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Oxidative stress significantly contributes to ageing and disease, with antioxidants holding promise in mitigating its effects. Functional foods rich in flavonoids offer a potential strategy to mitigate oxidative damage by free radicals. We investigated the protective effects of mulberry leaf flavonoids (MLF) against H2O2-induced oxidative damage in HepG2 cells. It assessed the inhibitory effect of MLF (62.5–500 μg/mL) on H2O2-induced oxidative damage by analyzing cellular morphology and oxidative stress markers, including ROS production, mitochondrial membrane potential, antioxidant enzyme levels, MDA, and apoptosis-related proteins. The results demonstrated that MLF prevented spiny cell formation triggered by 750 μM H2O2 and significantly reduced ROS levels, restored mitochondrial membrane potential, decreased lactate dehydrogenase and alanine transaminase leakage, and reduced MDA content induced by H2O2. MLF also modulated antioxidant enzymes and attenuated oxidative damage to HepG2 cell DNA, as confirmed by staining techniques. These findings indicate the potential of MLF as a hepatoprotective agent against oxidative damage in HepG2 cells. [ABSTRACT FROM AUTHOR]

IntroductionThis study aimed to investigate the regulatory effects of mulberry leaf flavonoids and carnosic acid complex (MCC) on the growth performance, intestinal morphology, antioxidant, and p38 MAPK/Nrf2 pathway in broilers.MethodsA total of 256 healthy 8-day-old female yellow-feathered broilers were randomly divided into 4 equal groups: a control group (CON) fed a basal diet, an antibiotic group (CTC) supplemented with 50 mg/kg chlortetracycline, and two experimental groups (MCC75, MCC150) fed basal diets with 75 mg/kg and 150 mg/kg of MCC, respectively. The experiment lasted for 56 days, with days 1–28 designated as the initial phase and days 29–56 as the growth phase.ResultsThe results on the growth performance showed that diets supplemented with MCC and CTC decreased the feed-to-gain ratio (F/G), diarrhea rate, and death rate, while significantly increasing the average daily weight gain (ADG) (p

Introduction: Horses are susceptible to oxidative stress during strenuous endurance exercise, leading to muscle fatigue and damage. Mulberry leaf flavonoids (MLFs) possess significant antioxidant properties. However, the antioxidant efficacy of MLFs can be influenced by the extraction process, and their impact on H2O2-induced oxidative stress in equine skeletal muscle satellite cells (ESMCs) remains unexplored.Methods: Our study employed three extraction methods to obtain MLFs: ultrasound-assisted extraction (CEP), purification with AB-8 macroporous resin (RP), and n-butanol extraction (NB-EP). We assessed the protective effects of these MLFs on H2O2-induced oxidative stress in ESMCs and analyzed the MLF components using metabolomics.Results: The results revealed that pre-treatment with MLFs dose-dependently protected ESMCs against H2O2-induced oxidative stress. The most effective concentrations were 0.8 mg/mL of CEP, 0.6 mg/mL of RP, and 0.6 mg/mL of NB-EP, significantly enhancing EMSC viability (p < 0.05). These optimized MLF concentrations promoted the GSH-Px, SOD and T-AOC activities (p < 0.05), while reducing MDA production (p < 0.05) in H2O2-induced ESMCs. Furthermore, these MLFs enhanced the gene expression, including Nrf2 and its downstream regulatory genes (TrxR1, GPX1, GPX3, SOD1, and SOD2) (p < 0.05). In terms of mitochondrial function, ESMCs pre-treated with MLFs exhibited higher basal respiration, spare respiratory capacity, maximal respiration, ATP-linked respiration compared to H2O2-induced ESMCs (p < 0.05). Additionally, MLFs enhanced cellular basal glycolysis, glycolytic reserve, and maximal glycolytic capacity (p < 0.05). Metabolomics analysis results revealed significant differences in mulberrin, kaempferol 3-O-glucoside [X-Mal], neohesperidin, dihydrokaempferol, and isobavachalcone among the three extraction processes (p < 0.05).Discussion: Our study revealed that MLFs enhance antioxidant enzyme activity, alleviate oxidative damage in ESMCs through the activation of the Nrf2 pathway, and improve mitochondrial respiration and cell energy metabolism. Additionally, we identified five potential antioxidant flavonoid compounds, suggesting their potential incorporation into the equine diet as a strategy to alleviate exercise-induced oxidative stress.

Mulberry (Morus alba L.) leaf is a well-established traditional Chinese botanical and culinary resource. It has found widespread application in the management of diabetes. The bioactive constituents of mulberry leaf, specifically mulberry leaf flavonoids (MLFs), exhibit pronounced potential in the amelioration of type 2 diabetes (T2D). This potential is attributed to their ability to safeguard pancreatic β cells, enhance insulin resistance, and inhibit α-glucosidase activity. Our antecedent research findings underscore the substantial therapeutic efficacy of MLFs in treating T2D. However, the precise mechanistic underpinnings of MLF's anti-T2D effects remain the subject of inquiry. Activation of brown/beige adipocytes is a novel and promising strategy for T2D treatment. In the present study, our primary objective was to elucidate the impact of MLFs on adipose tissue browning in db/db mice and 3T3-L1 cells and elucidate its underlying mechanism. The results manifested that MLFs reduced body weight and food intake, alleviated hepatic steatosis, improved insulin sensitivity, and increased lipolysis and thermogenesis in db/db mice. Moreover, MLFs activated brown adipose tissue (BAT) and induced the browning of inguinal white adipose tissue (IWAT) and 3T3-L1 adipocytes by increasing the expressions of brown adipocyte marker genes and proteins such as uncoupling protein 1 (UCP1) and beige adipocyte marker genes such as transmembrane protein 26 (Tmem26), thereby promoting mitochondrial biogenesis. Mechanistically, MLFs facilitated the activation of BAT and the induction of WAT browning to ameliorate T2D primarily through the activation of AMP-activated protein kinase (AMPK)/sirtuin 1 (SIRT1)/peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) signaling pathway. These findings highlight the unique capacity of MLF to counteract T2D by enhancing BAT activation and inducing browning of IWAT, thereby ameliorating glucose and lipid metabolism disorders. As such, MLFs emerge as a prospective and innovative browning agent for the treatment of T2D. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Food Safety & Quality is the property of Journal of Food Safety & Quality Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Oxidative stress significantly contributes to ageing and disease, with antioxidants holding promise in mitigating its effects. Functional foods rich in flavonoids offer a potential strategy to mitigate oxidative damage by free radicals. We investigated the protective effects of mulberry leaf flavonoids (MLF) against H2O2-induced oxidative damage in HepG2 cells. It assessed the inhibitory effect of MLF (62.5–500 μg/mL) on H2O2-induced oxidative damage by analyzing cellular morphology and oxidative stress markers, including ROS production, mitochondrial membrane potential, antioxidant enzyme levels, MDA, and apoptosis-related proteins. The results demonstrated that MLF prevented spiny cell formation triggered by 750 μM H2O2 and significantly reduced ROS levels, restored mitochondrial membrane potential, decreased lactate dehydrogenase and alanine transaminase leakage, and reduced MDA content induced by H2O2. MLF also modulated antioxidant enzymes and attenuated oxidative damage to HepG2 cell DNA, as confirmed by staining techniques. These findings indicate the potential of MLF as a hepatoprotective agent against oxidative damage in HepG2 cells.

Introduction: The incidence of inflammatory bowel disease (IBD) has been increasing year by year in recent years. Flavonoids are the main components of mulberry leaves exerting anti-inflammatory effects and have potential applications in drug screening for IBD treatment. Enzymatic hydrolysis can enhance the bioavailability and activity of flavonoid glycosides. No relevant reports on the potentiation of mulberry leaf flavonoids (MLF) after deglycosylation have been retrieved., Methods: An enzymatic method was used to prepare enzymatic hydrolysates of MLF (EMLF). The protective effect of EMLF on zebrafish with IBD was evaluated by observing zebrafish intestinal length and width, intestinal histopathological morphology, as well as neutrophil and goblet expression. Network pharmacology and metabolomics were performed to explore the mechanism of action of EMLF against IBD. Finally, the mechanism of EMLF against IBD was validated using Western Blot and real-time quantitative polymerase chain reaction (RT-qPCR)., Results: The EMLF was successfully prepared by hydrolyzing MLF with snailase for the first time, and the anti-inflammatory effect of EMLF was clarified to be superior to that of MLF in IBD zebrafish. The network pharmacology and metabolomics studies showed that the mechanism of EMLF for IBD may be related to regulation of purine metabolic pathway. The expression levels of adenosine deaminase (ADA), critical targets in purine metabolism, were significantly down-regulated, while the expression of adenine phosphoribosyltransferase (APRT) and xanthine dehydrogenase (XDH) was significantly increased when compared with the TNBS group., Conclusion: The results suggested that enzymatic deglycosylation is an effective measure to enhance the anti-inflammatory activity of MLF, which provides referable ideas and methods for the deep processing and utilization of natural drug resources., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Ethnopharmacological Relevance: Mulberry leaf (Folium Mori) is a dried leaf of the dicotyledonous mulberry tree and is a homologous food and medicine. Treating insomnia with it is a common practice in traditional Chinese medicine. But still, its potential sleep-improving mechanism remains to be elucidated., Aim of Review: Potential bioactive components and mechanisms of the sleep-improving effect of purified flavone from mulberry leaves (MLF) were explored through in vivo experiments, network pharmacology analysis, and molecular experimental validation., Materials and Methods: The mice model was established by pentobarbital sodium induction to evaluate the sleep-improving effect of MLF. The MLF's chemical composition was identified through a liquid chromatograph quadrupole time-of-flight mass spectrometer (Q-TOF LC/MS) to elucidate its sleep-improving active ingredient. At last, the underlying mechanism of MLF's sleep-improving effect was elucidated through neurotransmitter detection (ELISA), network pharmacology analysis, and molecular experimental validation (quantitative real-time PCR and western blotting)., Results: MLF could dramatically reduce sleep latency by 35%, prolong sleep duration by 123%, and increase the sleep rate of mice through increasing γ-aminobutyric acid (GABA) and serotonin (5-HT) release in serum, hypothalamus, and hippocampus. Q-TOF LC/MS identified 17 flavonoid components in MLF. Network pharmacological analysis suggested that the key sleep-improving active ingredients in MLF might be quercetin, kaempferol, morin, and delphinidin. The key path for MLF to improve sleep might be the tryptophan metabolism and neuroactive ligand-receptor interaction, and the key targets might be gamma-aminobutyric acid type A receptor subunit alpha2 Gene (GABRA2) and serotonin 1A (5-HT1A) receptors., Conclusions: MLF has shown significant sleep-improving effects in mice and may take effect through regulating the GABA and 5-HT receptors., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

To study the effect of mulberry leaf flavonoids on myocardial mitochondrial function and myocardial fibrosis in diabetic mice, and to explore the specific molecular mechanism of its effect. 45 ICR mice were randomly divided into 3 groups, namely normal group, model group and mulberry leaf flavonoid group. The model group and the mulberry leaf flavonoids group were intraperitoneally injected with alloxan saline solution to establish a diabetic model, and the mice in the normal group were intraperitoneally injected with saline. The diabetic mice in the mulberry leaf flavonoid group were treated with mulberry leaf flavonoids (1.0 g/kg/day) by gavage after the model was successfully established, and the normal group and the model group were treated with the same amount of normal saline. After 6 weeks of treatment, the levels of blood glucose, blood insulin, liver glycogen, liver hexokinase (HK) and pyruvate kinase (PK), myocardial mitochondria glutathione (GSH) and superoxide were measured and compared among the three groups of mice. Dismutase (SOD), total calcium and ATP content, and cardiac CD31, 琢-SMA and Collagen I mRNA expression. During the study period, 3 mice in the model group and 1 mouse in the mulberry leaf flavonoid group died. After 6 weeks of treatment with mulberry leaf flavonoids, the blood sugar of diabetic mice was significantly reduced, the blood insulin level, liver glycogen, liver HK and liver PK content, and myocardial mitochondrial GSH, SOD, total calcium and ATP content were significantly increased (P<0.05). Myocardial fibrosis indicators: the expression of CD31 mRNA in the myocardium of diabetic mice was significantly increased after treatment with mulberry leaf flavonoids, but the expression of 琢-SMA and Collagen I mRNA was significantly decreased (P<0.05). Mulberry leaf flavonoids can significantly reduce blood sugar levels in diabetic mice, improve their glucose metabolism and myocardial mitochondrial damage, and delay the progression of myocardial fibrosis. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Food Safety & Quality is the property of Journal of Food Safety & Quality Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Introduction: This study aimed to investigate the regulatory effects of mulberry leaf flavonoids and carnosic acid complex (MCC) on the growth performance, intestinal morphology, antioxidant, and p38 MAPK/Nrf2 pathway in broilers., Methods: A total of 256 healthy 8-day-old female yellow-feathered broilers were randomly divided into 4 equal groups: a control group (CON) fed a basal diet, an antibiotic group (CTC) supplemented with 50 mg/kg chlortetracycline, and two experimental groups (MCC75, MCC150) fed basal diets with 75 mg/kg and 150 mg/kg of MCC, respectively. The experiment lasted for 56 days, with days 1-28 designated as the initial phase and days 29-56 as the growth phase., Results: The results on the growth performance showed that diets supplemented with MCC and CTC decreased the feed-to-gain ratio (F/G), diarrhea rate, and death rate, while significantly increasing the average daily weight gain (ADG) ( p  < 0.05). Specifically, the MCC150 group enhanced intestinal health, indicated by reduced crypt depth and increased villus height-to-crypt depth ratio (V/C) as well as amylase activity in the jejunum. Both the MCC and CTC groups exhibited increased villus height and V/C ratio in the ileal ( p  < 0.05). Additionally, all treated groups showed elevated serum total antioxidant capacity (T-AOC), and significant increases in catalase (CAT) and glutathione peroxidase (GSH-Px) activities were observed in both the MCC150 and CTC groups. Molecular analysis revealed an upregulation of the jejunal mRNA expression levels of PGC-1α, Nrf2, and Keap1 in the MCC and CTC groups, as well as an upregulation of ileum mRNA expression levels of P38, PGC-1α, Nrf2, and Keap1 in the MCC150 group, suggesting activation of the p38-MAPK/Nrf2 pathway., Discussion: These findings indicate that dietary supplementation with MCC, particularly at a dosage of 150 mg/kg, may serve as a viable antibiotic alternative, enhancing growth performance, intestinal health, and antioxidant capacity in broilers by regulating the p38-MAPK/Nrf2 pathway., Competing Interests: YZ was employed by Geneham Pharmaceutical Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Liu, Huang, Chen, Zhou, Meng, Tan, He, Fu and Xiao.)

Introduction: Horses are susceptible to oxidative stress during strenuous endurance exercise, leading to muscle fatigue and damage. Mulberry leaf flavonoids (MLFs) possess significant antioxidant properties. However, the antioxidant efficacy of MLFs can be influenced by the extraction process, and their impact on H 2 O 2 -induced oxidative stress in equine skeletal muscle satellite cells (ESMCs) remains unexplored. Methods: Our study employed three extraction methods to obtain MLFs: ultrasound-assisted extraction (CEP), purification with AB-8 macroporous resin (RP), and n-butanol extraction (NB-EP). We assessed the protective effects of these MLFs on H 2 O 2 -induced oxidative stress in ESMCs and analyzed the MLF components using metabolomics. Results: The results revealed that pre-treatment with MLFs dose-dependently protected ESMCs against H 2 O 2 -induced oxidative stress. The most effective concentrations were 0.8 mg/mL of CEP, 0.6 mg/mL of RP, and 0.6 mg/mL of NB-EP, significantly enhancing EMSC viability ( p < 0.05). These optimized MLF concentrations promoted the GSH-Px, SOD and T-AOC activities ( p < 0.05), while reducing MDA production ( p < 0.05) in H 2 O 2 -induced ESMCs. Furthermore, these MLFs enhanced the gene expression, including Nrf2 and its downstream regulatory genes ( TrxR1 , GPX1 , GPX3 , SOD1 , and SOD2 ) ( p < 0.05). In terms of mitochondrial function, ESMCs pre-treated with MLFs exhibited higher basal respiration, spare respiratory capacity, maximal respiration, ATP-linked respiration compared to H 2 O 2 -induced ESMCs ( p < 0.05). Additionally, MLFs enhanced cellular basal glycolysis, glycolytic reserve, and maximal glycolytic capacity ( p < 0.05). Metabolomics analysis results revealed significant differences in mulberrin, kaempferol 3-O-glucoside [X-Mal], neohesperidin, dihydrokaempferol, and isobavachalcone among the three extraction processes ( p < 0.05). Discussion: Our study revealed that MLFs enhance antioxidant enzyme activity, alleviate oxidative damage in ESMCs through the activation of the Nrf2 pathway, and improve mitochondrial respiration and cell energy metabolism. Additionally, we identified five potential antioxidant flavonoid compounds, suggesting their potential incorporation into the equine diet as a strategy to alleviate exercise-induced oxidative stress., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zhang, Geng, Cao and Dugarjaviin.)

This study was aimed to evaluate the effect of mulberry leaf flavonoids (MLF) on oxidative stress, metabolic hormones, and milk production in Murrah buffaloes. Forty multiparous Murrah buffaloes (4 ± 1 lactations) with similar body weight (average 600 ± 50 Kg) and stage of lactation (90 ± 20 d) were randomly selected for this trial. Four treatment groups (10 buffaloes per group) with different doses of MLF included; control (0 g/d), MLF15 (15 g/d), MLF30 (30 g/d), and MLF45 (45 g/d). Buffaloes were fed with total mix ration consisting of grass (Pennisetum purpureum schum), brewery's grain and concentrate mixture for 5 weeks. Meteorological data including ambient temperature and relative humidity were recorded using the online dust monitoring system to calculate temperature-humidity index (THI). After 1 week of the adaptation, milk yield was recorded daily while physiological parameters (respiratory rate, rectal, and body surface temperature), and milk composition were measured weekly. At the end of the trial, blood samples were collected to analyze serum metabolic hormones including estradiol (E2), growth hormone (GH), prolactin (PRL), Tri-iodothyronine (T3), and Thyroxine (T4). Moreover, serum heat shock proteins (HSP), antioxidants enzymes including malondialdehyde (MDA), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) and blood biochemical indices were also analyzed. Results revealed a decrease (P = 0.012) in serum MDA level while increasing (P < 0.01) the HSP and serum GHS-Px contents in supplemented buffaloes. Treatment showed a linear and quadratic decrease (p = 0.001) in the serum T-AOC while reducing CAT contents linearly (p = 0.012) as compared to the control. However, no effect of treatment on serum SOD content was observed. Treatment resulted a linear increase (p = 0.001) in serum GH and PRL hormones while increasing serum E2 levels linearly (P < 0.001) and quadratically (P = 0.025). Treatment increased (p = 0.038) the daily milk yield as compared to the control. However, increase (P < 0.05) in serum T3 and T4 contents, fat corrected milk (4%) and milk protein (%) was observed only in MLF45. Moreover, we observed no change in serum biochemical indices except insulin which linearly increased (p = 0.002) in MLF45. Our findings indicated that MLF at 45 g per day is an appropriate level to enhance milk performance and alleviate heat stress in buffaloes.

As a new type of methane control agent, natural plant extract has been widely studied in recent years, but in vivo studies are few. This study was to investigate the effects of the dietary supplementation of 2 different polyphenols on the methane (CH4) emission and digestion metabolism in sheep. Ten healthy crossbred sheep (Dorper ♂ × small-tailed Han ♀; BW 60.0 ± 1.73 kg) were used in a change-over design. The sheep were fed the following 3 diets in the present study: the basal diet (CON) with no supplementation; the basal diet supplemented with 2 g mulberry leaf flavonoid (MLF) per day per sheep; the basal diet supplemented with 0.25 g resveratrol (RES) per day per sheep. Both MLF and RES reduced CH4 emission scaled to metabolic weight per kilogram of DMI and CO2 output scaled to metabolic weight, but the effect of RES was significant (P 0.05). In conclusion, MLF and RES can improve the digestibility of nutrients, the utilization of nutrients and energy, and reduce CH4 emission, but they are not conducive to nitrogen retention.

Mulberry (Morus alba L.) leaves have been widely applied to controlling blood glucose as a efficacious traditional Chinese medicine or salutary medical supplement. The extracts of mulberry leaf suppress inflammatory mediators and oxidative stress, protect the pancreatic β-cells and modulate glucose metabolism in diabetic rats. Our previous studies and others have shown that mulberry leaf extract has excellent therapeutic effects on type 2 diabetes mellitus (T2DM), however, the underlying mechanism remains to be studied. Skeletal muscle insulin resistance (IR) plays an important role in the pathogenesis of T2DM. The aim of this study was to investigate the effects and mechanisms of Mulberry leaf flavonoids (MLF) in L6 skeletal muscle cells and db/db mice. L6 skeletal muscle cells were cultured and treated with/without MLF for in vitro studies. For in vivo studies, the db/db mice with/without MLF therapy were used. Coomassie brilliant blue staining and α-SMA immunofluorescence staining were used to identify the differentiated L6 cells. Glucose level and ATP level of L6 myotubes were performed by optical density detection and cell viability was performed by MTT method. Mitochondrial membrane potential of L6 myotubes was detected by JC-1 fluorescent staining. ROS level of L6 myotubes was detected by DCFH-DA fluorescent staining. The body weight, food intake, and blood glucose of the mice were measured in different treatment days. Oral glucose tolerance test (OGTT), starch glucose tolerance test (STT) and insulin tolerance test (ITT) were performed in mice. Glycated hemoglobin, glycated serum protein, insulin, liver and muscle glycogen, total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-c) and low-density lipoprotein cholesterol (LDL-c) of the mice were detected by corresponding kit. The pathologic change of pancreas and skeletal muscle of mice were performed by H & E staining. Immunohistochemistry staining was used to detect the GLUT4 and p-AMPK expressions in skeletal muscle in mice. GLUT4, CPT-1, NRF1, COXIV, PGC-1α, and p-AMPK expression levels in L6 cells and mice were detected by western bolt assay. MLF and metformin significantly ameliorated muscle glucose uptake and mitochondrial function in L6 muscle cells. MLF also increased the phosphorylation of AMPK and the expression of PGC-1α, and up-regulated the protein levels of m-GLUT4 and T-GLUT4. These effects were reversed by the AMPK inhibitor compound C. In db/db mice, MLF improve diabetes symptoms and insulin resistance. Moreover, MLF elevated the levels of p-AMPK and PGC-1α, raised m-GLUT4 and T-GLUT4 protein expression, and ameliorated mitochondrial function in skeletal muscle of db/db mice. MLF significantly improved skeletal muscle insulin resistance and mitochondrial function in db/db mice and L6 myocytes through AMPK-PGC-1α signaling pathway, and our findings support the therapeutic effects of MLF on type 2 diabetes. Image 1 [ABSTRACT FROM AUTHOR]

This study was aimed to evaluate the effect of mulberry leaf flavonoids (MLF) on oxidative stress, metabolic hormones, and milk production in Murrah buffaloes. Forty multiparous Murrah buffaloes (4 ± 1 lactations) with similar body weight (average 600 ± 50 Kg) and stage of lactation (90 ± 20 d) were randomly selected for this trial. Four treatment groups (10 buffaloes per group) with different doses of MLF included; control (0 g/d), MLF15 (15 g/d), MLF30 (30 g/d), and MLF45 (45 g/d). Buffaloes were fed with total mix ration consisting of grass ( Pennisetum purpureum schum ), brewery's grain and concentrate mixture for 5 weeks. Meteorological data including ambient temperature and relative humidity were recorded using the online dust monitoring system to calculate temperature-humidity index (THI). After 1 week of the adaptation, milk yield was recorded daily while physiological parameters (respiratory rate, rectal, and body surface temperature), and milk composition were measured weekly. At the end of the trial, blood samples were collected to analyze serum metabolic hormones including estradiol (E2), growth hormone (GH), prolactin (PRL), Tri-iodothyronine (T3), and Thyroxine (T4). Moreover, serum heat shock proteins (HSP), antioxidants enzymes including malondialdehyde (MDA), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) and blood biochemical indices were also analyzed. Results revealed a decrease ( P = 0.012) in serum MDA level while increasing ( P < 0.01) the HSP and serum GHS-Px contents in supplemented buffaloes. Treatment showed a linear and quadratic decrease ( p = 0.001) in the serum T-AOC while reducing CAT contents linearly ( p = 0.012) as compared to the control. However, no effect of treatment on serum SOD content was observed. Treatment resulted a linear increase ( p = 0.001) in serum GH and PRL hormones while increasing serum E2 levels linearly ( P < 0.001) and quadratically ( P = 0.025). Treatment increased ( p = 0.038) the daily milk yield as compared to the control. However, increase ( P < 0.05) in serum T3 and T4 contents, fat corrected milk (4%) and milk protein (%) was observed only in MLF45. Moreover, we observed no change in serum biochemical indices except insulin which linearly increased ( p = 0.002) in MLF45. Our findings indicated that MLF at 45 g per day is an appropriate level to enhance milk performance and alleviate heat stress in buffaloes., (Copyright © 2020 Li, Hassan, Tang, Peng, Liang, Li, Peng, Xie and Yang.)

As a new type of methane control agent, natural plant extract has been widely studied in recent years, but in vivo studies are few. This study was to investigate the effects of the dietary supplementation of 2 different polyphenols on the methane (CH 4 ) emission and digestion metabolism in sheep. Ten healthy crossbred sheep (Dorper ♂ × small-tailed Han ♀; BW 60.0 ± 1.73 kg) were used in a change-over design. The sheep were fed the following 3 diets in the present study: the basal diet (CON) with no supplementation; the basal diet supplemented with 2 g mulberry leaf flavonoid (MLF) per day per sheep; the basal diet supplemented with 0.25 g resveratrol (RES) per day per sheep. Both MLF and RES reduced CH 4 emission scaled to metabolic weight per kilogram of DMI and CO 2 output scaled to metabolic weight, but the effect of RES was significant ( P  < 0.05). Both MLF and RES significantly improved apparent digestibility of DM, OM, NDF, ADF, and nitrogen, but the effect of RES was significant ( P  < 0.05). Both MLF and RES significantly improved ME ( P  < 0.05) and reduced energy losses in CH 4 emission ( P  > 0.05). In conclusion, MLF and RES can improve the digestibility of nutrients, the utilization of nutrients and energy, and reduce CH 4 emission, but they are not conducive to nitrogen retention.