway. Methods CCK-8 method was applied to detect the survival rate of A549 cells treated with different concentrations of limonin (0, 5, 10, 25, 50, 75, 100 μmol/L). A549 cells were separated into normal culture (NC) group, low-dose limonin group (treatment with 10 μmol/L limonin for 24 h), medium-dose limonin group (treatment with 25 μmol/L limonin for 24 h), high-dose limonin group (treatment with 50 μmol/L limonin for 24 h), coumermycin A1 group (treatment with 10 μmol/L JAK2 activator coumermycin A1+50 μmol/L limonin for 24 h), and AG490 group (treatment with 10 μmol/L JAK2 inhibitor AG490+50 μmol/L limonin for 24 h). Clone formation assay was applied to detect the clones of each group of cells. Transwell assay was applied to detect cell migration and invasion, and flow cytometry was applied to detect apoptosis. Western blot analysis was applied to detect the protein expression levels of JAK2, p-JAK2, STAT3, p-STAT3, E-cadherin, N-cadherin, and vimentin in each group. Results The viability of A549 cells decreased significantly in a limonin concentration-dependent manner (P < 0.05), with IC50 of 45.16±1.66 μmol/L. Concentrations of 10, 25, and 50 μmol/L were selected for subsequent experiments. The numbers of clones, migration, and invasion of A549 cells and the protein expression levels of IL-6, p-JAK2, p-STAT3, N-cadherin, and vimentin in the low-, medium-, and high-dose limonin groups significantly decreased, compared with those in the NC group, and the apoptosis rate and E-cadherin protein expression significantly increased (P < 0.05). The JAK2 activator coumermycin A1 attenuated the ability of limonin to inhibit the proliferation, migration, invasion, and other malignant biological behavior of A549 cells and attenuated the apoptosis ability. The JAK2 inhibitor AG490 enhanced the ability of limonin to inhibit the proliferation, migration, invasion, and other malignant biological behavior of A549 cells and enhanced the apoptosis ability. Conclusion Limonin can inhibit the malignant biological behavior of NSCLC cells, such as proliferation, migration, and invasion, by inhibiting the JAK2/STAT3 pathway.

Objective To investigate the influence of limonin on the malignant biological behavior of nonsmall cell lung cancer (NSCLC) cells by regulating the protein tyrosine kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling pathway. Methods CCK-8 method was applied to detect the survival rate of A549 cells treated with different concentrations of limonin (0, 5, 10, 25, 50, 75, 100 μmol/L). A549 cells were separated into normal culture (NC) group, low-dose limonin group (treatment with 10 µmol/L limonin for 24 h), medium-dose limonin group (treatment with 25 µmol/L limonin for 24 h), high-dose limonin group (treatment with 50 µmol/L limonin for 24 h), coumermycin A1 group (treatment with 10 μmol/L JAK2 activator coumermycin A1+50 μmol/L limonin for 24 h), and AG490 group (treatment with 10 μmol/L JAK2 inhibitor AG490+50 μmol/L limonin for 24 h). Clone formation assay was applied to detect the clones of each group of cells. Transwell assay was applied to detect cell migration and invasion, and flow cytometry was applied to detect apoptosis. Western blot analysis was applied to detect the protein expression levels of JAK2, p-JAK2, STAT3, p-STAT3, E-cadherin, N-cadherin, and vimentin in each group. Results The viability of A549 cells decreased significantly in a limonin concentration-dependent manner (P<0.05), with IC50 of 45.16±1.66 μmol/L. Concentrations of 10, 25, and 50 μmol/L were selected for subsequent experiments. The numbers of clones, migration, and invasion of A549 cells and the protein expression levels of IL-6, p-JAK2, p-STAT3, N-cadherin, and vimentin in the low-, medium-, and high-dose limonin groups significantly decreased, compared with those in the NC group, and the apoptosis rate and E-cadherin protein expression significantly increased (P<0.05). The JAK2 activator coumermycin A1 attenuated the ability of limonin to inhibit the proliferation, migration, invasion, and other malignant biological behavior of A549 cells and attenuated the apoptosis ability. The JAK2 inhibitor AG490 enhanced the ability of limonin to inhibit the proliferation, migration, invasion, and other malignant biological behavior of A549 cells and enhanced the apoptosis ability. Conclusion Limonin can inhibit the malignant biological behavior of NSCLC cells, such as proliferation, migration, and invasion, by inhibiting the JAK2/STAT3 pathway. [ABSTRACT FROM AUTHOR]

Carbonated drinks are popular due to their thirst-quenching and refreshing properties; however, they are deficient in nutritional components. An attempt has been made to develop a fruit-juice-based carbonated beverage by utilizing kinnow juice concentrate at different levels (2.5, 5.0, and 7.5%) along with sugar syrup and chilled carbonated water. Kinnow juice extracted with screw-type extractor had best quality with minimum bitterness (limonin 28 ppm) and maximum (56.50%) yield, which was concentrated from 11 to 70 ºB using rotary vacuum evaporator. Based on preliminary organoleptic trials, 5.0% juice concentrate was standardized for the formulation of carbonated beverage and shelf-life studies. Storage studies (at 30 ± 1ºC) revealed that physicochemical parameters remained unaltered, while a slight but significant (p < 0.05) decrease was observed in phytochemical parameters. Approximately 97% ascorbic acid and 95% of other bioactive compounds were found retained with high overall acceptability at the end of 4 months of storage. Also, significant decrease in CO2 content and CO2:volume was observed during storage. It can be concluded that the utilization of kinnow mandarins in carbonated beverage can provide dual benefits of providing convenient and natural flavoured carbonated drinks to the consumers and can boom the growth of the fruit-based carbonated beverage industry. [ABSTRACT FROM AUTHOR]

Background: Acute liver injury (ALI) is a common side effect of cisplatin treatment in the clinic and can lead to liver failure if not treated promptly. Previous studies have revealed that Limonin, a critical bioactive substance in citrus fruits, can protect multiple organs from various medical conditions. However, whether Limonin could ameliorate cisplatin-induced ALI remains unclear. Methods: In vivo and in vitro models were induced by cisplatin in the present study. Non-targeted metabolomics was employed to analyze the metabolic changes in the liver after ALI. In addition, molecular docking was utilized to predict the potential targets of Limonin. Results: Limonin attenuated hepatic histopathological injury by reducing hepatocyte apoptosis, lipid peroxidation, and inflammation in cisplatin-challenged mice. Employing metabolomics, we revealed that Limonin mediated the balance of various disturbed metabolic pathways in the liver after cisplatin-induced ALI. Integrating public data mining, molecular docking studies, and in vitro experiments demonstrated that Limonin suppressed the expression and activity of its direct target, 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), in the liver, thus reducing the production of corticosterone (CORT), a key metabolite promoted hepatocyte apoptosis. Conclusions: Limonin improves the liver metabolic microenvironment by inhibiting 11β-HSD1 to protect against cisplatin-induced ALI.

Background: Acute kidney injury (AKI) is a known complication of cisplatin administration; currently, there are no effective ways to prevent it. Therefore, it largely limited the use of cisplatin in chemotherapy in the clinic. In this study, we reported that Limonin, a triterpenoid compound extracted from citrus, alleviated cisplatin-induced AKI through metabolic reprogramming in the diseased kidneys. Methods: Cisplatin was employed to induce AKI in mice. Three groups were set up: Sham, cisplatin + vehicle, and cisplatin + Limonin. Using UHPLC-TOF/MS, we conducted metabolomics to profile the kidneys' endogenous metabolites and metabolic pathways. A network pharmacological method was performed to identify the targets of Limonin on AKI. The human proximal tubular epithelial cell line (HK-2) was applied for in vitro studies. Results: Limonin preserved serum creatinine and blood urea nitrogen levels after cisplatin-induced AKI. Employing metabolomics, we identified 33 endogenous differentially expressed metabolites and 7 significantly disturbed metabolic pathways in the diseased kidneys within three groups. After AKI, Limonin significantly reduced linoleic acid and its downstream product, arachidonic acid, thus exerting a protective effect on the kidney. The network pharmacological method identified CYP3A4 as a key target of Limonin in treating AKI, while CYP3A4 also serve as a mediator of arachidonic acid metabolism. In vitro, Limonin markedly reduced the level of arachidonic acid and HK-2 cell apoptosis triggered by cisplatin, mainly related to the targeted inhibition of CYP3A4-mediated arachidonic acid metabolism. Conclusion: Limonin ameliorates cisplatin-induced AKI by inhibiting CYP3A4 activity to regulate arachidonic acid metabolism, ultimately preserving kidney function.

Naringin and limonin are important antioxidant compounds in Citrus species. Due to the high biological effects of these compounds, in the present study, the variation of naringin and limonin content during fruit development was investigated at two fruit tissues of pulp and albedo in three Citrus genotypes (grapefruit, orange and mandarin). Also, the gene expression of involved enzymes in naringin and limonin metabolism was assayed. In all studied Citrus genotypes, the naringin content in albedo tissue was higher than the pulp and grapefruits showed the highest naringin content (940.26 mg per 100 g DW) followed by orange (791.17 mg per 100 g DW) and mandarin (602.10 mg per 100 g DW). The naringin content of studied genotypes was decreased during the fruit development and the highest decrement was observed at ripening stages. The decreasing of naringin content was attributed to the changes in genes expression of main involved enzymes in naringin biosynthesis pathways including chalcone-flavanone isomerase and naringenin glycoside biosynthesis (1.2 RhaT) during studied harvesting times. In both tissues of mandarin and orange fruits, the limonin content was increased up to 80 days after full bloom then decreased. In grapefruit, due to the late-ripening of the fruit, the peak of limonin increment was occurred later, which can be attributed to the long growth process of grapefruits compared to orange and mandarin. The reduction of limonin content with fruit ripening was in accordance with the increased gene expression of limonoid UDPglucosyl transferase that converts limonin to tasteless compounds. [ABSTRACT FROM AUTHOR]

E. rutaecarpa var. officinalis is a traditional Chinese medicinal plant known for its therapeutic effects, which encompass the promotion of digestion, the dispelling of cold, the alleviation of pain, and the exhibition of anti-inflammatory and antibacterial properties. The principal active component of this plant, limonin, is a potent triterpene compound with notable pharmacological activities. Despite its significance, the complete biosynthesis pathway of limonin in E. rutaecarpa var. officinalis remains incompletely understood, and the underlying molecular mechanisms remain unexplored. The main purpose of this study was to screen the reference genes suitable for expression analysis in E. rutaecarpa var. officinalis, calculate the expression patterns of the genes in the limonin biosynthesis pathway, and identify the relevant enzyme genes related to limonin biosynthesis. The reference genes play a pivotal role in establishing reliable reference standards for normalizing the gene expression data, thereby ensuring precision and credibility in the biological research outcomes. In order to identify the optimal reference genes and gene expression patterns across the diverse tissues (e.g., roots, stems, leaves, and flower buds) and developmental stages (i.e., 17 July, 24 August, 1 September, and 24 October) of E. rutaecarpa var. officinalis, LC-MS was used to analyze the limonin contents in distinct tissue samples and developmental stages, and qRT-PCR technology was employed to investigate the expression patterns of the ten reference genes and eighteen genes involved in limonin biosynthesis. Utilizing a comprehensive analysis that integrated three software tools (GeNorm ver. 3.5, NormFinder ver. 0.953 and BestKeeper ver. 1.0) and Delta Ct method alongside the RefFinder website, the best reference genes were selected. Through the research, we determined that Act1 and UBQ served as the preferred reference genes for normalizing gene expression during various fruit developmental stages, while Act1 and His3 were optimal for different tissues. Using Act1 and UBQ as the reference genes, and based on the different fruit developmental stages, qRT-PCR analysis was performed on the pathway genes selected from the "full-length transcriptome + expression profile + metabolome" data in the limonin biosynthesis pathway of E. rutaecarpa var. officinalis. The findings indicated that there were consistent expression patterns of HMGCR, SQE, and CYP450 with fluctuations in the limonin contents, suggesting their potential involvement in the limonin biosynthesis of E. rutaecarpa var. officinalis. This study lays the foundation for further research on the metabolic pathway of limonin in E. rutaecarpa var. officinalis and provides reliable reference genes for other researchers to use for conducting expression analyses. [ABSTRACT FROM AUTHOR]

BackgroundA 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.ResultHere, 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.ConclusionOur 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.

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.

Objectives Limonin has received significant attention due to its multiple biological effects, intervertebral disc degeneration (IDD) is also of interest due to the high prevalence of this disease. In this study, we determined the effects of limonin on IDD and the underlying mechanism of action to find novel ways to treat IDD. Methods An IL-1β-induced cell inflammation model and a lumbar instability model inducing IDD were established to assess the progression of IDD with or without limonin treatment. We further evaluated MAPK/NF-κB and necroptosis pathways and alterations in the extracellular matrix specific within the disc. Key findings Limonin suppresses inflammation in the nucleus pulposus in vitro by reducing the production of pro-inflammatory markers such as iNOS and COX-2. Limonin reduced the activation of the MAPK/NF-κB signalling pathway and the RIP1/RIP3/MLKL necroptosis pathway in the NP cells. Moreover, limonin delays the IDD progression in the lumbar instability model. Conclusions Limonin could potentially delay IDD by inhibiting NP cell necroptosis and modulating peripheral matrix proteins within the intervertebral disc and is a potential pharmacological research direction for the therapy in patients with IDD. [ABSTRACT FROM AUTHOR]

Background: Nonsteroidal anti‐inflammatory drugs (NSAIDs) can cause intestinal damage and ulcers and the incidence is increasing. Limonin plays an important role in the regulation of inflammatory diseases, but it has not been reported in the treatment of intestinal injury and ulcers. Methods: Indomethacin (INDO) induced intestinal injury and ulcer model in rats. The indexes related to intestinal injury were detected. Western blot and molecular docking techniques were used to detect the docking between Limonin and Nrf2. Next, ML385, an inhibitor of Nrf2/ARE signaling pathway, was applied to treat intestinal epithelial IEC‐6 cells induced by INDO. And CCK8, Western blot, TUNEL, ELISA, DCFH‐DA assay, kits, and immunofluorescence were conducted to detect cell activity, apoptosis, inflammatory response, oxidative stress, and tight junction again. Results: INDO can significantly induce intestinal ulcerative lesions in rats. Limonin could improve intestinal ulcerative lesions induced by INDO in rats. Limonin could reduce INDO‐induced inflammatory response and oxidative stress in the small intestine of rats, and improve the intestinal barrier dysfunction induced by INDO. Limonin could dock with Nrf2 structure and activate Nrf2/ARE signaling pathway. ML385 could reverse the protective effect of Limonin against INDO‐induced cell damage. Conclusion: Limonin ameliorates INDO‐induced intestinal damage and ulcers through Nrf2/ARE pathway. [ABSTRACT FROM AUTHOR]

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Limonin is a natural triterpenoid isolated from citrus fruit. In the present study, we examined the effects of limonin on cardiometabolic alterations in diet-induced metabolic syndrome. Metabolic syndrome was induced in rats by feeding them a high-fat (HF) diet plus 15% fructose in drinking water for 16 weeks. They were treated with limonin (50 or 100 mg/kg) (n = 8/group) for the final 4 weeks. Increases in body weight (BW), fasting blood glucose (FBG), serum insulin, total cholesterol (TC), blood pressure (BP), liver fat accumulation, and adipocyte hypertrophy, as well as oral glucose tolerance in rats with metabolic syndrome were alleviated by limonin treatment (p

Abstract Background Nonsteroidal anti‐inflammatory drugs (NSAIDs) can cause intestinal damage and ulcers and the incidence is increasing. Limonin plays an important role in the regulation of inflammatory diseases, but it has not been reported in the treatment of intestinal injury and ulcers. Methods Indomethacin (INDO) induced intestinal injury and ulcer model in rats. The indexes related to intestinal injury were detected. Western blot and molecular docking techniques were used to detect the docking between Limonin and Nrf2. Next, ML385, an inhibitor of Nrf2/ARE signaling pathway, was applied to treat intestinal epithelial IEC‐6 cells induced by INDO. And CCK8, Western blot, TUNEL, ELISA, DCFH‐DA assay, kits, and immunofluorescence were conducted to detect cell activity, apoptosis, inflammatory response, oxidative stress, and tight junction again. Results INDO can significantly induce intestinal ulcerative lesions in rats. Limonin could improve intestinal ulcerative lesions induced by INDO in rats. Limonin could reduce INDO‐induced inflammatory response and oxidative stress in the small intestine of rats, and improve the intestinal barrier dysfunction induced by INDO. Limonin could dock with Nrf2 structure and activate Nrf2/ARE signaling pathway. ML385 could reverse the protective effect of Limonin against INDO‐induced cell damage. Conclusion Limonin ameliorates INDO‐induced intestinal damage and ulcers through Nrf2/ARE pathway.

Random skin flap is commonly used in plastic and reconstructive surgery, however, distal part of skin flap often occurs ischemia and necrosis. Limonin, with bioactivities of anti-inflammation, anti-apoptosis and anti-oxidative stress, may be effective for skin flap survival. In our study, random flap model was performed in mice to explore the role of limonin in the survival of skin flap. On postoperative day 7, the necrosis of skin flaps was observed, while visualization of blood flow below the tissue surface was detected through Laser Doppler blood flow imaging (LDBFI). Then flap tissues were acquired to assess and levels of angiogenesis, apoptosis and oxidative stress. The results showed that limonin decreased necrosis and edema of skin flaps compared with the control group, with more blood flow in the flap under LDBFI detection. Limonin treatment also increased the mean vessels density, elevated the expression levels of angiogenic proteins (matrix metallopeptidase 9, vascular endothelial growth factor, Cadherin5) and antioxidant proteins [superoxide dismutase 1 (SOD1), endothelial nitric oxide synthase, heme oxygenase], and reduced the expression of apoptotic factors (BAX, CYC, Caspase3). In summary, limonin could effectively enhance the survival of random skin flap, the potential mechanism may attribute to the induction of angiogenesis, and inhibition of apoptosis and oxidative stress.

Limonin is a tetracyclic triterpenoid compound enriched in citrus fruits. Whether it is effective against obesity and the underlying molecular mechanisms remain unknown. Herein, a high-fat diet (HFD) was given to male C57BL/6J mice for four weeks to induce obesity. After that, limonin (50 mg/kg) was orally administered to mice for another 8 weeks along with the HFD. Limonin treatment remarkably decreased fat mass and body weight gain, and simultaneously alleviated HFD-induced fatty liver and hyperlipidemia. Limonin prevented the whitening of brown adipose tissue (BAT) and promoted the browning of inguinal white adipose tissues (iWAT) in mice. Besides, the genes associated with thermogenesisincluding Ucp1, Fatp1 and Cpt1b and mitochondrial maker TOM20 were upregulated by limonin treatment in the BAT and iWAT. The promotion of thermogenic program was associated with the activation of PGC-1α. Overall, these results demonstrate that limonin holds great potential for treating obesity by regulating energy balance.

Hair loss remains a significant problem that is difficult to treat; therefore, there is a need to identify safe natural materials that can help patients with hair loss. We evaluated the hair anagen activation effects of limonin, which is abundant in immature citrus fruits. Limonin increased the proliferation of rat dermal papilla cells (rDPC) by changing the levels of cyclin D1 and p27, and increasing the number of BrdU-positive cells. Limonin increased autophagy by decreasing phosphorylated mammalian target of rapamycin levels and increasing the phospho-Raptor, ATG7 and LC3B. Limonin also activated the Wnt/β-catenin pathway by increasing phospho-β-catenin levels. XAV939, a Wnt/β-catenin inhibitor, inhibited these limonin-induced changes, including induced autophagy, BrdU-positive cells, and cell proliferation. Limonin increased the phosphorylated AKT levels in both two-dimensional cultured rDPC and three-dimensional spheroids. Treatment with the PI3K inhibitor wortmannin inhibited limonin-induced proliferation, and disrupted other limonin-mediated changes, including decreased p27, increased BrdU-positive cells, induced autophagy, and increased ATG7 and LC3B levels. Wortmannin also inhibited limonin-induced cyclin D1 and LC3 expression in spheroids. Collectively, these results indicate that limonin can enhance anagen signaling by activating autophagy via targeting the Wnt/β-catenin and/or PI3K/AKT pathways in rDPC, highlighting a candidate nutrient for hair loss treatment. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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Lung cancer as one of the highest incident malignant tumors did not receive satisfactory chemotherapy due to lack of specific drug targets and targeted drugs. This study screened a new effective lung tumor inhibitor limonin from herbal medicine, which inhibited proliferation and promoted apoptosis of lung adenocarcinoma cells by targeting specific high expressed TMEM16A ion channel. Moreover, a novel biodegradable self-healing hydrogel was prepared from acylhydrazide functionalized carboxymethyl cellulose (CMC-AH) and oxidized pectin (pec-CHO) to reduce the side effects of the limonin to the body. The hydrogels showed fast gelation, good biocompatibility and sustained limonin release property. The limonin-loaded hydrogel significantly inhibited the growth of lung adenocarcinoma in xenografts mice because the limonin inhibited the proliferation, migration and promoted apoptosis of LA795 cells, and eliminated the acute toxicity through sustained release from the hydrogel. Combined the antitumor performance of the limonin and sustained release of pec-CHO/CMC-AH hydrogel, this limonin/hydrogel system achieved satisfactory antitumor effect and eliminated side effects in vivo. Therefore, this system has great potential application for enhanced lung adenocarcinoma therapy. • A novel tumor inhibitor of limonin was screened to target the specific high expressed TMEM16A in lung adenocarcinoma. • A fully biodegradable self-healing hydrogel was fabricated from CMC-AH and pec-CHO. • The pec-CHO/CMC-AH hydrogel showed fast gelation reaction for drug localization. • The limonin loaded hydrogel showed excellent lung tumor suppression with reduced biotoxicity. • This limonin loaded pec-CHO/CMC-AH hydrogel is highly promising in targeted long-term lung cancer therapy. [ABSTRACT FROM AUTHOR]

Formation of delayed bitterness is an intractable problem in the processing of citrus juice. This study analyzed the distribution of main bitter-causing substances in sweet orange and the effect of heat treatment, storage temperature, clarification, and homogenization on the limonin content in sweet orange juice. The segment membrane accounted for 34–42% of limonin in the fruit and 21–45% of limonoate A-ring lactone (LARL). Heat treatment accelerated the formation of limonin in juice. Pectinase treatment appeared to facilitate the formation of limonin by reducing the pH of juice. In addition, refrigeration at 4 °C was found to restrain the formation of limonin in juice by inhibiting the activity of limonin D-ring lactone hydrolase (LDLH). In conclusion, segment membrane may have a great contribution to the delayed bitterness of sweet orange juice due to its abundant LARL. These results may provide some guidance for citrus processing and bitterness removal. [Display omitted] • The segment membrane of sweet orange contained abundant LARL and limonin. • High temperature exacerbated the formation of limonin in sweet orange juice. • Refrigeration (4 ℃) inhibited the formation of limonin in sweet orange juice. • Pectinase treatment facilitated the formation of limonin by reducing the juice pH. [ABSTRACT FROM AUTHOR]

The current study focused on novel approaches for plasma-activated water treated sweet orange peel waste through the use of various food-grade debittering agents (salt, alkali, and solvent). Among the different methods investigated, solvent treatment with acetone at a ratio of 1:10 (sample: solvent, DSL 2) achieved a greater reduction in bitterness causing compounds naringin from 0.59 to 0.41 mg/g and limonin 1.30 to 1.17 mg/g. Moreover, the hurdle impact of both and debittering treatment was effective in reducing the total terpenoid content from 290.89 mg LE/100 g to 230 mg LE/100 g and anti-nutrients, tannin (289.20 mg/100 g to 109.63 mg/100 g), phytic acid (52.22 mg/100 g to 22.55 mg/100 g). The saponin was not found in the treated samples. Solvent treatment decreased total phenols from 163.46 ± 2.54 to 115.81 ± 5.02 mg GAE/100 g and total flavonoids from 662.23 ± 4.53 to 548.48 ± 2.98 mg QE/100 g. Similarly, FRAP (Ferric reducing antioxidant power) decreased from 58.87 ± 1.57 to 38.08 ± 1.96 μg AAE/g and DPPH (2, 2-Diphenyl-1-picrylhydrazyl) (%) from 151.06 ± 3.07 to 123.81 ± 2.46. Optimization of all treatment conditions was accomplished using a full factorial design. Treatment at a ratio of 1:10 (sample: salt, DS 2) was determined to be the most optimized condition, taking into account all parameters (total phenols, total flavonoids, total terpenoid content, naringin, limonin, hesperidin, antinutrients and antioxidant activities) including sensory evaluation. Debittered sweet orange peel powder obtained through this process demonstrated higher acceptability for functional and novel food product development. [Display omitted] • The utilization of sweet orange peel in conjunction with plasma-activated water (PAW) and debittering agents can transform into a product with added value. • The optimized treatment, DS 2 was determined to be the most effective in reducing bitter compounds such as Naringin and Limonin while maintaining an acceptable overall acceptability. • The DS 2 treatment proved effective in preserving phenols, flavonoids, antinutrients, and antioxidants. [ABSTRACT FROM AUTHOR]

Limonin is an intensely bitter and highly oxidized tetracyclic triterpenoid secondary metabolite, which is abundant in the Rutaceae and Meliaceae, especially in Citrus. In order to detect limonin content in complex substrates such as citrus and traditional Chinese medicine, monoclonal antibodies specifically recognizing limonin were prepared and an indirect competitive enzyme-linked immunosorbent assay (icELISA) was established. The median inhibition concentration (IC 50) was 5.40 ng/mL and the linear range was 1.25–23.84 ng/mL. The average recoveries from citrus peel and pulp samples were 95.9%–118.8% and 77.5%–113.1%, respectively. Moreover, the contents of limonin in 6 citrus samples and 4 herbal samples were analyzed by icELISA and UPLC-MS, and the results of the two methods were consistent. This validation is sufficient to demonstrate that the developed immunoassay is applicable for the detection of limonin in citrus and herbal samples and has the advantage of high efficiency, sensitivity, and convenience. • A sensitive and specific mAb 2E10 against limonin was prepared, which had no cross-reactivity with obacunon and nomilin. • A sensitive icELISA for limonin was developed, and its detection results were consistent with UPLC-MS/MS method. • The developed icELISA can be used for the detection of limonin in complex matrix samples such as citrus and herbal. [ABSTRACT FROM AUTHOR]

E. rutaecarpa var. officinalis is a traditional Chinese medicinal plant known for its therapeutic effects, which encompass the promotion of digestion, the dispelling of cold, the alleviation of pain, and the exhibition of anti-inflammatory and antibacterial properties. The principal active component of this plant, limonin, is a potent triterpene compound with notable pharmacological activities. Despite its significance, the complete biosynthesis pathway of limonin in E. rutaecarpa var. officinalis remains incompletely understood, and the underlying molecular mechanisms remain unexplored. The main purpose of this study was to screen the reference genes suitable for expression analysis in E. rutaecarpa var. officinalis, calculate the expression patterns of the genes in the limonin biosynthesis pathway, and identify the relevant enzyme genes related to limonin biosynthesis. The reference genes play a pivotal role in establishing reliable reference standards for normalizing the gene expression data, thereby ensuring precision and credibility in the biological research outcomes. In order to identify the optimal reference genes and gene expression patterns across the diverse tissues (e.g., roots, stems, leaves, and flower buds) and developmental stages (i.e., 17 July, 24 August, 1 September, and 24 October) of E. rutaecarpa var. officinalis, LC-MS was used to analyze the limonin contents in distinct tissue samples and developmental stages, and qRT-PCR technology was employed to investigate the expression patterns of the ten reference genes and eighteen genes involved in limonin biosynthesis. Utilizing a comprehensive analysis that integrated three software tools (GeNorm ver. 3.5, NormFinder ver. 0.953 and BestKeeper ver. 1.0) and Delta Ct method alongside the RefFinder website, the best reference genes were selected. Through the research, we determined that Act1 and UBQ served as the preferred reference genes for normalizing gene expression during various fruit developmental stages, while Act1 and His3 were optimal for different tissues. Using Act1 and UBQ as the reference genes, and based on the different fruit developmental stages, qRT-PCR analysis was performed on the pathway genes selected from the “full-length transcriptome + expression profile + metabolome” data in the limonin biosynthesis pathway of E. rutaecarpa var. officinalis. The findings indicated that there were consistent expression patterns of HMGCR, SQE, and CYP450 with fluctuations in the limonin contents, suggesting their potential involvement in the limonin biosynthesis of E. rutaecarpa var. officinalis. This study lays the foundation for further research on the metabolic pathway of limonin in E. rutaecarpa var. officinalis and provides reliable reference genes for other researchers to use for conducting expression analyses.

Khasi Mandarin (Citrus reticulata) is a unique fruit usually cultivated in high terrains regions of North-East India. They are different from ordinary orange in terms of its biochemical content (limonin, secondary metabolites), pharmacological activities, and physicochemical appearance (bright orange in color, hard peel, sweet aromatic juice). It has high nutritional values (NV) by enriching beta carotene as powerful antioxidant for scavenging free radicals, magnesium for blood pressure regulation, potassium for healthy cardio vascular functions, thiamin for energy conversion from food and vitamin C as immune enhancer. Despite its NV, limonin an aglycones and glycosides are largely available in Khasi Mandarin. Presence of this compound in Khasi Mandarin display numerous pharmacological activities like antimicrobial, antioxidant, anticancer, antidiabetic, anti-inflammatory, analgesic, antibacterial, anti-virus, liver protection and pesticidal. In this review, we discussed not only about the morphological aspects but detailed focused has been given on the nutritional value of Khasi mandarin, post-harvest disease and its management, metabolism and bioactive functions of the limonin as therapeutic applications.

Background and Purpose: Limonin, a naturally occurring tetracyclic triterpenoid, has extensive pharmacological effects. Its role in cardiac hypertrophy remains to be elucidated. We investigated its effects on cardiac hypertrophy along with the potential mechanisms involved.Experimental Approach: The effects of limonin on cardiac hypertrophy in C57/BL6 mice caused by aortic banding, plus neonatal rat cardiac myocytes (NRCMs) stimulated with phenylephrine to induce cardiomyocyte hypertrophy in vitro were investigated.Key Results: Limonin markedly improved the cardiac function and heart weight in aortic banded mice. Limonin-treated mice and NRCMs also produced fewer cardiac hypertrophy markers than those treated with the vehicle in the hypertrophic groups. Sustained aortic banding- or phenylephrine-stimulation impaired cardiac sirtuin 6 (SIRT6) protein levels, which were partially reversed by limonin associated with enhanced activity of PPARα. Sirt6 siRNA inhibited the anti-hypertrophic effects of limonin in vitro. Interestingly, limonin did not influence Sirt6 mRNA levels, but regulated ubiquitin levels. Thus, the protein biosynthesis inhibitor, cycloheximide and proteasome inhibitor, MG-132, were used to determine SIRT6 protein expression levels. Under phenylephrine stimulation, limonin increased SIRT6 protein levels in the presence of cycloheximide, but it did not influence SIRT6 expression in the presence of MG-132, suggesting that limonin promotes SIRT6 levels by inhibiting its ubiquitination degradation. Furthermore, limonin inhibited the degradation of SIRT6 by activating ubiquitin-specific peptidase 10 (USP10), while Usp10 siRNA prevented the beneficial effects of limonin.Conclusion and Implications: Limonin mediates the ubiquitination and degradation of SIRT6 by activating USP10, providing an attractive therapeutic target for cardiac hypertrophy. [ABSTRACT FROM AUTHOR]

Limonin is one of the research hotspots in natural drug development. However, its low solubility in water leads to poor oral bioavailability, discouraging the further study of its potential as a candidate compound. In order to overcome this limitation, and to enhance its biological activities, a novel limonin derivative—HY-071085—was synthesized by structural modification, and has exhibited strong anti-inflammatory and analgesic activity. In order to achieve a thorough understanding of the biological actions of HY-071085 in vivo, this study evaluated the pharmacokinetics and bioavailability of HY-071085 in rats and beagle dogs, and the distribution and excretion in rats. Using ultra-high-performance liquid chromatography-tandem mass spectrometry, the kinetic profiles of HY-071085 in the plasma of healthy rats and beagle dogs after a single gavage, repeated gavages and the intravenous injection of HY-071085 were studied. The tissue distribution (heart, liver, spleen, lung, kidney, gastric tissue, intestine, brain, skin, testis, ovary and womb) and excretion of HY-071085 were also studied. These results showed that HY-071085 has nonlinear dynamic characteristics in rat and beagle dog plasma. It was found that the plasma concentrations of HY-071085 in female rats were significantly higher than those in male rats after a single oral administration. There were gender differences in the kinetic behavior of HY-071085 in rats; however, there was no difference identified in dogs. HY-071085 was mainly eliminated as metabolites in rats, and was distributed in most of the tissues except the brain, with the highest content being in the gastric tissue and intestinal arease, followed by the liver, spleen, fat, lung, kidney, ovary and heart. The bioavailability of HY-071085 in male and female rats was 2.8% and 10.8%, respectively, and was about 13.1% in beagle dogs. The plasma protein binding rate of HY-071085 in rats, beagle dogs and humans ranged from 32.9% to 100%, with obvious species differences. In conclusion, our study provides useful information regarding the absorption, distribution and excretion of HY-071085, which will provide a good base for the study of the mechanism of its biological effects. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Citrus fruits are grown commercially throughout the world. They are widely consumed due to their nutrients, use in energy supplements, and numerous health benefits. There is significant interest among consumers about this naturally available source, rich in flavonoids and antioxidants. However, underutilized citrus varieties remain unexplored due to the lack of information about the pool of nutritive properties they confer. Ten underutilized citrus varieties were collected from Nokrek Biosphere Reserve, West Garo Hills, Meghalaya, India, identified by UNESCO as a Biosphere reserve, to study the diversity in terms of limonin, ascorbic acid, carotenoids, browning, flavonoids, total phenol, and antioxidant activity, the contents of which varied significantly among different citrus cultivars. The results indicated that Citron and Pomelo were good sources of ascorbic acid (29.50 and 45.09 mg/100 mL), and that Khasi papeda was found to contain lower limonin content (9.21 ppm). However, in terms of flavonoids, Khasi papeda and Pomelo were found to exhibit a higher naringin content (189.13 ppm and 32.15 ppm), whereas the hesperidin content was highest in Kachai lemon, Khasi papeda, and Chinotto, at 199.51 ppm, 148.04 ppm, and 135.88 ppm, respectively. Antioxidant activity was assessed by three antioxidant assays (ABTS+ (radical cation azino-bis [3-ethylbenzthiazoline-6-sulfonic acid]) (ABTS), 2,2-diphenyl-1-picrylhydrazyl radical (DPPH), and Ferric Reducing Antioxidant Power (FRAP)). Khasi papeda (7.48 mM L−1 Trolox), Chinotto (7.48 mM L−1 Trolox), and Pomelo (7.48 mM L−1 Trolox) exhibited the highest reducing power with DPPH radical scavenging activity, and Khasi papeda (15.41 mg GAE L−1) possessed a higher phenolic content, whereas the antioxidant activity when assessed with ABTS and FRAP assays was highest among the underutilized species of Khasi papeda (4.84 mM L−1 Trolox, 1.93 mM L−1 Trolox) and Ada Jamir (4.96 mM L−1 Trolox, 2.03 mM L−1 Trolox), respectively. To the best of our knowledge, this is among the very few papers presenting comprehensive data on the metabolic diversity of flavonoids and antioxidant potential to characterize the underutilized citrus species. This study also demonstrated that Khasi papeda, Pomelo, Chinotto, and Kachai lemon can serve as potential sources of functional components, bioactive compounds, and antioxidants, which can be explored for further application in the processing industry for nutritional security. [ABSTRACT FROM AUTHOR]

Abstract Limonin is an important bioactive substance in citrus fruits, especially in seeds, which has great potential in cancer prevention and treatment. In order to explore the anticancer activity based on interaction between limonin and NQO1, Human promyelocytic leukemia cells (HL‐60) were studied in vitro. We found that limonin could inhibit proliferation and promote apoptosis of HL‐60 cells, and the effect was positively correlated with its dosage. Western blot results showed that limonin could activate the endogenous apoptosis pathway mediated by mitochondria via up‐regulating pro‐apoptotic proteins (Bax, cytochrome c, Caspase3, and Caspase9) and down‐regulating anti‐apoptotic proteins (Bcl‐2), thus inhibiting the proliferation of HL‐60 cells and promoting apoptosis, which further proved the anticancer activity of limonin from the molecular mechanism. At the same time, limonin down‐regulated the expression of NQO1, indicating that limonin may indirectly act on the apoptosis pathway by regulating the expression activity of antioxidant enzymes in vivo, thus exerting its inhibitory effect on tumor cells, which provides an idea for the molecular mechanism that natural products can indirectly exert their anticancer effect by regulating the activity of antioxidant enzymes.

[Display omitted] • The effects of berberine/curcumin on HSA-limonin interactions were first compared. • Cell experiments showed berberine and curcumin affect HSA-limonin binding affinity. • Provided insights into HSA's role as a multi-drug transporter. The competition among drugs for binding to plasma proteins is regarded as a pharmacokinetic drug interaction. Competition between antitumor agents and other drugs for plasma protein binding can alter the free concentration of the drug, potentially impacting its efficacy and increasing the risk of toxic side effects. Through a range of spectroscopic techniques, this study examined the interaction between limonin and human serum albumin (HSA) in the context of berberine (Ber) and curcumin (Cur) under physiological conditions to clarify the binding mechanisms of binary and ternary systems at the molecular level. As demonstrated by fluorescence quenching experiments, Static quenching was identified as the mechanism of interaction between HSA and limonin. The results of site competition experiments indicated that the binding site between limonin and HSA was site I, a result further supported by molecular docking simulations. Through the use of thermodynamic data calculations, it was determined that limonin forms a stable complex with HSA by establishing hydrogen bonds and van der Waals forces. Circular dichroism (CD) spectroscopy, three-dimensional (3D) fluorescence spectroscopy, and synchronous fluorescence spectroscopy (SFS) employed to validate the notion that limonin perturbed the microenvironment of amino acids and induced conformational changes in HSA. What's more, the presence of Ber or Cur was found to have further modified the alterations observed in the interaction between the original HSA-limonin binary system. In vitro cellular experiments showed that interaction with HSA reduced the antitumor activity of limonin. In contrast, adding Ber or Cur increased the inhibition rate of tumor cells. The coexistence of both Ber and Cur significantly diminished limonin's binding affinity to HSA. The current investigation enhances comprehension regarding the binding characteristics and interaction mechanisms involving limonin, Ber, Cur, and HSA. It explores the potential of HSA as a versatile drug carrier and furnishes theoretical underpinnings for co-administrative strategies. [ABSTRACT FROM AUTHOR]

Plant natural products provide considerable resources for the innovation of agrochemicals or pharmaceuticals due to their extensive biological potential and scaffold diversity. Limonin, a naturally occurring tetracyclic triterpenoid found in non-edible wastes of the citrus industry is a promising lead for developing new pesticide candidates. Reported herein is the limonin-directed analogues with scaffold diversity and their agricultural insecticidal activity evaluation. Pesticidal bioassay results revealed that 29 target compounds displayed varying degrees of enhanced larvicidal activity against Plutella xylostella Linnaeus compared to their precursor limonin. Among them, compounds IIId , Vd , VIb , VId , VIIa , VIId , and VIIf exhibited more pronounced insecticidal potential with LC 50 values of 6.97, 10.07, 13.50, 13.74, 7.43, 8.63, and 12.82 μg/mL, respectively, far exceeding that of limonin (LC 50 = 110.67 μg/mL) and the two positive controls rotenone (LC 50 = 33.29 μg/mL) and toosendanin (LC 50 = 31.60 μg/mL). Notably, the larvicidal activity of IIId , VIIa , and VIId against P. xylostella was greater than 15.8-, 14.8-, and 12.8-fold that of limonin, respectively. This project also disclosed that 27 target compounds demonstrated insecticidal activity against Mythimna separata Walker superior to limonin. Compounds IIIa , IIIc , IIIf , Vd , VIa , and VIId showed potent growth inhibitory activity against M. separata with LC 50 values of 0.63, 0.42, 0.51, 0.69, 0.43, and 0.71 mg/mL, respectively, whereas the LC 50 value of limonin was 1.20 mg/mL. It was illustrated that these heterocyclic scaffolds played an important role in improving the insecticidal performance of limonin. Further cytotoxicity assay indicated that the tested compounds showed almost no cytotoxicity to human keratinocyte cell line (HaCaT), thus showing a good safety profile. Overall, this work will provide valuable insight into agrochemical innovation based on limonin and will also promote the industrial value of limonoids from citrus industrial wastes. [Display omitted] • Limonin-type oxime ether derivatives with scaffold diversity were constructed by a combinatorial synthesis strategy. • Compounds IIId , Vd , VIb , VId , VIIa , VIId , and VIIf were found to be potent against Plutella xylostella Linnaeus. • Analogues IIIa , IIIc , IIIf , Vd , VIa , and VIId exhibited pronounced insecticidal activity against Mythimna separata Walker. • The results will be beneficial for promoting the redevelopment and utilization of citrus industrial wastes. [ABSTRACT FROM AUTHOR]

Pomelo seed as a by-product from pomelo consumption is rich in bioactive compounds, however, a huge volume of pomelo seed was disposed as wastes, the comprehensive utilization of pomelo seed could not only generate valued-added products/ingredients, but also decrease the environmental pollution. In this study, the main active substance limonin in pomelo seed was considered as a high-value bioactive compound. The purification of limonin from pomelo seed was investigated, and the neuroprotective and mechanism were characterized. The UPLC-MS/MS results indicated that 29 compounds in pomelo seed were identified, including 14 flavonoids, 3 limonids, 9 phenols and 3 coumarins. Moreover, high purity of limonin was obtained by crystallization and preparative-HPLC. Furthermore, limonin pretreatment can antagonize the cell damage mediated by Aβ25−35 in a concentration-dependent relationship. The regulation of Bax/Bcl-2, expression of caspase-3 protein and the activation of PI3K/Akt signaling pathway were observed in the cells pretreated with limonin. Treatment of PC12 cells with PI3K inhibitor LY294002 weakened the protective effect of limonin. These results indicated that limonin prevented Aβ25−35-induced neurotoxicity by activating PI3K/Akt, and further inhibiting caspase-3 and up-regulating Bcl-2. This study enables comprehensive utilization of pomelo seed as by-product and offers a theoretical principle for a waste-to-wealth solution, such as potential health benefits of food ingredient and drug.

A potential bioactive compound limonin is found in citrus fruits, especially tremendous extent in seeds. In this study, an attempt has been made to develop a novel and proficient extraction method by using the hydrotrope solution to recover limonin from lemon (Citrus limon L.) seeds. Initially, the extraction time (4 h) was screened by varying the time from 3 to 6 h by fixing the concentration of hydrotrope (sodium salicylate), raw material loading and temperature of the extraction. To enhance the yield of hydrotrope extraction, Box–Behnken experimental design (BBD) was proposed. The maximum yield of limonin recovered was 6.41 mg g−1 at optimized process conditions such as the hydrotrope concentration of 1.65 M, raw material loading of 8.08%, and temperature of 44°C. The obtained limonin yield in this study was higher compared to the other studies using different citrus seeds. The recovered limonin was identified by using Fourier Transform-Infrared Spectroscopy (FTIR) and high-performance liquid chromatography (HPLC). Furthermore, the scavenging ability was evaluated for the extracted limonin, and it is proved to be a potent antioxidant. Therefore, the results revealed that the extraction of bioactive compounds using hydrotrope is efficient, eco-friendly and can also lower the usage of organic chemicals. [ABSTRACT FROM AUTHOR]

NAFLD is the most prevalent liver disease in human history. The treatment is still limited yet. In the current study, we reported that limonin inhibited hepatic lipid accumulation and fatty acid synthesis in HFD fed mice. Using AMPK inhibitor and AMPK deficient C. elegans , we revealed the effect was dependent on the activation of AMPK. We found that limonin activated AMPK through inhibition of cellular energy metabolism and increasing ADP:ATP ratio. Furthermore, the treatment of limonin induced AMPK mediated suppression of the transcriptional activity of SREBP1/2. Our study suggests that limonin may a promising therapeutic agent for the treatment of NAFLD. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease and continues to rise in the worldwide. Limonin is a triterpenoid compound widely found in the fruits of citrus plants with a wide range of pharmacological effects, including anti-cancer, anti-inflammation, anti-viral, anti-oxidation and liver protection properties. However, the potential molecular mechanism of limonin on NAFLD in zebrafish remains unknown. In this study, zebrafish larvae were exposed to thioacetamide to establish an NAFLD model and the larvae were treated with limonin for 72 h simultaneously. The human liver cell line was stimulated with lipid mixture and meanwhile incubated with limonin for 24 h. The results showed that Limonin significantly reduced the accumulation of lipid droplets in the liver and down-regulated the levels of lipogenic transcription factors FASN and SREBP1 in NAFLD. Limonin suppressed macrophages infiltration and the down-regulated the relative expression levels of the pro-inflammatory factors IL-6, IL-1β and TNF-α secreted by macrophages. Besides, limonin could reversed the reduction of glutathione and the accumulation of reactive oxygen species through up-regulating NRF2/HO-1 signaling pathway in the liver. In conclusion, this study revealed that limonin has a protective effect on NAFLD due to its resistance to lipid deposition as well as antioxidant and anti-inflammatory actions. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease and continues to rise in the worldwide. Limonin is a triterpenoid compound widely found in the fruits of citrus plants with a wide range of pharmacological effects, including anti-cancer, anti-inflammation, anti-viral, anti-oxidation and liver protection properties. However, the potential molecular mechanism of limonin on NAFLD in zebrafish remains unknown. In this study, zebrafish larvae were exposed to thioacetamide to establish an NAFLD model and the larvae were treated with limonin for 72 h simultaneously. The human liver cell line was stimulated with lipid mixture and meanwhile incubated with limonin for 24 h. The results showed that Limonin significantly reduced the accumulation of lipid droplets in the liver and down-regulated the levels of lipogenic transcription factors FASN and SREBP1 in NAFLD. Limonin suppressed macrophages infiltration and the down-regulated the relative expression levels of the pro-inflammatory factors IL-6, IL-1β and TNF-α secreted by macrophages. Besides, limonin could reversed the reduction of glutathione and the accumulation of reactive oxygen species through up-regulating NRF2/HO-1 signaling pathway in the liver. In conclusion, this study revealed that limonin has a protective effect on NAFLD due to its resistance to lipid deposition as well as antioxidant and anti-inflammatory actions.

NAFLD is the most prevalent liver disease in human history. The treatment is still limited yet. In the current study, we reported that limonin inhibited hepatic lipid accumulation and fatty acid synthesis in HFD fed mice. Using AMPK inhibitor and AMPK deficient C. elegans, we revealed the effect was dependent on the activation of AMPK. We found that limonin activated AMPK through inhibition of cellular energy metabolism and increasing ADP:ATP ratio. Furthermore, the treatment of limonin induced AMPK mediated suppression of the transcriptional activity of SREBP1/2. Our study suggests that limonin may a promising therapeutic agent for the treatment of NAFLD.

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‐1β, 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. [ABSTRACT FROM AUTHOR]

The bitterness of citrus juice is one of the most important factors in the reduction of marketability in juice industry. Limonin and Narengin are known as two important compounds in bitter citrus juice during storage. Therefore, various processes, including chemical, microbial and enzymatic methods, have been used to reduce these bitter compounds. The purpose of this study was to investigate the effect of adding Pseudomonas putida bacteria on the reduction of the level of limonin and also the measurement of physicochemical and organoleptic properties. Pseudomonas putida bacteria with two concentrations of 102 and 104 cfu/mL as a causative agent of orange and grapefruit concentrate were studied. Also, the physicochemical properties of concentrates were measured at 120 h (growth time and bacterial activity). The results showed that the growth rate of bacteria had a significant effect on the reduction of lemonin, pH and turbidity levels (p < /em>

Abstract Background Ovarian cancer therapy generally involves systemic chemotherapy with anticancer drugs; however, chemotherapy with a platinum-based drug has often been shown to cause adverse reactions and drug resistance in ovarian cancer patients. Evodia rutaecarpa (ER) reportedly shows anticancer activity against various types of cancer cells. However, the effects of ER have not yet been fully uncovered in ovarian cancer. Methods In the present study, we investigated the anticancer effects of an ER extract and its components against the ovarian cancer cell lines SKOV-33, A2780, RMUG-S and a cisplatin-resistant SKOV-3 cell line (CisR SKOV-3). Cell viability and colony formation assays along with subcellular fractionation analysis, immunoblotting, and immunofluorescence staining were performed. Results ER treatment led to a significant reduction in the viability of SKOV-3 cells. Moreover, limonin, a compound found in ER, reduced the viability of both serous-type (SKOV-3 and A2780) and mucinous-type (RMUG-S) ovarian cancer cells by inducing apoptosis via activation of the p53 signaling pathway. Furthermore, limonin reversed the drug resistance through activation of apoptosis in CisR SKOV-3. Conclusion Taken together, our findings suggest that limonin contributes to the anti-ovarian cancer effects of ER by inducing apoptosis via activation of the p53 signaling pathway.

Background: Myocardial infarction (MI) resulting from acute coronary ischemia may cause significant morbidity and mortality, and microRNAs play a vital role in this pathophysiology. Limonin (LIM) is a natural medicine from citrus fruit that protects organs against ischemic diseases, but the candidate genes and pathways associated with cardioprotection are unknown. Methods: MI was induced by ligating the left anterior descending coronary in male Sprague-Dawley rats. LIM was orally administered for 7 days after the induction of MI. Subsequently, the hearts were collected to examine significant changes in microRNAs and mRNAs among the control (CON), MI, and LIM + MI groups. Gene Ontology (GO) terms, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and protein-protein interaction (PPI) networks were used to identify the biological functions and signaling pathways of differentially expressed mRNAs. Candidate genes were validated by RT-qPCR. Results: Compared to the CON group, MI caused significant changes in the expression of 26 microRNAs and 1979 mRNAs. The bioinformatics analysis showed that inflammation, apoptosis, and oxidation were enriched in GO terms, while RAP1, PI3K/AKT, RAS, and cGMP-PKG were enriched in KEGG pathways. In addition, compared to the MI group, LIM induced significant changes in the expression of 4 microRNAs and 173 mRNAs. The differentially expressed mRNAs were related to collagen biosynthesis, the immune response, extrinsic apoptosis, and tight junctions. One microRNA (rno-miR-10a-5p) and 2 mRNAs (IGLON5 and LMX1A) were differentially expressed among the CON, MI, and LIM + MI groups. Conclusions: Our results suggest that the rno-miR-10a-5p-IGLON5/LMX1A axis may be a candidate pathway and promising target through which LIM alleviates MI-induced cardiac dysfunction.