Response Surface Methodology for Optimization of Process Parameters and Antioxidant Properties of Mulberry (Morus alba L.) Leaves by Extrusion

Mulberry (Morus alba L.) leaves (MLs), originally used to feed silkworms, have recently been recognized as a food ingredient containing health-beneficial, bioactive compounds. In this study, the extrusion process was applied for the enhancement of the amount of extractable flavonoids from MLs. Extrusion conditions were optimized by water solubility index, total phenolic content, and total flavonoid content (TF) using response surface methodology, and antioxidative stress activities were evaluated in macrophage cells. According to the significance of regression coefficients of TF, the optimal extrusion parameters were set as barrel temperature of 114 °C, moisture feed content of 20%, and screw speed of 232 rpm. Under these conditions, the TF of extruded ML reached to 0.91% and improved by 63% compared with raw ML. Fifteen flavonoids were analyzed using ultra-high-performance liquid chromatograph coupled with photodiode array detection and quadrupole time-of-flight mass spectrometry (UPLC-PDA-QTOF/MS), and the extrusion resulted in increases in quercetin-3-gentiobioside, quercetin-3,7-di-O-glucoside, kaempferol-3,7-di-O-glucoside, rutin, isoquercitrin, and moragrol C. Besides, regarding antioxidative activity, extruded ML water extract inhibited the production of H₂O₂-induced reactive oxygen species and attenuated nuclear morphology alterations in macrophage cells. The findings of this study should be useful in food processing design to improve the extractable functional compounds in MLs.     

Evaluation of the Chemical Composition and Antioxidant Activity of Mulberry (Morus alba L.) Fruits from Different Varieties in China

Mulberry (Morus alba L.) fruit is a fruit with nutritional and medicinal value. It is widely cultivated in different regions of China, which may result in differences in its chemical composition. In this research, 25 mulberry fruit samples from six provinces in China were investigated. The contents of anthocyanins were evaluated by high-performance liquid chromatography (HPLC). The contents of two main anthocyanins, cyanidin-3-O-glucoside (C3G) and cyanidin-3-O-rutinoside (C3R), ranged from 0.656 ± 0.006 mg/g to 4.367 ± 0.243 mg/g and from 0.226 ± 0.007 mg/g to 1.649 ± 0.013 mg/g, respectively. Additionally, the contents of total phenolic, total flavonoid, vitamin C, titratable acids, reducing sugars and antioxidant capacity (FRAP, DPPH, scavenging and hydroxyl radical scavenging activity) were also assessed. The results and principal component analysis showed that the Zhongsang 5801 variety from Sichuan, Dechang had the greatest health value with the highest active compound contents. Based on our analysis, the variety from Sichuan, Dechang is a high-quality plant source for mulberry fruit cultivation. This research provides a basis for the rational development and utilization of mulberry fruit resources in China.     

Impact of Tea Processing on Tryptophan,Melatonin, Phenolic and Flavonoid Contents in Mulberry (Morus alba L.) Leaves: Quantitative Analysis by LC-MS/MS

Mulberry (Morus alba L.) leaves from two cultivars, Yai-Burirum (YB) and Khunphai (KP), were prepared into green tea (GT) and black tea (BT). Compared to fresh leaf (FL) extract, GT and BT extracts were evaluated for their total phenolic and total flavonoid contents. Total phenolic content (TPCs) in all samples ranged between 129.93 and 390.89 mg GAE/g extract. The processing of tea decreased the levels of TPC when compared to FL extracts in both cultivars. The total flavonoid content (TFCs) in all samples was found in the range of 10.15–39.09 mg QE/g extract and TFCs in GT and BT extracts were higher than FL extracts. The change in tryptophan, melatonin, phenolic and flavonoid contents was investigated by liquid chromatography–mass spectroscopy (LC-MS). The results exhibited that tryptophan contents in all samples were detected in the range 29.54–673.72 µg/g extract. Both GT and BT extracts increased tryptophan content compared to FL extracts. BT extracts presented the highest amounts of tryptophan among others in both cultivars. Phenolic compounds were found in mulberry leaf extracts, including gallic acid, caffeic acid, gentisic acid, protocatechuic acid and chlorogenic acid. Chlorogenic acid presented the highest amount in all samples. Almost all phenolic acids were increased in the processed tea extracts except chlorogenic acid. Rutin was the only flavonoid that was detected in all extracts in the range 109.48–1009.75 mg/g extract. The change in phenolic and flavonoid compounds during tea processing resulted in the change in antioxidant capacities of the GT and BT extracts. All extracts presented acetylcholinesterase enzyme (AChE) inhibitory activity with IC₅₀ in the range 146.53–165.24 µg/mL. The processing of tea slightly increased the AChE inhibitory effect of GT and BT extracts. In conclusion, processed tea from mulberry leaves could serve as a new alternative functional food for health-concerned consumers because it could be a promising source of tryptophan, phenolics and flavonoids. Moreover, the tea extracts also had antioxidative and anti-AChE activities.     

The Effect of Maturity and Extraction Solvents on Bioactive Compounds and Antioxidant Activity of Mulberry (Morus alba) Fruits and Leaves

Cultivation location, maturity levels, and extraction solvents could affect the bioactive compounds and biological activities of mulberry (Morus alba Linnaeus). The lack of study on Malaysia-grown mulberry causes its underutilization. This study investigated the bioactive compound content and the antioxidant activity of Sabah-grown mulberry at two different maturity stages (fruits: red mature and black fully ripe; leaves: young and mature) extracted using 70% (v/v) methanol, 60% (v/v) ethanol, and 65% (v/v) acetone. Analyses showed that mulberry fruits demonstrated maturity-dependent increment (except UHPLC-DAD quantification), while the leaves revealed maturity-dependent reduction. Principal component analysis (PCA) displayed 65% (v/v) acetone black fully ripe fruits as the best phenolics and antioxidant sources. However, the 60% (v/v) ethanol black fully ripe fruits contained 20.08–68.43% higher total anthocyanins. Meanwhile, the 65% (v/v) acetone and 70% (v/v) methanol red mature fruits were higher in chlorogenic acid (27.53–47.12%) and rutin (31.42–35.92%) than other fruit extracts, respectively. For leaves, 65% (v/v) acetone young leaves were the best phenolics and antioxidant sources. However, the 60% (v/v) ethanol young leaves possessed greater chlorogenic acid (19.56–74.11%) than other leaf extracts. Overall, Malaysia-grown mulberry is rich in phenolics and antioxidants, suggesting its potential application in food and pharmaceutical products.     

Anti-inflammatory effects of mulberry (Morus alba L.) root bark and its active compounds

Abstract

Mulberry (Morus alba L.) root bark (MRB) was extracted using methanol and the extracts were subjected to tests of anti-inflammatory effects. The ethyl acetate fraction demonstrated the best anti-inflammatory effects. Purified compounds, sanggenon B, albanol B and sanggenon D, showed inhibitory effects on NO production in LPS-stimulated RAW264.7 cells and albanol B demonstrated the best anti-inflammatory effects. Regarding the underlying molecular mechanisms, further investigations showed that treatments with Albanol B reduced production of pro-inflammatory cytokines and decreased expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). These results would contribute to development of novel anti-inflammatory drugs from MRB.     

Analgetic Activity of Papaya (Carica papaya L.) Leaves Extract

The analgetic activity of Carica papaya leaves (CPL) extracts (n-hexane, ethyl acetate, ethanol) was investigated in mice model using acetic acid induced pain (Siegmund method). Experimental animals were divided into 11 groups and received n-hexane, ethyl acetate, and ethanol extracts at doses of 0.175, 0.35, 0.70 mg/kg bw orally; CMC-Na 0.5% (control group); 50 mg/ kg bw of aspirin. The results showed that all extracts at the doses of 0.175, 0.35 and 0.70 mg/kg bw gave significant analgetic activity (p<0.05) compared to control group. Ethanol extract of CPL dose of 0.70 mg/kg bw showed the best analgetic activity that was comparable to aspirin.     

Quantitative analysis of aglycone quercetin in mulberry leaves (Morus alba L.) by capillary zone electrophoresis.

A capillary zone electrophoresis method was established for analysis of aglycone quercetin in mulberry leaves (Morus alba L.). The influence of, e.g., background electrolyte concentrations and pH, surfactant concentrations, organic solvents, temperature, and voltage on the separation of aglycone quercetin, rutin, quercitrin, kaempferol, catechin, and gallic acid was systematically investigated. The optimum condition providing baseline separation of all compounds within 16.5 min was obtained in 150 mM boric acid (pH 10.0) using a fused-silica capillary with an effective length of 42.5 cm (50 microm inner diameter), temperature of 32 degrees C, and voltage of 15 kV. Method assessment was performed by standard addition method using rutin as an internal standard. Linearity of the method was excellent (r(2) > 0.999) over the concentration tested (40-160 microg/mL). The relative standard deviations (%RSDs) from injection, intraday, and interday precision were less than 2.5%. Recoveries were good (asymptotically equal to 100.0%,%RSD = 0.8%) with a limit of detection (LOD) and limit of quantitation (LOQ) of 0.86 and 3.16 microg/mL (%RSD = 1.8%), respectively. The aglycone quercetin found in the mulberry leaves was 0.452 g/100 g (%RSD = 0.6%) on dry weight.     

Beneficial Effects of Laurel (Laurus nobilis L.) and Myrtle (Myrtus communis L.) Extract on Rat Health

Polyphenols of Laurel and Myrtle exhibit structural diversity, which affects bioavailability, metabolism, and bioactivity. The gut microbiota plays a key role in modulating the production, bioavailability and, thus the biological activities of phenolic metabolites, particularly after the intake of food containing high-molecular-weight polyphenols. The aim of this study was to investigate whether the polyphenolic components of Laurel and Myrtle aqueous extract have beneficial effects on rat health. The growth of lactic acid bacteria (LAB), β-glucuronidase, β-glucosidase, β-galactosidase activity, pH value, body weight change and food efficacy ratio after intragastric treatment of rats with Laurel and Myrtle extract at doses of 50 and 100 mg/kg for two weeks were investigated. The endogenous populations of colonic probiotic bacteria (Lactobacilli and Bifidobacteria) were counted on selective media. According to the obtained data, Laurel extract in the applied dose of 50 and 100 and Myrtle extract (100 mg/kg) positively affects the rats health by increasing the number of colonies of Lactobacilli and Bifidobacteria compared to the control group, causes changes in glycolytic enzymatic activity and minor change in antioxidative tissue activity. In addition, high doses of Laurel increase food efficiency ratio, while Myrtle has the same effect at a lower dose.     

Improvement of stilbenoid production by 2-hydroxypropyl-β-cyclodextrin in white mulberry (Morus alba L.) callus cultures

Mulberroside A, oxyresveratrol and resveratrol, commonly found in Morus alba L., are potent anti-aging phytostilbenes. In this study, the effect of the addition of 2-hydroxypropyl-β-cyclodextrin on the levels of phytostilbenes in M. alba callus cultures was investigated. Commercial cyclodextrin was used in the hydrolytic and culture processes of the M. alba callus cultures. The hydrolytic study indicated that 2-hydroxypropyl-β-cyclodextrin acted as a retardant for stilbenoid hydrolysis. It reduced mulberroside A deglycosylation and stabilised oxyresveratrol. The elicitation result showed that extracellular oxyresveratrol was increased by adding 2-hydroxypropyl-β-cyclodextrin to the culture media of both free and immobilised M. alba callus (>730-fold and >169-fold, respectively) compared with those of the control. However, the intracellular mulberroside A levels in the treatment groups did not increase compared with those of the control. The results show that the addition of 2-hydroxypropyl-β-cyclodextrin significantly changed the patterns and levels of the stilbenoids in M. alba callus cultures.     

柱前衍生化高效液相色谱荧光检测法测定桑叶中的1-脱氧野尻霉素

建立了芴甲氧酰氯(FMOC—Cl)柱前荧光衍生一高效液相色谱法测定桑叶中1-脱氧野尻霉素(DNJ)的方法。桑叶经0．05mol／L,盐酸提取,在pH8．5的硼酸盐缓冲溶液条件下,DNJ反应生成荧光产物,然后用高效液相色谱-荧光检测器测定。流动相为乙腈-0．1％醋酸(55：45,V/V)。线性范围为0．567～34mg／L(相关系数r=0．9999);检出限为0．03mg／L。实验测得桑叶中DNJ含量为0．24％;平均回收率为97．1％,RSD为1．35％(n=6)。     

Lipolytic effect of compounds isolated from leaves of mulberry (Morus alba L.) in 3T3-L1 adipocytes

In this study, 19 known compounds were isolated from mulberry (Morus alba L.) leaves. The lipid accumulation inhibitory activity of the isolated compounds was investigated. Compounds 4 and 12 showed good anti-adipogenic activity based on 3T3-L1 adipocytes with values of 36.6 ± 9.0 and 34.7 ± 4.0%, respectively. In addition, compounds 3, 6 and 15 showed significant inhibitory activity with values from 15.4 to 21.2% and compounds 2, 8–9 and 17–18 exhibited weak activity with values ranging from 2.1 to 10.7% at a concentration of 40.0 μM. These results show the potentiality that mulberry leaf is an excellent inhibitory phytochemical source against lipid accumulation.     

柱前衍生化高效液相色谱-荧光检测法测定桑叶中的1-脱氧野尻霉素

采用柱前衍生化高效液相色谱-荧光检测法测定了桑叶中的1-脱氧野尻霉素(DNJ)。用0.05 mol/L HCl提取桑叶中的DNJ,采用6-氨基喹啉基-N-羟基琥珀酰亚氨基甲酸酯(AQC)试剂在pH 8.5硼酸盐缓冲液下对DNJ进行衍生化,以0.02 mol/L磷酸二氢钾缓冲液(pH 5.0)-乙腈(体积比为85∶15)为流动相,利用C18色谱柱(5 μm,250 mm×4.6 mm)分离,在激发波长为250 nm、发射波长为395 nm条件下进行荧光检测,DNJ的AQC衍生物与衍生化试剂的水解产物分离良好。方法的线性范围为0.5～25 mg/L,检出限为0.02 mg/L（S/N=3）。实验测得桑叶中DNJ含量为0.12%;回收率为96.1%～98.6%。     

LC-MS Metabolite Profiling and the Hypoglycemic Activity of Morus alba L. Extracts

Morus alba L. is used in traditional Chinese medicine for its anti-diabetic activity; however, the part of the hypoglycemic activity and related active metabolites are still not fully clarified. In this study, the metabolites in the M. alba roots, leaves, twigs, and fruits extracts (70% ethanol extracts) were systematically identified, and their hypoglycemic activity was evaluated by the high-fat diet/streptozotocin-induced 2 diabetes mellitus (T2D) mouse model. A total of 60 high-level compounds, including 16 polyphenols, 43 flavonoids, and one quinic acid, were identified by high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS) combined with the fragmentation pathways of standards and the self-established database. Among them, 23 metabolites were reported for the first time from this plant. In contrast to the extracts of M. alba leaves and fruits, the extracts of roots and twigs displayed significant hypoglycemic activity The glycemia was significantly reduced from 32.08 ± 1.27 to 20.88 ± 1.82 mmol/L and from 33.32 ± 1.98 to 24.74 ± 1.02 mmol/L, respectively, after 4 weeks of treatment with roots and twigs extracts. Compound 46 (morusin), which is a high-level component identified from the extracts of M. alba roots, also displayed significant activity in decreasing the blood glucose level of T2D mice reduced from 31.45 ± 1.23 to 23.45 ± 2.13 mmol/L. In addition, the extracts of roots and twigs displayed significant activity in reducing postprandial glycemia. This work marks the first comparison of the metabolites and hypoglycemic activity of M. alba roots, leaves, twigs, and fruits extracts, and provides a foundation for further development of M. alba extracts as anti-diabetic drugs.     

Activity-guided investigation of Carissa carandas (L.) roots for anti-inflammatory constituents

The present study was structured to investigate the anti-inflammatory potential of the extracts, fractions and compounds isolated from Carissa carandas (L.) roots. Bioassay guided fractionation of methanol extract based on inhibitory potential towards proinflammatory mediators (TNF-α, IL-1β and nitric oxide (NO)) led to the identification of stigmasterol (1), lupeol (2), oleanolic acid (3), carissone (4) and scopoletin (5) as potential anti-inflammatory agents. Carissone (4) (IC₅₀ = 20.1 ± 2.69 μg/mL) and scopoletin (5) (IC₅₀ = 24.6 ± 1.36 μg/mL) exhibited significant inhibition of NO production comparable to specific NO inhibitor (L-NAME; IC₅₀ = 19.82 ± 1.64 μg/mL) without affecting the cell viability. Also, 4 and 5 at a concentration of 30 μM were found to inhibit 41.88–53.44% of TNF-α and IL-1β. To the best of our knowledge, this is the first report displaying the anti-inflammatory effects of C. carandas (L.) roots, partially mediated by inhibition of TNF-α, IL-1β and NO.     

高效液相色谱法同时测定烟叶中茄尼醇和辅酶Q10的含量

高效液相色谱法同时测定烟叶中茄尼醇和辅酶Q10的含量;反相高效液相色谱;二级管阵列检测器;烟叶;茄尼醇;辅酶Q10     

Antioxidant,Antibacterial, and Antifungal Activities of the Ethanolic Extract Obtained from Berberis vulgaris Roots and Leaves

This work assessed the phenolic and flavonoid components and their antioxidant, antifungal, and antibacterial effects in the ethanolic extract of barberry leaf and roots. The antibactericidal activity of root and leaf extracts against pathogenic bacteria was tested using agar diffusion and microdilution broth production for the lowest inhibitory concentration (MIC). Berberis vulgaris root and leaf extracts inhibited Staphylococcus aureus ATCC9973, Escherichia coli HB101, Staphylococcus enteritis, and Escherichia coli Cip812. The disc assay technique was used to assess the bactericidal activity of the extracts versus both pathogenic Gram-positive and Gram-negative strains. Hydro alcoholic extract was more effective against bacterial than fungal strains. The results showed that Berberis vulgaris leaf and roots extract had similar antifungal activities. Berberis vulgaris root extract inhibited the mycelial growth of Penicillium verrucosum, Fusarium proliferatum, Aspergillus ochraceous, Aspergillus niger, and Aspergillus flavus. Berberis vulgaris root extract has excellent antioxidant, antibacterial, and antifungal effects. Berberis vulgaris exhibited antimicrobial activity in vitro, and MIC showed that Berberis vulgaris parts efficiently affected pathogens in vitro. In conclusion, both Berberis vulgaris roots and leaves have considerable antibacterial activity and can be used as a source of antibacterial, antioxidant, and bioactive compounds to benefit human health.     

Chemical Profile of Launaea nudicaulis Ethanolic Extract and Its Antidiabetic Effect in Streptozotocin-Induced Rats

Launaea nudicaulis is used in folk medicine worldwide to treat several diseases. The present study aimed to assess the antidiabetic activity of L. nudicaulis ethanolic extract and its effect on diabetic complications in streptozotocin-induced hyperglycemic rats. The extract was orally administrated at 250 and 500 mg/kg/day for 5-weeks and compared to glibenclamide as a reference drug at a dose of 5 mg/kg/day. Administration of the extract exhibited a potential hypoglycemic effect manifested by a significant depletion of serum blood glucose concurrent with a significant elevation in serum insulin secretion. After 5-weeks, extract at 250 and 500 mg/kg/day decreased blood glucose levels by about 53.8 and 68.1%, respectively, compared to the initial values (p ≤ 0.05). The extract at the two dosages prevented weight loss of rats from the 2nd week till the end of the experiment, compared to diabetic control rats. The extract further exhibited marked improvement in diabetic complications including liver, kidney and testis performance, oxidative stress, and relative weight of vital organs, with respect to diabetic control. Histopathological examinations confirmed the previous biochemical analysis, where the extract showed a protective effect on the pancreas, liver, kidney, and testis that degenerated in diabetic control rats. To characterize extract composition, UPLC-ESI–qTOF-MS identified 85 chromatographic peaks belonging to flavonoids, phenolics, acyl glycerols, nitrogenous compounds, and fatty acids, with four novel phenolics reported. The potential anti-diabetic effect warrants its inclusion in further studies and or isolation of the main bioactive agent(s).     

Biosynthesis of Zinc Oxide Nanoparticles from Acacia nilotica (L.) Extract to Overcome Carbapenem-Resistant Klebsiella Pneumoniae

Recently, concerns have been raised globally about antimicrobial resistance, the prevalence of which has increased significantly. Carbapenem-resistant Klebsiella pneumoniae (KPC) is considered one of the most common resistant bacteria, which has spread to ICUs in Saudi Arabia. This study was established to investigate the antibacterial activity of biosynthesized zinc oxide nanoparticles (ZnO-NPs) against KPC in vitro and in vivo. In this study, we used the aqueous extract of Acacia nilotica (L.) fruits to mediate the synthesis of ZnO-NPs. The nanoparticles produced were characterized by UV-vis spectroscopy, zetasizer and zeta potential analyses, X-ray diffraction (XRD) spectroscopy, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), and transmission electron microscopy (TEM). The antimicrobial activity of ZnO-NPs against KPC was determined via the well diffusion method, and determining minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC), the results showed low MIC and MBC when compared with the MIC and MBC of Imipenem and Meropenem antibiotics. The results of in vitro analysis were supported by the results upon applying ZnO-NP ointment to promote wound closure of rats, which showed better wound healing than the results with imipenem ointment. The biosynthesized ZnO-NPs showed good potential for use against bacteria due to their small size, applicability, and low toxicity to human cells.     

Two new phenolic constituents from the root bark of Morus alba L. and their cardioprotective activity

A new biphenyl-furocoumarin, named morescoumarin A (1), and a new prenylated flavanone, named morflavanone A (2) were isolated from the root bark of Morus alba L., together with four known compounds (3–6). Their structures were determined by extensive spectroscopic analyses and comparison with literature data. The cardioprotective effects of these compounds against doxorubicin-induced cell death were evaluated by MTT method.     

Adlay Seed (Coix lacryma-jobi L. var. Ma-yuen Stapf.) Ethanolic Extract Fractions and Subfractions Induce Cell Cycle Arrest and Apoptosis in Human Breast and Cervical Cancer Cell Lines

The antitumor effects of Coix lacryma-jobi L. var. ma-yuen Stapf. (adlay seed) ethanolic extract have been increasingly shown. This study aimed to investigate the beneficial effects of both the fractions and subfractions of adlay seed ethanolic extract on the human breast (MCF-7) and cervical (HeLa) cancer cell lines, as well as exploring their possible mechanisms of action. The ethanolic extracts were obtained from different parts of adlay seed, including AHE (adlay hull extract), ATE (adlay testa extract), ABE (adlay bran extract) and PAE (polished adlay extract). The results of a 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl- tetrazolium bromide (MTT) assay showed that AHE-Ea and ATE-Ea showed significant growth inhibitory effects in a dose-dependent manner. The results also showed that the AHE-Ea-K, AHE-Ea-L, ATE-Ea-E and ATE-Ea-F subfractions inhibited cell proliferation, induced cell cycle arrest in the G0/G1 phase and decreased CDK4/Cyclin D1 protein expression. Finally, the extract activated caspase-3 activity and PARP protein expression, which induced MCF-7 and HeLa cell apoptosis. We then used liquid chromatography–mass spectrometry (LC/MS) to identify the potential active components., Quercetin showed an anticancer capacity. In conclusion, the AHE-Ea-K, AHE-Ea-L, ATE-Ea-E and ATE-Ea-F subfractions showed antitumor effects through the inhibition of MCF-7 and HeLa cell line viability, as well as inducing apoptosis and cell cycle arrest.