Research progress on chemical constituents and pharmacological activities of Morus alba / 中国中药杂志

Morus alba, a traditional economic crop, is also a significant medicinal plant. The branches(Mori Ramulus), leaves(Mori Folium), roots and barks(Mori Cortex), and fruits(Mori Fructus) of M. alba are rich in chemical components, such as alkaloids, flavonoids, flavanols, anthocyanins, benzofurans, phenolic acids, and polysaccharides, and possess hypoglycemic, hypolipidemic, anti-inflammatory, anti-tumor, anti-microbial, liver protective, immunoregulatory, and other pharmacological activities. This study analyzed the sources, classification, and functions of the main chemical components in M. alba and systematically summarized the latest research results of essential active components in M. alba and their pharmacological effects to provide references for in-depth research and further development as well as utilization of active components in M. alba.

Mori Folium Improves Glucose and Lipid Metabolism Disorders in Rats with Type 2 Diabetes Mellitus by Regulating PI3K/Akt/PPARα/CPT-1 Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect and mechanism of Mori Folium extract on the glucose and lipid metabolism disorders in the liver of rats with type 2 diabetes mellitus （T2DM） through the phosphatidylinositol 3-kinase/protein kinase B/peroxisome proliferation-activated receptor α/carnitine palmitoyl transferase-1 （PI3K/Akt/PPARα/CPT-1） signaling pathway. MethodThe T2DM model was induced by the high-fat diet combined with the intraperitoneal injection of streptozotocin （STZ）. The model rats were randomly divided into a model group， a metformin （0.2 g·kg-1） group， and a Mori Folium water extract （4.0 g·kg-1） group according to blood glucose and body weight. In the 8-week administration， fasting blood glucose was measured at the same time every week. The histomorphological and fat changes in the rat liver were observed by hematoxylin-eosin （HE） staining and oil red O staining. The levels of total cholesterol （TC）， triglyceride （TG）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， alanine aminotransferase （ALT）， and aspartate aminotransferase （AST） in the serum were measured by biochemical methods. Western blot （WB） was used to quantitatively detect the protein expression of p-PI3K，PI3K，p-Akt，Akt，PPARα，and CPT-1 in the rat liver. ResultAfter 8-week administration， the blood glucose of rats was higher in the model group than that in the control group （P<0.01）， and lower in the Mori Folium water extract group than that in the model group （P<0.01）. The results of HE staining showed that the liver tissue structure of the control group was complete， and the hepatocytes were arranged radially around the central vein， while the hepatocyte injury in the model group was obvious. Compared with the model group， the Mori Folium water extract group showed improved vacuolar degeneration and no lesions such as small bile duct hyperplasia. Oil red O staining showed that there was no obvious steatosis and necrosis in the hepatocytes of rats in the control group， and no lipid droplets in the hepatocytes were observed， while the model group showed increased lipid droplets. Mori Folium significantly reduced the lipid droplets in the liver. Biochemical analysis showed that the levels of TC， TG， LDL-C， AST， and ALT in the model group were significantly higher than those in control group （P<0.01）. The levels of TC， TG， LDL-C， AST， and ALT in the Mori Folium water extract group were significantly lower than those in the model group （P<0.05，P<0.01）. WB showed that the protein expression of p-PI3K/PI3K， p-Akt/Akt， PPARα， and CPT-1 in the model group were lower than those in the control group （P<0.01）. Mori Folium water extract could increase the protein expression of p-PI3K/PI3K， p-Akt/Akt， PPARα， and CPT-1 （P<0.05 or P<0.01）. ConclusionThe hypoglycemic mechanism of Mori Folium water extract may be related to the regulation of the PI3K/Akt/PPARα/CPT-1 signaling pathway.

Herbal Textual Research on Mori in Famous Classical Formulas / 中国实验方剂学杂志

Through consulting the ancient herbs， medical books and modern literature， this paper made textual research on the name， origin， producing area， quality evaluation， collection and processing of medicinal materials of Sang （Mori Folium， Mori Cortex， Mori Ramulus， Mori Fructus） in famous classical formulas， in order to provide a basis for the development of famous classical formulas containing medicinal materials of Sang. According to the research， Mori Folium and Mori Cortex were first used as medicines in Shengnong Bencaojing ， Mori Ramulus was first used as medicine in Jinxiaofang， and Mori Fructus was first used as medicine in Xinxiu Bencao. Before the Tang dynasty， there were Nyusang and Shansang. Since Tang dynasty， there were many sources of medicinal materials of Sang， including Baisang （Morus alba）， Jisang （M. australis）， Shansang （M. mongolica）， etc. According to textual research， the mainstream varieties were M. australis， M. alba and their cultivated varieties. In modern times， according to the relevant information and the Chinese Pharmacopoeia， M. alba is the original base. In ancient times， the origin of mulberry changed with the development of sericulture， mulberry has been widely planted since the Song dynasty. In the Ming and Qing dynasties， mulberry has been planted most in Jiangsu and Zhejiang. In modern times， they are mainly produced in Jiangsu， Zhejiang， Anhui， Hunan and other places. In recent years， due to the related policies and strategies such as "moving silkworms from east to west"， the center of silkworm breeding has gradually transferred to the west. As for the quality evaluation and harvesting and processing of mulberry medicinal materials， Most of the ancient and modern records of Mori Folium are the same. They are harvested after frost， and dried after removing impurities. The quality is better when the leaves are large and thick， yellowish green， holding prickly hands and undergoing frost. The harvesting period of Mori Cortex is slightly different in ancient and modern records. Ancient books record that it can be harvested all the year round， but in modern times， it is mostly harvested from late autumn to the next spring. The processing methods include removing soil and fibrous roots， scraping off yellow-brown rough skin， peeling off white skin and drying in the sun. The quality is better when they are white， thick， flexible， free of rough skin and full of powder. There are few records about the collection， processing and quality evaluation of Mori Ramulus and Mori Fructus in ancient Chinese herbal books. According to modern literature， Mori Ramulus is usually collected in late spring and early summer， with leaves removed， slightly dried， sliced while fresh， and dried in the sun. The best quality of Mori Ramulus is fine and tender with the yellow and white section. Mori Fructus is harvested from April to June when the fruit turns red， and dried in the sun， or slightly steamed and dried in the sun， and it is better to be big， dark purple， oily and thick. There are many processing methods of mulberry medicinal materials. Ancient books record stir frying， baking， burning and steaming of Mori Folium， in modern times， there is honey-roasted method， but most of them are used as raw products. In ancient materia medica， Mori Cortex has firing method， baking method， stir-frying method， honey-fried method， etc. In modern times， there are stir-fried and honey-fried methods， and most of them are used as raw products. Ancient books record that Mori Ramulus has cutting and frying methods， while modern ones have cutting， frying， wine-processed and bran-processed methods. Processing methods of Mori Fructus are consistent in ancient and modern times， and they are mostly dried after being cleaned or steamed. Based on the research results， it is suggested that M. alba should be selected as mulberry medicinal materials in the famous classical formulas， and appropriate medicinal parts and processing methods can be selected according to the indications of the famous classical formulas.

Mechanism of effective components of Mori Folium in alleviating insulin resistance based on JNK signaling pathway / 中国中药杂志

A stable hepatoma cell line(Hep G2 cell) insulin resistance model was established and used to analyze the effect of effective components of Mori Folium in alleviating insulin resistance,and preliminary explore the mechanism for alleviating insulin resistance. The Hep G2 insulin action concentration and the duration of action were investigated using the glucose oxidase method(GOD-POD method) to establish a stable Hep G2 insulin resistance model. Normal control group,model group,Mori Folium polysaccharide group,Mori Folium flavonoid group and rosiglitazone group were divided to determine the glucose consumption. The effect of Mori Folium effective components on Hep G2 insulin resistance was analyzed. The mRNA expressions of JNK,IRS-1 and PDX-1 in each group were detected by Real-time quantitative PCR(qRT-PCR). The protein expressions of p-JNK,IRS-1 and PDX-1 were detected by Western blot. And the mechanism of effective components of Mori Folium in alleviating insulin resistance was investigated. The results showed that the glucose consumption was significantly decreased in the insulin resistance cells after incubation with 25. 0 mg·L-1 insulin for 36 h(P<0. 01),and the model was relatively stable within 36 h. Mori Folium polysaccharides and flavonoids all alleviated insulin resistance,among which Mori Folium flavonoids had better effect in alleviating Hep G2 insulin resistance(P<0. 05). The qRT-PCR analysis showed that Mori Folium polysaccharides and flavonoids could inhibit JNK and IRS-1 mRNA expressions,while enhancing PDX-1 mRNA expression. Western blot analysis displayed that Mori Folium polysaccharides and flavonoids could inhibit p-JNK and IRS-1 protein expressions,while enhancing PDX-1 protein expression. Mori Folium polysaccharides and flavonoids can alleviate insulin resistance in Hep G2 cells,and its mechanism may be the alleviation of insulin resistance by inhibiting JNK signaling pathway.

Study on Frosting Quality Markers of Mori Folium Based on Component Characteristic / 中国实验方剂学杂志

Objective: To screen the frosting qualitymarkers (Q-marker) of Mori Foliumand identify Mori Folium after frost. Method: The HPLC-DAD-MSn fingerprints of Mori Folium before and after frost were established,the common peaks were markedand the characteristic components were selected by orthogonal partial least squares discrimination analysis (OPLS-DA). The components with a large content change were used as marker components,and the relatively stable component was used as an internal reference component. The ratio of the marker components to the peak area of the internal reference component was used as component characteristic to identify the frost of Mori Folium,andqualitative study of frosting markerswere performed by MSn and authentic standards. Result: The fingerprints were established and 33 common peaks were marked. Through OPLS-DA,peaks 1,23,14 were determined as marker components,peak 12 was determined as internal reference component. The frosting quality markers of Mori Folium were characterized as citric acid derivatives,saponin F,tryptophan and neochlorogenic acid by MSn and standards. The ratios of the peak areas of citric acid derivatives,saponin F,tryptophan and neochlorogenic acid before and after frost were 0.15±0.054,1.0±0.48; 0.14±0.073,0.98±0.48,0.13±0.088,0.89±0.49,respectively. Conclusion: In the fingerprints established in this study,the peak area ratios of citric acid derivatives,saponin F,tryptophan to new chlorogenic acid had specificity,which can be used as frosting quality markers for identification of Mori Folium. This study expanded the connotation of the Q-marker specificity of traditional Chinese medicine. The results can provide experimental data for the quality evaluation and drug-forming properties of Mori Folium,and provide reference for similar research.

Effect and Mechanism of Mori Folium on Insulin Resistance in 3T3-L1 Cells / 中国实验方剂学杂志

Objective: To observe effect of Mori Folium-containing serum on glucose consumption and cell activity of fat cell line 3T3-L1 insulin resistance (IR) model, in order to screen out the optimal concentration of drug-containing serum, detect effect of Mori Folium on the content of inflammatory factors, and explore the possible mechanism. Method: 3T3-L1 preadipocytes in logarithmic growth phase were selected, and induced with 10 mg·L-1 insulin (Ins), 0.25 mmol·L-1 dexamethasone (DEX) and 0.5 mmol·L-1 3-isobutyl-methylxanthine(IBMX) for 48 h and then with 10 mg·L-1 Ins for 48 h. After the cells were differentiated into mature adipocytes, they were induced with 1 μmol·L-1 DEX for 96 h to establish IR model. Glucose content in the supernatant of cells was detected by glucose oxidase after serum containing Mori Folium cultured for 12,24,36, 72 h. Methyl-thiazdyl-tetrazolium(MTT) was used to detect the effect of serum containing Mori Folium on IR cells activity. The content of tumor necrosis factor-α(TNF-α) was determined by enzyme-linked immunosorbent assay (ELISA). Meanwhile, the effects of inflammatory factors on the expressions of insulin signaling pathway proteins insulin receptor (InsR), insulin receptor substrate (IRS), p-IRS1 and glucose transporter 4 (GLUT4) were determined by Western blot. Result: Serum containing Mori Folium could significantly increase the glucose consumption rate and cell activity of IR cells (Pα (PPPConclusion: Mori Folium can significantly improve IR status of 3T3-L1 cells, and its mechanism may be related to inhibiting TNF-α and promoting the expressions of insulin signaling pathway proteins.

Fast Identification Analysis on Chemical Constituents in Mori Folium by Using UPLC-Q-TOF/MS Method / 中国中医药信息杂志

Obejective To conduct fast identification analysis on chemical constituents of Mori Folium UPLC-Q-TOF/MS method.Methods ACQUITY UPLC-Q-TOF/MS spectrometer was used, acetonitrile and water (containing 0.1％ formic acid) as mobile phase with gradient elution. The flow rate was 0.3 mL/min, with ESI negative ion mode detection, data analysis with Masslynx4.1 software.Results According to the retention time provided by the mass spectrometry, the exact molecular weight and the secondary mass spectrometry were used to analyze the fragments by referring to literature, and 19 chemical constituents of Mori Folium were identified and deduced.Conclusion The method can analyze the chemical constituents of Mori Folium rapidly, sensitively and comprehensively, and provide the basis for the study of the pharmacological basis of Mori Folium.

Inhibitory effects on the production of inflammatory mediators and reactive oxygen species by Mori folium in lipopolysaccharide-stimulated macrophages and zebrafish

ABSTRACT Mori folium, the leaf of Morus alba L. (Moraceae), has been traditionally used for various medicinal purposes from ancient times to the present. In this study, we examined the effects of water extract of Mori folium (WEMF) on the production of inflammatory mediators, such as nitric oxide (NO) and prostaglandin E2 (PGE2), and reactive oxygen species (ROS) in lipopolysaccharide (LPS)-stimulated murine RAW 264.7 macrophages. Our data indicated that WEMF significantly suppressed the secretion of NO and PGE2 in RAW 264.7 macrophages without any significant cytotoxicity. The protective effects were accompanied by a marked reduction in their regulatory gene expression at the transcription level. WEMF attenuated LPS-induced intracellular ROS production in RAW 264.7 macrophages. It inhibited the nuclear translocation of the nuclear factor-kappa B p65 subunit and the activation of mitogen-activated protein kinases in LPS-treated RAW 264.7 macrophages. Furthermore, WEMF reduced LPS-induced NO production and ROS accumulation in zebrafish. Although more efforts are needed to fully understand the critical role of WEMF in the inhibition of inflammation, the findings of the present study may provide insights into the approaches for Mori folium as a potential therapeutic agent for inflammatory and antioxidant disorders.

Effects of Mori Folium extract on diet-induced obesity mechanism in rats / 中国中药杂志

To study the anti-obesity effect of Mori Folium extract on diet-induced obesity(DIO) and to explore the preliminary mechanism in rats. DIO rat models were established by high glucose and high fat diet for 8 weeks. Then high(10 mg•kg⁻¹) and low(5 mg•kg⁻¹) does Mori Folium extracts were given by intragastric administration for 13 weeks. After the last administration, their body weight, 24 h food intake, water intake, Lee's index, liver/body mass index, and fat/body mass index were determined. The levels of lipoprotein lipase(LPL), CCAAT/enhancer-binding protein alpha(C/EBPα) and peroxisome proliferator-activated receptor gamma(PPARγ) were measured by enzyme-linked immunosorbent assay(ELISA). Phosphorylated AMP-activated protein kinase alpha(p-AMPKα), C/EBPα and PPARγ expression levels in adipose tissues were detected by Western blot. The hematoxylin-eosin staining(HE) was used to observe the histopathological changes of adipose tissues. The results showed that both high dose and low dose Mori Folium extract can decrease body weight, Lee's index, renal fat/body mass ratio and testicle fat/body mass ratio, and the high dose group could decrease the total fat/body mass ratio. Both high dose and low dose groups had no significant effect on the food intake and water intake; however, they could decrease levels of LPL in fat, up-regulate p-AMPKα protein expression, down-regulate C/EBPα and PPARγ protein expression, and reduce fat cell volume. In conclusion, Mori Folium extract had a slimming effect on DIO rats, and its mechanism may be associated with up-regulating the expression of p-AMPKα, down-regulating the expression of PPARγ, C/EBPα and LPL, inhibiting the differentiation of preadipocytes into mature fat cells, and reducing the volume of fat cells.

Pharmacokinetics of Mori Folium flavones and alkaloids in normal and diabetic rats / 中国中药杂志

To study the pharmacokinetic effect of Mori Folium flavones and alkaloids in normal and diabetic rats. An UPLC-TQ-MS method was developed for the simultaneous determination of rutin, isoquercitrin, astragalin, kaempferol, quercetin, chlorogenic acid, cryptochlorogenic acid, neochlorogenic acid, DNJ and fagomine in plasma of rats. The diabetic rat model was induced through intravenous injection with alloxan and high-fat diet. Samples of plasma of rats were obtained at different time points, after the rats were administrated with Mori Folium flavones and alkaloids. After the deproteinization with acetonitrile, the concentrations of Mori Foliam constituents in rats at different time points were detected by UPLC-TQ-MS method, and pharmacokinetic parameters were calculated by DAS 2.0 software. The results showed that quercetin and kaempferol reached peak at 0.333 h, indicating that Mori Folium flavonoid constituents were absorbed and distributed quickly. At about 4 h after administration, both of them reached the peak concentrations for the second time, suggesting that they stayed in intestine for a long time. DNJ and fagomine in gastrointestinal tract can be quickly absorbed into blood, and the concentration in plasma reached peak after 0.667 h, suggesting that both of them could be rapidly distributed in the systemic circulation of rats. Cryptochlorogenic acid, neochlorogenic acid, quercetin, kaempferol and rutin were found to have a higher Cmax and AUC0-t in normal rats than those in diabetic rats. The t1/2values of cryptochlorogenic acid and neochlorogenic acid were shorter in diabetic rats, while quercetin, kaempferol and rutin had a longer t1/2value in diabetic rats. Chlorogenic acid, astragalin, isoquercitrin, fagomine had a higher Cmax in diabetic rats, and the t1/2values of astragalin and fagomine were longer, which suggested differences in absorption of active ingredients under normal and diabetic conditions.

Protective effects of total flavonoid from Mori folium on renalfibrosis in type 1 diabetic mice and specific mechanisms of these effects / 中国药理学通报

Aim To observe the effect of total flavonoid from Mori folium(TFMF) on renal interstitial fibrosis in type 1 diabetic mice and its possible mechanism.Methods Diabetic mice were induced by intraperitoneal injection of streptozotocin(STZ) dissolved in 0.01 mol·L-1 citrate buffer(pH 4.5) at 150 mg·kg-1 body weight after 12 h of food deprivation.Forty model mice were divided randomly into four groups: model group, and low-(0.25 g·kg-1), moderate-(0.5 g·kg-1), high-dose groups(1 g·kg-1) fed with TFMF once daily.In addition, eight normal mice were used as normal group.After 12 weeks, the fasting blood glucose(FBG), serum creatinine(Cr), blood urea nitrogen(BUN) and microalbuminuria(mAlb) were measured.Masson staining, Sirius red staining and collagen type Ⅳ immunohistochemical staining were used to detect the expression of collagen protein in the cortex, while laminin staining to assess the degree of glomerular and renal tubular basement membrane thickening.The protein expressions related to epithelial-mesenchymal transition and PI3K/Akt/mTOR in the renal cortex of mice were detected by Western blot.Results The moderate and high dose of TFMF could significantly decrease the levels of FBG, Cr, BUN and mAlb in diabetic mice, meanwhile decreasing the expression of α-SMA protein by inhibiting the activation of PI3K/Akt/mTOR signaling pathway, which led to the amelioration of the pathological alterations of renal tissue.Conclusions The moderate and high dose of TFMF can reduce the level of renal interstitial fibrosis in type 1 diabetic mice, and its mechanism may be related to the inhibition of activation of PI3K/Akt/mTOR signaling pathway.

The Immunomodulatory Activity of Mori folium, the Leaf of Morus alba L., in RAW 264.7 Macrophages In Vitro

BACKGROUND: Immunoregulatory elements have emerged as useful immunotherapeutic agents against cancer. In traditional medicine, Mori folium, the leaf of Morus alba L. (Moraceae), has been used for various medicinal purposes; however, the immunomodulatory effects have not been fully identified. We evaluated the immunoenhancing potential of water extract of Mori folium (WEMF) in murine RAW264.7 macrophages. METHODS: RAW264.7 cells were treated with WEMF for 24 hours and cell viability was detected by an MTT method. Nitric oxide (NO) levels in the culture supernatants were assayed using Griess reagent. The productions of prostaglandin E2 (PGE2) and immune-related cytokines was measured using ELISA detection kits. The mRNA and protein expression levels of Inducible NO synthase, COX-2, and cytokines were assayed by reverse transcription-PCR and Western blotting, respectively. The effect of WEMF on phagocytic activity was measured using a Phagocytosis Assay Kit. RESULTS: WEMF significantly stimulated the production of NO and PGE2 as immune response parameters at noncytotoxic concentrations, which was associated with the increased expression of inducible NO synthase and COX-2. The release and expression of cytokines, such as TNF-α, interleukin (IL)-1β, IL-6, and IL-10, were also significantly increased in response to treatment with WEMF. Moreover, WEMF promoted the macrophagic differentiation of RAW264.7 cells and the resulting phagocytosis activity. CONCLUSIONS: WEMF has the potential to modulate the immune function by regulating immunological parameters. Further studies are needed to identify the active compounds and to support the use of WEMF as an immune stimulant.

Research progress on bioactive component groups and their action mechanisms of Mori Folium for prevention and treatment of diabetes / 中草药

Mori Folium (mulberry leaves or the leaves of Morus alba) as one of Chinese materia medica and food raw materials, its research and application in the treatment of diabetes and diabetic complications have become an increasing concern to the majority of scientists. With deepening the basic research and industrial development of Mori Folium, the bioactive component groups and their action mechanisms of Mori Folium used for the prevention and treatment of diabetes and diabetic complication continue to be revealed. In this paper, based on the previous research, we summarize and analyze the understanding of domestic and foreign scholars about the pathogenesis of diabetes and the active component groups and their action mechanisms of Mori Folium for the prevention and treatment of diabetes and diabetic complication in recent years in order to provide the guidance for the further study and the reference for the development and utilization of Mori Folium resource.

Improvement of Assay Method for Mori Folium in Chinese Pharmacopoeia / 中国药师

To optimize the extract method and chromatographic conditions for the determination of rutin in Mori Folium in Chinese pharmacopoeia. Methods:The HPLC analysis was performed on a ZORBAX SB-C18 (250 mm × 4. 6 mm, 5 μm) column. The mobile phase was acetonitrile-0. 2% phosphorice acid solution with gradient elution and the flow rate was 1. 0 ml·min-1 . The de-tection wavelength was 354 nm, and the column temperature was 30 ℃. Results: The content of rutin determined by the method in Chinese pharmacopoeia was actually the total contents of rutin and isoquercitrin, while the optimized method could separate the two components effectively, and the good linearity of rutin and isoquercitrin was within the range of 2.76-27.60 μg·ml-1(r=0.999 9) and 4. 74-47. 39 μg·ml-1(r=0. 999 8), respectively, and the average recovery was 100. 31% (RSD=0. 83%) and 100. 32%, re-spectively (RSD=1. 04%, n=6). Conclusion:The optimized method is simple, stable and reproducible,which can be used in the quality control of Mori Folium.

Role of mulberry resources in prevention and treatment of type 2 diabetes / 中草药

Mulberry resources including Mori Folium, Mori Fructus, Mori Ramulus, and Mori Cortex are used in clinic of traditional Chinese medicine, and have important functional effects on type 2 diabetes. In order to better explore diabetic drugs and take better advantage of mulberry resources, we review the chemical constituents and pharmacological effects of mulberry including hypoglycemic, hypolipidemic, anti-inflammatory, and anti-oxidant activities.