How Does In Vitro Digestion Change the Amount of Phenolics in Morus alba L. Leaf? Analysis of Preparations and Infusions.

The application of Morus alba L. in traditional oriental medicine and cuisine has resulted in numerous studies on its health-promoting effects. However, if the process is not monitored by the manufacturers, the processing of the leaves alters the obtained health-promoting properties and results in different health qualities in the final composition of dietary supplements. This article aims to analyze changes (using the HPLC/DAD method) in the proposed conditioned mulberry leaves in terms of key compounds (phenolic acids and flavonols) responsible for antioxidant activity after being digested in in vitro conditions. The analyzed material was leaves of white mulberry (Morus alba L.) cv. Zólwinska wielkolistna, conditioned (1-4 h) and non-conditioned. The conditioning process of mulberry proposed here, e.g., for industry production, resulted in variable transformations of polyphenols during in vitro digestion. For many polyphenols, especially those shown in the highest amounts, significant correlations were found between their content and conditioning, as well as the stage of digestion. In the case of mulberry infusions, the amounts of individual polyphenols were several times lower than in the preparations, which was due to the degree of dilution. Their amounts tended to decrease in the course of digestion. Taking this into account, it seems justified to continue research on the in vivo bioavailability of bioactive components from conditioned Morus alba L. leaves.

Morus alba L. Leaves (WML) Modulate Sweet (TAS1R) and Bitter (TAS2R) Taste in the Studies on Human Receptors - A New Perspective on the Utilization of White Mulberry Leaves in Food Production?

From the nutritional perspective, the main direction of the utilization of white mulberry (Morus alba L.) parts so far has been to produce dietary supplements or functional foods for individuals with diabetes or over-weight. Its leaves are widely known as a valuable source of bioactive compounds responsible for its antioxidant and antidiabetic effects, both in animals and humans. The authors found that processed leaves can also be investigated as potential bitter and/or sweet taste modulators-an important property of new functional foods. The study aimed to validate the inhibitory effect of Morus alba L. on the TAS2R3 and TAS2R13 bitter taste and TSA1R2/TSA1R3 receptors and determine the changes that the conditioning process caused in such receptors. The effect on the receptors was evaluated in specially transfected HEK293T cells, and the inhibition ratio was measured using the calcium release test. Moreover, the stability of phenolics in the simulated intestinal in vitro digestion process was determined. Results showed that the Morus alba leaf extracts were rich in gallic, chlorogenic and caffeic acids together with rutin and quercetin 3-(6-malonyl)-glucoside, while the conditioning process positively affected their amount. Most identified phenolics were reduced during in vitro digestion. In the taste receptors test, it was found that the phytochemicals from conditioned Morus alba leaf extract enhanced sweet taste, together with a reduction of bitter taste receptor activity in some cases. To conclude, the study has found that Morus alba, especially when conditioned for 4 h, seems to be a valuable modulator of taste, which should be considered in future research as a crucial reason for its new utilization.

Antioxidant and Anti-Inflammatory Activity on LPS-Stimulated RAW 264.7 Macrophage Cells of White Mulberry (Morus alba L.) Leaf Extracts.

The white mulberry (Morus alba L.) is widely used as a medicinal plant in Asia. In this study, the bioactive compounds of ethanolic extracts of white mulberry leaves from the Sakon Nakhon and Buriram cultivars were evaluated. The ethanolic extracts of mulberry leaves from the Sakon Nakhon cultivar showed the highest total phenolic content of 49.68 mg GAE/g extract and antioxidant activities of 4.38 mg GAE/g extract, 4.53 mg TEAC/g extract, and 92.78 mg FeSO4/g extract using 2,2 diphenyl-1-picrylhydrazyl (DPPH), 2,20-azinobis-(3-ethylbenzothiazolin-6-sulfonic acid) (ABTS), and ferric reducing antioxidant power (FRAP) assays, respectively. The resveratrol and oxyresveratrol compounds in mulberry leaves were also investigated by high-performance liquid chromatography (HPLC). The mulberry leaf extracts from the Sakon Nakhon and Buriram cultivars showed oxyresveratrol contents of 1.20 ± 0.04 mg/g extract and 0.39 ± 0.02 mg/g extract, respectively, whereas resveratrol was not detected. It was also found that the potent anti-inflammatory properties of mulberry leaf extracts and its compounds, resveratrol and oxyresveratrol, suppressed the LPS-stimulated inflammatory responses in RAW 264.7 macrophage cells by significantly reducing nitric oxide production in a concentration-dependent manner. These compounds further inhibited interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) production and suppressed the mRNA and protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated RAW 264.7 macrophage cells. Therefore, it is established that mulberry leaf extract and its bioactive compounds contribute to its anti-inflammatory activity.

Protective role of the dried white mulberry extract on the reproductive damage and fertility in rats treated with carmustine.

The present study investigated the protective effect of dried white Mulberry extract (DWME) against carmustine (Crm) induced biochemical alterations and spermatological, histopathological, and fertility damage in Wistar albino rats. Male rats were divided into four groups (control, Crm, Crm + DWME, and DWME group). It was found that Crm decreased the motility. Crm decreased the concentration (not different from control group) compared to DWME groups. Total blood MDA levels were reduced during the recovery period. Also, the recovery period reduced the MDA levels in the Crm group/testicular tissue. The GSH levels in the Crm + DWME group were the highest among all groups in the testicular tissue/experiment period. In the immunohistochemical evaluation of the testicular tissue, a high level of caspase-3 was observed in the cells that underwent meiosis in the Crm group. The most pronounced DNA damage was also detected in the Crm group. The Crm + DWME group showed the highest number of offspring born during recovery period. In conclusion, dried white mulberry extract protects against the spermatological damages caused by carmustine. Moreover, recovery period played a positive effect on spermatological parameters and fertility.

Some Common Medicinal Plants with Antidiabetic Activity, Known and Available in Europe (A Mini-Review).

Diabetes is a metabolic disease that affected 9.3% of adults worldwide in 2019. Its co-occurrence is suspected to increase mortality from COVID-19. The treatment of diabetes is mainly based on the long-term use of pharmacological agents, often expensive and causing unpleasant side effects. There is an alarming increase in the number of pharmaceuticals taken in Europe. The aim of this paper is to concisely collect information concerning the few antidiabetic or hypoglycaemic raw plant materials that are present in the consciousness of Europeans and relatively easily accessible to them on the market and sometimes even grown on European plantations. The following raw materials are discussed in this mini-review: Morus alba L., Cinnamomum zeylanicum J.Presl, Trigonella foenum-graecum L., Phaseolus vulgaris L., Zingiber officinale Rosc., and Panax ginseng C.A.Meyer in terms of scientifically tested antidiabetic activity and the presence of characteristic biologically active compounds and their specific properties, including antioxidant properties. The characteristics of these raw materials are based on in vitro as well as in vivo studies: on animals and in clinical studies. In addition, for each plant, the possibility to use certain morphological elements in the light of EFSA legislation is given.

Effects of Growth Period and Storage Methods on Primary Metabolite Contents and Antioxidant Activities of Morus alba L. Leaf.

(1) Background: Mulberry leaves have been widely consumed due to their richness in bioactive substances and high antioxidant activity. The choice of storage method to ensure the quality of mulberry leaves is a challenge in the supply process. (2) Methods: The differences in primary metabolites of freeze-dried mulberry leaf powder after 30 days of storage under different storage conditions (i.e., vacuum or non-vacuum, 4 °C or room temperature) were investigated. (3) Results: A low temperature and vacuum had better preservation effects on the types and activity of the primary metabolites of mulberry leaves, with vacuum preservation being the best. However, the types of primary metabolites in mulberry leaves were significantly reduced after non-vacuum storage at room temperature compared to those with other storage methods. Among the metabolites detected, including dehydroascorbic acid, various phenolic acids, amino acids, lipids, and carbohydrates showed a significant decrease in their contents of more than 40%, and there was a significant increase in the contents of various compounds of the muconic acid biosynthetic pathway compared to those in other storage methods. Moreover, the antioxidant activity of mulberry leaves stored at room temperature under non-vacuum conditions was also significantly reduced. (4) Conclusions: Vacuum storage is the most ideal storage method for preserving mulberry leaves.

Metabolic Actions of a Supplement of Ilex Paraguariensis (An Extract of the Leaf Standardized to 2% I-Deoxinojirimcina), White Mulberry and Chromium Picolinate in Nondiabetic Subects with Dysglycemia: A Randomized Trial.

AIM: To prove if a nutraceutical containing Ilex paraguariensis (Ilex L. spp. Aquifoliales) (an extract of the leaf standardized to 2% I-deoxinojirimcina), white mulberry (Morus spp., Moraceae), and chromium picolinate can be effective in improving glycemic status in subject with dysglycemia. METHODS: We randomized patients to consume placebo or the nutraceutical, self-administered once a day, one tablet at breakfast, for 3 months. RESULTS: A reduction in fasting plasma glucose, postprandial glucose, and glycated hemoglobin was observed with the nutraceutical combination, both compared to baseline and placebo. Data suggested a decrease in the Homeostasis Model Assessment index with the nutraceutical, both compared to baseline and placebo. The M value, an index of insulin sensitivity, obtained after nutraceutical treatment was higher compared to baseline. We recorded a decrease in total cholesterol, low-density lipoprotein-cholesterol, and triglycerides with the nutraceutical combination compared to baseline and placebo. A decrease in high-sensitivity C-reactive protein was observed with the nutraceutical combination compared to baseline and placebo. CONCLUSIONS: A nutraceutical containing Ilex paraguariensis, white mulberry, and chromium picolinate can be helpful in improving glycemic status and lipid profile in dysglycemic subjects.

Antioxidant and Anti-Inflammatory Effects of White Mulberry (Morus alba L.) Fruits on Lipopolysaccharide-Stimulated RAW 264.7 Macrophages.

In this study, the protective effects of white mulberry (Morus alba) fruits on lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages were investigated. The ethanol (EtOH) extract of white mulberry fruits and its derived fractions contained adequate total phenolic and flavonoid contents, with good in vitro antioxidant radical scavenging activity. The extract and fractions also markedly inhibited ROS generation and antioxidant activity. After treatment with the EtOH extract and its fractions, LPS stimulation-induced elevated nitric oxide (NO) production was restored, which was primarily mediated by downregulation of inducible NO synthase expression. A total of 20 chemical constituents including flavonoids, steroids, and phenolics were identified in the fractions using ultra-high-performance liquid chromatography (UHPLC)-quadrupole time-of-flight (QTOF) high-resolution mass spectrometry (HRMS). These findings provide experimental evidence of the protective effects of white mulberry fruit extract against oxidative stress and inflammatory responses, suggesting their nutraceutical and pharmaceutical potential as natural antioxidant and anti-inflammatory agents.

Ilex paraguariensis, white mulberry and chromium picolinate in patients with pre-diabetes.

AIM: to evaluate if a nutraceutical compound containing Ilex paraguariensis, White Mulberry and Chromium Picolinate can ameliorate glycemic status in patients with pre-diabetes. METHODS: we enrolled patients with IFG or IGT, not taking other hypoglycemic compounds. Patients were randomized to take placebo or the nutraceutical compound for 3 months, in a randomized, double-blind, placebo-controlled design. Both treatments were self-administered once a day, 1 tablet during the breakfast. RESULTS: a reduction of FPG was observed with the nutraceutical combination (-7.8%). Furthermore, there was a decrease of HOMA-IR with the nutraceutical combination (-7.9%). M value was higher (p < 0.05 vs baseline and p < 0.05 vs placebo) at the end of the treatment. We obtained a reduction of Tg with the nutraceutical combination (-8.3%). About 16.6% of patients treated with nutraceutical returned to have a normal glycemia (< 100 mg/dL), and all patients had an improvement of insulin-resistance, in particular 67% of patients returned to have a M value inside range of normal insulin sensitivity. CONCLUSIONS: a nutraceutical containing Ilex paraguariensis, White Mulberry and Chromium Picolinate at 500 mg can be helpful in improving glycemia and Tg value, in patients with pre-diabetes.

Identification and characterization of naturally occurring inhibitors against human carboxylesterase 2 in White Mulberry Root-bark.

White Mulberry Root-bark (WMR) is an edible Chinese herbal used for the treatment of inflammation, nephritis and asthma. This study aimed to investigate the inhibitory effects of ethanol extract from WMR against human carboxylesterase 2 (hCE2), as well as to identity and character natural hCE2 inhibitors in this herbal. Our results demonstrated that the ethanol extract of WMR displayed potent inhibitory effects against hCE2, while three major bioactive constitutes in WMR were identified on the basis of LC fingerprinting combined with activity-based screening of LC fractions. Three bioactive compounds including SD, KG and SC were efficiently identified by comparison of LC retention times, UV and MS spectral data, with the help of authentic standards. The inhibition potentials and inhibition types of these natural compounds against hCE2 were further investigated in human liver microsomes. The results demonstrated that these bioactive compounds are potent non-competitive inhibitors against hCE2, with the Ki values ranging from 0.76µM to 1.09µM. All these findings suggested that three abundant natural compounds in WMR displayed potent inhibitory effects against hCE2, which could be used as lead compounds to develop more potent hCE2 inhibitors for the alleviation of hCE2-mediated severe delayed-onset diarrhea.

A 28-Day Repeated Dose Toxicological Study of an Aqueous Extract of Morus Alba L.

Morus alba L. (white mulberry) leaves are one of the oldest recognized traditional Chinese medicines. More recently, M alba leaves and their constituents, particularly iminosugars (or azasugars), have garnered attention for their ability to maintain normal blood glucose concentrations, an effect identified in both animal studies and human clinical trials. Reducose (Phynova Group Limited) is a commercial water-soluble extract of M alba leaves standardized to 5% 1-deoxynojirimycin (DNJ), an iminosugar with α-glucosidase inhibition properties. Although there is an extensive history of consumption of M alba leaves by humans and animals worldwide, suggesting that the leaves and their extracts have a relatively good safety profile, we are unaware of safety assessments on an extract containing a higher amount of DNJ than that occurs naturally. The current 28-day repeated dose oral toxicity study in rats, conducted according to Organisation for Economic Co-operation and Development guidelines, was carried out to assess the safety of Reducose. Male and female Hsd.Han Wistar rats (4 groups of 10 animals/sex) were administered Reducose via gavage at doses of 0, 1,000, 2,000 and 4,000 mg/kg body weight (bw)/d. No treatment-related mortality or adverse effects (per clinical observations, body weight/weight gain, food consumption, ophthalmoscopy, clinical pathology, gross pathology, organ weights, or histopathology) were observed, and no target organs were identified. The no observed adverse effect level was determined to be 4,000 mg/kg bw/d for both male and female rats, the highest dose tested.

New potential phytotherapeutics obtained from white mulberry (Morus alba L.) leaves.

The present work demonstrates the profound and unique phyto-pharmacological and nutritional profile of white mulberry (Morus alba L.) leaves which containing considerable amounts of easy digestive proteins, carbohydrates, micro- and macronutrients, polyphenols, free amino acids, organic acids. The wide range of significant biopharmaceutical activities of the aqueous and polar organic solvents extracts from mulberry leaves - including antidiabetic, antibacterial, anticancer, cardiovascular, hypolipidemic, antioxidant, antiatherogenic, and anti-inflammatory - have been critically discussed. The main objective was to demonstrate the results of recently published study on the components of white mulberry leaves exhibiting their biological activity in the various pathological and health human ailments. In addition, we intend to drawn the attention of researchers and public health workers for the extended exploration of this deciduous plant leaves as the source of potential indigenous nutraceuticals and functional food products to enable development of alternative prevention and treatment protocols offered in therapy of the common non-communicable diseases and malignances.

Evaluation of acute toxicity, genotoxicity and inhibitory effect on acute inflammation of an ethanol extract of Morus alba L. (Moraceae) in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Morus alba L. (white mulberry) is used in traditional medicine worldwide, including Brazil. The leaves of this plant are used to treat inflammatory disorders. Universal interest in this plant necessitates studies on the toxicological safety and scientific substantiation of the medicinal properties of M. alba. In previous work, we investigated the acute toxicity of orally administered M. alba ethanol extract in mice. AIM OF THE STUDY: This work was designed to investigate the ethanol extract obtained from M. alba leaves for acute toxicity when intraperitoneally administered, in vivo genotoxicity, and potential to reduce acute inflammation. In order to further investigate the constituents of the extract, we also obtained the high-performance liquid chromatography (HPLC) fingerprint of the extract. MATERIALS AND METHODS: Phytochemical analysis by thin layer chromatography (TLC) was performed and the results were used to obtain the HPLC fingerprint. Acute toxicity of 300 and 2000mg/kg b.w. i.p. doses administered to mice for 14 days was evaluated. Genotoxicity was evaluated by counting the number of micronucleated polychromatic erythrocytes in the blood of mice that either received or did not receive the extract at 75, 150 and 300mg/kg b.w. per os. The anti-inflammatory effect of the same doses administered per os was investigated using the carrageenan air pouch model. RESULTS: The TLC analysis of the extract revealed the presence of a remarkable amount of flavonoids and cinnamic acids. The HPLC fingerprint showed the presence of one major peak corresponding to chlorogenic acid and two smaller peaks corresponding to flavonoids. In the toxicity assays, there were no deaths or deviations in behavior of treated mice as compared to the control at any dose. However, biochemical, hematological, and histological analyses showed that intraperitoneal injection caused several forms of damage to the mice, which were not observed in case of oral administration, studied in our previous work. Oral administration of the extract did not result in genotoxicity and considerably reduced (58.6-65.6% inhibition) leukocyte migration in all doses evaluated, in comparison with the negative control. CONCLUSIONS: The ethanol extract from M. alba leaves administered intraperitoneally possesses a greater degree of toxicity in mice when compared to per os administration. The extract was not genotoxic when ingested by mice and exhibited a highly inhibitory effect against acute inflammation, which is probably linked to the presence of chlorogenic acid and flavonoids in the composition. This work contributes to the determination of safety of the medicinal use of M. alba leaves.