Medicinal properties of Morus alba for the control of type 2 diabetes mellitus: a systematic review

Background: The objective of this review was to evaluate the medicinal potential of Morus alba leaves on the control of type 2 diabetes mellitus (DM2). The research question was: what is the therapeutic potential of Morus alba leaves for the control of DM2? Methods: This systematic review was based on the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) guidelines. The included studies were extracted from Scopus, Pubmed, ScienceDirect, Scielo, and Google Scholar; January 2015 to July 2021. Key search terms were MeSH and DeCS: Morus alba, mulberry, hypoglycemic agent. The inclusion criteria were: studies in rats administered Morus alba leaf extracts; studies that included the dimensions of lipidemia and glycemia; studies that included indicators such as fasting glucose, postprandial glucose, glycosylated hemoglobin, triglycerides, low-density lipoproteins, total cholesterol, and insulin resistance. Exclusion criteria: studies in which Morus alba leaves were administered with other plants; studies with other parts of the Morus alba plant; proteomic studies, cancer, duplicate studies, in vitro studies, and evaluation of included studies. All included investigations were evaluated for biases. Results: Of 253 studies found, 29 were included. The extracts of Morus alba leaves at the phytochemical level improve glucose uptake. Chlorogenic acid, isoquercitrin, and quercitrin, present in the leaves of Morus alba, have hypoglycemic properties and an ameliorating effect on diabetic nephropathy. This leaf has pharmacological effects such as glucose absorption, insulin secretion production, antioxidant and anti-inflammatory agent, antihyperglycemic and antihyperlipidemic activities, and obesity management. Conclusions: Morus alba leaves have pharmacological effects on DM2 that include glucose absorption, production of insulin secretion, antioxidant agent, antihyperglycemic and antihyperlipidemic activities, and obesity control. Beyond these results, there is a lack of studies on the potential and synergistic effects of Morus alba leaves' components, limiting the possibility of a more effective therapy using the plant's leaves.


Introduction
The World Health Organization (WHO) in 2014 globally reported that 422 million adults had diabetes. 1 Diabetes mellitus (DM) has become a major global problem with a significant health impact because the number of cases is constantly increasing and also because the long-term projections are less and less optimistic, 2 becoming one of the diseases selected by the world health authorities that require priority attention. In 2019, DM was the leading cause of 1.5 million deaths, and in 2012 2.2 million people died due to hyperglycemia. 2 By 2030, it is estimated that the global prevalence of diabetes will be 10.2% (578 million people) and 10.9% (700 million) by 2045, 3 Of the three types of diabetes mellitus, the most common is type 2 (T2DM).
T2DM is frequently linked to obesity. A predominant insulin resistance, accompanied by deficient hormone activity, leads to an increase in its secretion. 4 T2DM is a disease associated with multiple factors and of a chronic nature, so its treatment must be seen integrally in the patient's life. Therefore, continuous adjustments are needed according to the specific requirements of each patient. 5 An approach to diabetes care has been proposed by Kenneth S. Polonsky 6 and is based on the formation of work teams that involve health professionals for patient care and on the development of care delivery models for diseases using this approach, the Diabetes Prevention Program (USA) 6 demonstrated that physical exercise and weight loss could reduce the risk of diabetes in predisposed people by 58%. The main effects are also seen after treatment with metformin or pioglitazone.
The WHO, in its latest report on diabetes, points out that there are a variety of antidiabetic pharmacological therapies. However, most of them are expensive and cause adverse effects, impacting health systems, especially diabetic patients. The other problem is that only a minority in developing countries generally have insulin and drugs considered essential in controlling diabetes, such as antihypertensives and lipid-lowering drugs, within reach. Hence, it is necessary to establish regulatory policies and programs to improve equity in access. 1 The National Demographic and Family Health Survey (ENDES) carried out in 2013 in Peru indicated a progressive increase in the number of T2DM cases, mainly due to the lifestyle of the Peruvian population. Epidemiological data suggest that there is a prevalence of 33.8% of overweight and 18.3% of obesity. 7 In Peru, diabetes affects 7% of the population, representing 31.5% of acute myocardial infarctions and another 25% of cerebrovascular accidents. T2DM is equivalent to 96.8% of outpatient visits with this condition. The prevalence of glucose intolerance is 8.11%, and the variation in fasting glucose is 22.4%. The prevalence data for overweight, MS in adults, and obesity is 34.7%, 25%, and 17.5%, respectively. 8 The present review focuses its study on the leaves of the Morus alba plant, which is distinguished by being used in traditional medicine and which is also a very promising species. During the last 20 years, the great majority of studies have addressed the isolation, and active principles of the polysaccharides of this plant have focused mainly on its leaves and fruits, which are both medicinal and edible. 9 The results of preclinical investigations of the last decade suggest that Morus alba, as a therapeutic agent, is of utmost importance for treating DM; the polyhydroxy alkaloids, flavonoids, and polysaccharides of Morus may be; active components of great potentiality. 10 In addition, this product, especially in oriental medicine, has been used for fever, liver damage, vision problems, and to lower blood sugar levels. However, the studies carried out to date on the antidiabetic compounds in this product are insufficient. 11 Different parts of Morus alba are used in the treatment of various diseases, are required for their medicinal properties such as antioxidant, antibacterial, antidiabetic, and antiviral. In addition, it has active components in the leaf, such as polysaccharides, that can lower the glucose level, blood, and blood pressure. The leaves are used for hypoglycemia and also as antioxidants, which contain the chemical components: scopolin, isoquercitrin, astragalin, rose oxide, benzyl glucopyranoside, and phenolic acids such as gallic, protocatechuic, p-hydroxybenzoic, vanillic, chlorogenic, syringic, p-coumaric, ferulic, and m-coumaric, also the polysaccharide deoxynojirimycin (DNJ). 12 The present systematic review focuses on the active principles found in the leaves of the Morus alba plant and its medicinal properties, taking into account its use in traditional medicine. It is based on the fact that the extracts of Morus alba leaves contain flavonoids as main constituents, which also include antioxidant, antidiabetic, antihyperlipidemic, antiobesity, cardioprotective activities, and are rich in anthocyanins and alkaloids. 13 The purpose of this systematic review was to evaluate the medicinal potential of Morus alba leaves on the control of T2DM. The research question was: what is the medicinal potential of Morus alba leaves in the control of T2DM?

Methods
This review is based on the methodology established in the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) statement. (14,41) All the scientific information included in the study was extracted from international databases such as Scopus, Pubmed, ScienceDirect, Scielo, and Google Scholar from January 2015 -July 2021; for identification, filtering, eligibility, and inclusion, the Mendeley Desktop 1.19.8 web application was used. The protocol of this systematic review was registered in the PROSPERO database, ID: 274164, on 18 th August, 2021 (pending).
The primary auxiliary strategy (applied to all declared databases) to identify studies that correspond to the research question was the following: The auxiliary search strategy (applied to all declared databases) to identify other studies that correspond to the research question was the following: The inclusion criteria were: studies in rats administered Morus alba leaf extracts, studies that included dyslipidemia and glycemic dimensions, studies that included indicators such as fasting glucose, postprandial glucose, glycated hemoglobin, triglycerides, low-density lipoproteins, total cholesterol, and insulin resistance. Only empirical articles, systematic reviews, case studies were taken into account; Due to its importance, a descriptive analysis was included. The exclusion criteria were: studies in which Morus alba leaves were administered with other plants, studies with other parts of the Morus alba plant (fruits, roots, stems), proteomic analyses, cancer, duplicate studies, inward studies, in vitro and evaluation of included studies. Letters to the editor, narrative reviews, or scientific journal editorials were excluded from this study.
To correct biases, the PRISMA 2020 checklist 14 was first used. The empirical articles were evaluated through an analytical rubric prepared according to the parameters declared in the SSAHS scale of López-López E, Tobón S, Juárez-Hernández LG) to consider scientific articles. 15 Systematic reviews were assessed using the Quality Assessment of Systematic Reviews and Meta-Analyses, using an observation guide (checklist style according to the parameters of National Heart, Lung, and Blood Institute). 16 The checklists can be found as extended data. 41 Two experienced researchers on the team reviewed the systematic reviews-the original investigations by the three remaining members. The coordination and development of these activities were through the Zoom video chat software.
To include the studies, the relationship of each one with the research question was verified, based -fundamentallyon the terms: Morus alba; mulberry; and hypoglycemic agent. Then, Quality Assessment of Systematic Reviews and Meta-Analyzes and a scale for evaluating scientific articles were used to guarantee strict compliance with the inclusion and exclusion criteria stated in previous paragraphs. The web application used throughout the identification, screening, eligibility, and inclusion process was Mendeley. To collect the relevant data from each report, PRISMA was used. In the final process of compiling/coding the reports already examined, the entire research team was involved. The variables for which relevant information was sought were diabetes mellitus, type 2 diabetes mellitus, blood glucose, lipid-lowering agent, lipoproteins, blood lipids, glycemic control, postprandial hyperglycemia. Figure 1 shows that of an initial sample of 261, 29 investigations (17 originals and 12 systematic reviews) were included for 11.11%. The articles rejected for not meeting the inclusion criteria were 232, mainly because they lacked theoretical-methodological support, detected through the quality assessment instruments. Table 1 shows the distribution of the search results by author, article title, and type of study/characteristics of the studies finally included. 58,62% (original research); 41,38% (systematic reviews). All investigations were included based on the inclusion and exclusion criteria provided in the study methodology.

Results
Of the set of excluded studies, the proposal by Hwang SH, Li HM, Lim SS, Wang Z, Hong JS, Huang B, 17 despite meeting most of the inclusion criteria, was a clinical study that did not specify clearly if the extract was obtained from Morus alba leaves. Also, the research of Qi S, Li N, Tuo Z, Li J, Xing S, Li B, et al., 18 despite meeting most of the inclusion criteria, was excluded because the extract used did not specify if it had been made with Morus alba leaves. Finally, the research of Todd SB, Fallah S, Salemi Z, Seifi M. 19 was excluded because it measured the work, measured the level of specific hormones, but not that of the constituents of the Morus alba leaf that participate in the obesity.

Discussion
The genus Morus includes several species, such as Morus alba, M. nigra, both native to Asia, and M. rubra of North American origin. With wide distribution in the world, this genus, both in temperate and tropical areas, reports various medicinal properties, one of the most studied being Morus alba.
A study based on the network approach or network pharmacology found that the alkaloids, flavonoids, and polysaccharides of Morus alba leaves impart regulatory effects on blood sugar levels, identifying 202 components, of which 22 can have significant curative effects on DM. 20 To study its main bioactive components such as anthocyanins, polysaccharides, phenols, alkaloids, and flavonoids, at the phytochemical level, various techniques have been used that include extraction and separation processes such as solid-liquid extraction, solid-phase extraction, pressurized liquid  extraction, extraction by supercritical fluid, microwave-assisted extraction, ultrasonic-assisted extraction, enzyme assisted extraction, macroporous resin adsorption, silica gel chromatography, ion-exchange chromatography gel filtration chromatography, preparative liquid chromatography, and countercurrent chromatography. 21 Triterpenes and steroids are found in Morus alba leaf, fruit, and root extracts, as well as phenols and tannins and secondary metabolites with pharmacological properties for the treatment of many diseases associated with oxidative stress, such as Diabetes. 22 The leaves of Morus alba contain flavonoids as main constituents, which, in addition, possess various biological activities, including antioxidant, antidiabetic, antihyperlipidemic, antiobesity, cardioprotective activities, and are rich in anthocyanins and alkaloids. 13 Rutin or quercetin-3-rutinoside is a flavonoid glycoside obtained from the leaves of Morus alba, and its antiobesity effect has been verified by studying the lipid accumulation mechanism in 3T3-L1 adipocyte and C57BL mouse models. 23 Both compounds, rutin, and quercetin-3-O-β-D-glucoside (Q3G), have been shown to enhance glucose uptake through the AKT-mediated insulin signaling pathway or AMP-activated protein kinase activation (AMPK) in 3T3L1 adipocytes and could potentially be used in therapy for T2DM management. 11 In addition to rutin, the leaves of Morus alba contain apigenin, quercetin-3-triglyceride, lupeol, β-sitosterol, moracetin, isoquercitrin, coumarin, volatile oil, alkaloids, amino acids, and organic acids, proving the hypoglycemic properties and their effect enhancer on diabetic nephropathy. 24 Four main chemical components were identified in the ethanol extract of Morus alba: chlorogenic acid, rutin, isoquercitrin, and quercitrin. MLE was found to improve hyperglycemia, insulin resistance, and dyslipidemia in T2 DM rats with a significant therapeutic effect. 25 1-deoxynojirimycin (DNJ) is another compound present in the leaves of Morus alba. It is an effective inhibitor of α-glucosidase, providing a significant hypoglycemic effect. Its primary mechanism of action occurs after 1deoxynojirimycin enters the human body, with the inhibition of sucrose, maltase, α-glucosidase enzyme, and α-amylase decomposing starch, so it can block the absorption of carbohydrates in the body, inhibiting the increase in blood sugar to achieve the effect of the prevention and treatment of DM. 26 DNJ controls postprandial hyperglycemia by acting as a natural inhibitor of α-glucosidase in patients with T2DM. 27 Experimental models and laboratory techniques that evaluate various pharmacological activities have been included in this review. Several investigators have used streptozotocin-induced Wistar rats for the empirical part, administering Morus alba leaf extract. One of them demonstrated antioxidant activity, and antidiabetic effect of ethanol extracts with high levels of phenolic acids, chlorogenic acid, and flavonol glycosides, finding efficacy in the correction of hyperglycemia in the increased secretion of insulin. Two other investigations corroborated the improvement of the antioxidant situation 28 and that administering high doses (0.25%) and low doses (0.5%) of Morus alba leaf tea demonstrated hypolipidemic, hypoglycemic, and improvement on diabetic nephropathy. 24,29 Another study mentions that administering Morus alba ethanol extracts in doses of 0.5-1g/kg and oxyresveratrol (0.6 g/kg), an essential compound in the extract, could be considered as a potential adjunct therapy for the management of Diabetes since both reduced the level of glucose from fasting blood and plasma glucose. 30 Likewise, in another study, ethanolic extract of Morus alba leaves (100 mg/kg) was orally administered for eight weeks to female Wistar rats that were fed a high-cholesterol diet to evaluate the lowering effect of obesity, dyslipidemia, and insulin resistance, showing significant results since it decreased body weight gain, hypercholesterolemia, hypertriglyceridemia, atherogenic indices, based on lipid profile, and coronary artery, glucose level, and the insulin resistance index. 31 The effect of Morus alba leaf extract (FME) on insulin resistance through IRS-1 / PI3K / Glut-4 signaling pathway in rats with T2DM was also evaluated; the results obtained showed that the FME showed improvement in insulin resistance, in addition, a decrease in body weight, blood glucose, triglycerides, total cholesterol, and low-density lipoprotein levels were observed, significant antihyperglycemic and antihyperlipidemic activities via the pathway signaling IRS-1/PI3K/Glut-4, 33 Khyade (2018) obtained positive pharmacological effects on specimens of Rattus norvegicus induced with Diabetes, using a decoction of Morus alba leaves in doses of 20 g/L per day as treatment. The results found were decreased blood glucose levels and altered regulation of metabolic processes. 32 The effects of dietary supplementation with Morus alba leaf extracts with acetone-water (AE) and ethanol-water (EE) were also evaluated on the situation of the trace element (Fe, Zn, and Cu) about the management of Diabetes and antioxidant indices in diabetic rats fed the high-fat STZ diet and biochemical analyzes (glucose, thiobarbituric acid reactive substances, ferric reducing ability of plasma) were performed in blood serum, which showed that EA decreased the hepatic and renal storage of EF, while the EE increased the levels. 33 Laboratory tests were performed using mice induced to Diabetes and treatment with flavonoids extracted from the leaves of Morus alba on skeletal muscle cells, resulting in an improvement in mitochondrial function by activating AMPK in type 2 diabetes, improving resistance to the Skeletal muscle insulin significantly lowering blood glucose levels. 34 Likewise, it was investigated whether flavonoids, which are extracted from the leaves of Morus alba, can regulate glycolipid metabolism and to investigate whether flavonoids could regulate the signal of the IRS1 / PI3K / AKT pathway to affect the expression of FAS and GLUT4 membrane transfer capacity in 3T3. The results showed that the flavonoids from Morus alba leaf extracts alleviated the metabolic abnormalities of glycolipids in the IR model of 3T3-L1 adipocytes.
The effect was associated with the activation of the IRS1/PI3K/AKT pathway. 31 A double-blind, randomized clinical trial was conducted in animals and in patients with glucose intolerance, who were administered a standardized extract to evaluate the antihyperglycemic effect, the inhibitory effect of α-glucosidase, the acute single oral toxicity, and the reduction of blood glucose in a test. The standardized extract of Morus alba leaves was found to inhibit α-glucosidase at a level four times higher than the positive control (acarbose), in a concentrationdependent manner, proving to be an excellent potential material to improve postprandial hyperglycemia. 35 Work has been carried out with the interaction of the leaves of Morus alba and other plants. One of them used fenugreek seed (Trigonella foenum-graecum), Morus alba Leaf, and American ginseng root (Panax quinquefolius) can improve glycemia in humans and animals with impaired glucose and T2DM metabolism, improving insulin sensitivity and glucose absorption in human adipocytes. 36  One method used to standardize the Morus alba extract is high-performance reversed-phase liquid chromatography (RP HPLC). The plant extract and the single bioactive molecule were studied for their potential for inhibition of combined CYP450, as well as forms of recombinant human CYP450, such as the isoenzymes CYP3A4, CYP2D6, CYP2C9, and CYP1A2, the findings suggested that the plant extract and its bioactive compound are safe to use in the management of Diabetes. 38 Likewise, experimental studies in diabetic rats and normal rats have shown that the fruit and bark of the Morus alba root can potentially be used as an effective treatment for DM2 39 and effectively improve hyperlipidemia and inhibit diacylglycerol acyltransferase. 18 A systematic review indicates that several studies have shown that the aqueous extracts of the bark of roots, leaves, and stems contain polyphenols and polysaccharides with antihyperglycemic and antihyperlipidemic properties; polysaccharides have been further studied to assess their antidiabetic bioactivity in T2DM-induced rats, clarifying the underlying mechanism of these activities. 9 A systematic review is presented on the chemical composition of the different parts of Morus alba and its pharmacological effects on DM, which include the influence on glucose absorption, production on insulin secretion as an antioxidant and anti-inflammatory agent. The study found that Morus alba leaf contains steroids and triterpene compounds, flavonoids, coumarin, essential oils, amino acids, alkaloids, and organic acids. The branch of Morus alba, for its part, contains tannin, fructose, stachyose, glucose, maltose, honeydew, and arabinose. The bark of the Morus alba root contains flavonoids, such as mulberry, mulberrochromene, and cyclomulberrine. The different parts of the plant contain active principles such as Polyhedral alkaloids, polysaccharides, and flavonoids are components that are found in all aspects of the Morus alba plant; the fruits also contain vitamin B1, B2, and carotene, also, fatty acids (linoleic, oleic and stearic). 10 Another review to evaluate the potential of Morus alba as a natural agent in the treatment of obesity found that the inhibitory effects of Morus alba on digestive enzymes and adipocyte differentiation and its stimulating effects on energy expenditure and Lipid metabolism are mechanisms responsible for the management of obesity in obese patients. 40

Conclusions
It has been shown that the aqueous extracts of the leaves of Morus alba at a phytochemical level contain 202 constituents, among them routine with proven antiobesity effect, rutin together with quercetin-3-O-β-D-glucoside (Q3G) improve the glucose uptake and demonstrated antioxidant activity and antidiabetic effect.
The compound in the Morus alba leaf, 1-deoxynojirimycin (DNJ), has been identified as an effective inhibitor of α-glucosidase and provides a significant hypoglycemic effect, which could improve postprandial hyperglycemia and also prevent and treat in DM, oxyresveratrol could be considered as a potential adjunct mechanism for the management of Diabetes.
Chlorogenic acid, routine, isoquercitrin, and quercitrin present in Morus alba leaves have been shown to have hypoglycemic properties and an ameliorating effect on diabetic nephropathy, insulin resistance, and dyslipidemia in rats.
Various systematic reviews have evaluated that Morus alba leaves have various pharmacological effects on T2DM, including glucose absorption, production in insulin secretion, antioxidant and anti-inflammatory agent, antihyperglycemic and antihyperlipidemic activities, and obesity management.
The results of this research have a theoretical and scientific relevance because it manages to systematize scientific research relevant to the cognitive needs that the need for the use of Morus alba leaf for the treatment of type 2 diabetes mellitus raises today, which, as explained in this research, is a public health problem worldwide.

Limitations
The results achieved allow identifying a lack of studies on the potential and synergistic effects of the components of the Morus alba leaves; this limits the possibility of a more effective therapy using the leaves of the plant; Furthermore, there are few studies on the extraction methodology of the components of the Morus alba leaf. These limitations must be taken into account for the orientation and development of new research.

Recommendations
T2DM and the risk factors involved in the appearance of aggravating conditions of this disease, it should be considered that the extracts of the leaves of Morus alba constitute an effective alternative to keep glycemia and lipidemia under control in patients with T2DM, being an excellent complement to your usual treatment. It would also be necessary to expand pharmacological safety studies and preclinical and clinical studies of the use of Morus alba leaf extracts to generate more scientific evidence, which allows new forms of administration to be known.

Data availability
Underling data All data underlying the results are available as part of the article and no additional source data are required.