The Optimization of Extraction Process, Antioxidant, Whitening and Antibacterial Effects of Fengdan Peony Flavonoids.

Flavonoids have important biological activities, such as anti-inflammatory, antibacterial, antioxidant and whitening, which is a potential functional food raw material. However, the biological activity of Fengdan peony flavonoid is not particularly clear. Therefore, in this study, the peony flavonoid was extracted from Fengdan peony seed meal, and the antioxidant, antibacterial and whitening activities of the peony flavonoid were explored. The optimal extraction conditions were methanol concentration of 90%, solid-to-liquid ratio of 1:35 g:mL, temperature of 55 °C and time of 80 min; under these conditions, the yield of Fengdan peony flavonoid could reach 1.205 ± 0.019% (the ratio of the dry mass of rutin to the dry mass of peony seed meal). The clearance of Fengdan peony total flavonoids to 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical, hydroxyl radical and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) free radical could reach 75%, 70% and 97%, respectively. Fengdan peony flavonoid could inhibit the growth of the Gram-positive bacteria. The minimal inhibitory concentrations (MICs) of Fengdan peony flavonoid on S. aureus, B. anthracis, B. subtilis and C. perfringens were 0.0293 mg/mL, 0.1172 mg/mL, 0.2344 mg/mL and 7.500 mg/mL, respectively. The inhibition rate of Fengdan peony flavonoid on tyrosinase was 8.53-81.08%. This study intensely illustrated that the antioxidant, whitening and antibacterial activity of Fengdan peony total flavonoids were significant. Fengdan peony total flavonoids have a great possibility of being used as functional food materials.

Cytological Effects of Bleaching Agent (Quneex) on Plant Cells and Plant DNA.

BACKGROUND AND OBJECTIVE: There have been a number of reported drawbacks and efficacy issues regarding the use of bleaching agents in the plant industry. This study was conducted to determine the cytological effects of the bleaching agent (Quneex) on the plant cells and plant DNA using the Allium cepa assay. MATERIALS AND METHODS: It was subjected sixteen root meristems of A. cepa to different concentrations of the bleaching agent (0.1, 0.2, 0.3, 0.4 and 0.5%) with different periods of time (6, 12 and 24 h). Recovery was done for 6, 12 and 24 h after exposure. RESULTS: The mitotic index significantly decreased with time and also decreased with increase in the concentration of the bleaching agent. Abnormal chromosomal changes reflecting mutagenesis including stickiness, laggards, bridges, C-metaphase, star-metaphase, binucleation, polyploidy, disturbance and multinucleation were observed in the different concentrations and periods of time. After recovery, a slow increase in the mitotic index was observed. All treatments with or without recovery for 12 and 24 h resulted in reduction in the amount of DNA. CONCLUSION: Bleaching agents similar to Quneex containing sodium hypochlorite have mutagenic properties that can be potentially hazardous to the environment and also to humans. Thus, there is a need to regulate the use and disposal of such chemicals into the environment particularly to the sewers, to prevent contamination of potable water, plant and biodiverse aquatic animals.

The effect of bleaching agents on the degradation of vitamins and carotenoids in spray-dried whey protein concentrate.

Previous research has shown that bleaching affects flavor and functionality of whey proteins. The role of different bleaching agents on vitamin and carotenoid degradation is unknown. The objective of this study was to determine the effects of bleaching whey with traditional annatto (norbixin) by hydrogen peroxide (HP), benzoyl peroxide (BP), or native lactoperoxidase (LP) on vitamin and carotenoid degradation in spray-dried whey protein concentrate 80% protein (WPC80). An alternative colorant was also evaluated. Cheddar whey colored with annatto (15 mL/454 L of milk) was manufactured, pasteurized, and fat separated and then assigned to bleaching treatments of 250 mg/kg HP, 50 mg/kg BP, or 20 mg/kg HP (LP system) at 50°C for 1 h. In addition to a control (whey with norbixin, whey from cheese milk with an alternative colorant (AltC) was evaluated. The control and AltC wheys were also heated to 50°C for 1 h. Wheys were concentrated to 80% protein by ultrafiltration and spray dried. The experiment was replicated in triplicate. Samples were taken after initial milk pasteurization, initial whey formation, after fat separation, after whey pasteurization, after bleaching, and after spray drying for vitamin and carotenoid analyses. Concentrations of retinol, a-tocopherol, water-soluble vitamins, norbixin, and other carotenoids were determined by HPLC, and volatile compounds were measured by gas chromatography-mass spectrometry. Sensory attributes of the rehydrated WPC80 were documented by a trained panel. After chemical or enzymatic bleaching, WPC80 displayed 7.0 to 33.3% reductions in retinol, ß-carotene, ascorbic acid, thiamin, α-carotene, and α-tocopherol. The WPC80 bleached with BP contained significantly less of these compounds than the HP- or LP-bleached WPC80. Riboflavin, pantothenic acid, pyridoxine, nicotinic acid, and cobalamin concentrations in fluid whey were not affected by bleaching. Fat-soluble vitamins were reduced in all wheys by more than 90% following curd formation and fat separation. With the exception of cobalamin and ascorbic acid, water-soluble vitamins were reduced by less than 20% throughout processing. Norbixin destruction, volatile compound, and sensory results were consistent with previous studies on bleached WPC80. The WPC80 colored with AltC had a similar sensory profile, volatile compound profile, and vitamin concentration as the control WPC80.

New Whitening Constituents from Taiwan-Native Pyracantha koidzumii: Structures and Tyrosinase Inhibitory Analysis in Human Epidermal Melanocytes.

Nontoxic natural products useful in skin care cosmetics are of considerable interest. Tyrosinase is a rate-limiting enzyme for which its inhibitor is useful in developing whitening cosmetics. Pyracantha koidzumii (Hayata) Rehder is an endemic species in Taiwan that exhibits tyrosinase-inhibitory activity. To find new active natural compounds from P. koidzumii, we performed bioguided isolation and studied the related activity in human epidermal melanocytes. In total, 13 compounds were identified from P. koidzumii in the present study, including two new compounds, 3,6-dihydroxy-2,4-dimethoxy-dibenzofuran (9) and 3,4-dihydroxy-5-methoxybiphenyl-2'-O-ß-d-glucopyranoside (13), as well as 11 known compounds. The new compound 13 exhibited maximum potency in inhibiting cellular tyrosinase activity, the protein expression of cellular tyrosinase and tyrosinase-related protein-2, as well as the mRNA expression of Paired box 3 and microphthalmia-associated transcription factor in a concentration-dependent manner. In the enzyme kinetic assay, the new compound 13 acted as an uncompetitive mixed-type inhibitor against the substrate l-3,4-dihydroxyphenylalanine and had a Km value against this substrate of 0.262 mM, as calculated using the Lineweaver-Burk plots. Taken together, our findings show compound 13 exhibits tyrosinase inhibition in human melanocytes and compound 13 may be a potential candidate for use in cosmetics.

Sensory and Functionality Differences of Whey Protein Isolate Bleached by Hydrogen or Benzoyl Peroxide.

UNLABELLED: Whey protein is a highly functional food ingredient used in a wide variety of applications. A large portion of fluid whey produced in the United States is derived from Cheddar cheese manufacture and contains annatto (norbixin), and therefore must be bleached. The objective of this study was to compare sensory and functionality differences between whey protein isolate (WPI) bleached by benzoyl peroxide (BP) or hydrogen peroxide (HP). HP and BP bleached WPI and unbleached controls were manufactured in triplicate. Descriptive sensory analysis and gas chromatography-mass spectrometry were conducted to determine flavor differences between treatments. Functionality differences were evaluated by measurement of foam stability, protein solubility, SDS-PAGE, and effect of NaCl concentration on gelation relative to an unbleached control. HP bleached WPI had higher concentrations of lipid oxidation and sulfur containing volatile compounds than both BP and unbleached WPI (P < 0.05). HP bleached WPI was characterized by high aroma intensity, cardboard, cabbage, and fatty flavors, while BP bleached WPI was differentiated by low bitter taste. Overrun and yield stress were not different among WPI (P < 0.05). Soluble protein loss at pH 4.6 of WPI decreased by bleaching with either hydrogen peroxide or benzoyl peroxide (P < 0.05), and the heat stability of WPI was also distinct among WPI (P < 0.05). SDS PAGE results suggested that bleaching of whey with either BP or HP resulted in protein degradation, which likely contributed to functionality differences. These results demonstrate that bleaching has flavor effects as well as effects on many of the functionality characteristics of whey proteins. PRACTICAL APPLICATIONS: Whey protein isolate (WPI) is often used for its functional properties, but the effect of oxidative bleaching chemicals on the functional properties of WPI is not known. This study identifies the effects of hydrogen peroxide and benzoyl peroxide on functional and flavor characteristics of WPI bleached by hydrogen and benzoyl peroxide and provides insights for the product applications which may benefit from bleaching.

Extraction of palm tree cellulose and its functionalization via graft copolymerization.

The work in this paper was planned with the aim of extracting the cellulosic component of palm tree waste and functionalizing this cellulose through graft copolymerization with acrylic acid. The cellulose extraction included hot alkali treatment with aqueous sodium hydroxide to remove the non-cellulosic binding materials. The alkali treatment was followed by an oxidative bleaching using peracid/hydrogen peroxide mixture with the aim of removing the rest of non-cellulosic materials to improve the fiber hydrophilicity and accessibility towards further grafting reaction. Optimum conditions for cellulose extraction are boiling in 5% (W/V) NaOH in a material to liquor ratio of 1:20 for 1 h then bleaching with 60 ml/l bleaching mixture at initial pH value of 6.5 for 30 min. The pH of the bleaching medium is turned to the alkaline range 11 and bleaching continues for extra 30 min. Graft copolymerization reaction was initiated by potassium bromate/thiourea dioxide redox system. Optimum conditions for grafting are 30 mmol of potassium bromate, 30 mmol of thiourea dioxide and 150 g of acrylic acid (each per 100 g of cellulose). The polymerization reaction was carried out for 120 min at 50°C using a material to liquor ratio of 1:20.

Hypopigmentary effects of ethyl P-methoxycinnamate isolated from Kaempferia galanga.

We isolated crystals from the chloroform fraction of an ethanol extract of Kaempferia galanga and identified it as ethyl p-methoxycinnamate through nuclear magnetic resonance analysis. In the present study, we found that ethyl p-methoxycinnamate significantly decreased melanin synthesis in B16F10 murine melanoma cells stimulated with α-melanocyte stimulating hormone (α-MSH). In a cell-free system, however, ethyl p-methoxycinnamate did not directly inhibit tyrosinase, the rate-limiting enzyme of melanogenesis. Instead, it inhibited tyrosinase activity in B16F10 cells in a dose-dependent manner. Furthermore, Western blot analysis showed that ethyl p-methoxycinnamate decreased microphthalmia-associated transcription factor and tyrosinase levels in α-MSH-stimulated B16F10 cells. These results indicate that the pigment-inhibitory effect of ethyl p-methoxycinnamate results from downregulation of tyrosinase. Ethyl p-methoxycinnamate isolated from K. galanga could be developed as a skin whitening agent to treat hyperpigmentary disorders.

Antioxidant capacity of leaf extracts from two Stevia rebaudiana Bertoni varieties adapted to cultivation in Mexico.

The recent introduction of the cultivation of Stevia rebaudiana Bertoni in Mexico has gained interest for its potential use as a non-caloric sweetener, but some other properties of this plant require studies. Extracts from two varieties of S. rebaudiana Bertoni adapted to cultivation in Mexico were screened for their content of some phytochemicals and antioxidant properties. Total pigments, total phenolic and flavonoids contents of the extracts ranged between 17.7-24.3 mg/g, 28.7-28.4 mg/g, and 39.3-36.7 mg/g, respectively. The variety "Criolla" exhibited higher contents of pigments and flavonoids. Trolox equivalent antioxidant capacity ranged between 618.5-623.7 mM/mg and DPPH decolorization assay ranged between 86.4-84.3%, no significant differences were observed between varieties. Inhibition of ß-carotene bleaching ranged between 62.3-77.9%, with higher activity in the variety "Criolla". Reducing power ranged between 85.2-86% and the chelating activity ranged between 57.3-59.4% for Cu²âº and between 52.2-54.4% for Fe²âº, no significant differences were observed between varieties. In conclusion, the results of this study showed that polar compounds obtained during the extraction like chlorophylls, carotenoids, phenolic compounds and flavonoids contribute to the antioxidative activity measured. The leaves of S. rebaudiana Bertoni could be used not only as a source of non-caloric sweeteners but also naturally occurring antioxidants.

La reciente introducción del cultivo de Stevia rebaudiana Bertoni en México ha ganado interés debido a su potencial uso como fuente de edulcorantes no calóricos, pero otras propiedades de esta planta aun requieren de estudios. Extractos de hojas de dos variedades de S. rebaudiana Bertoni adaptadas al cultivo en México fueron evaluados en cuanto a su contenido de algunos fitoquímicos y sus propiedades antioxidantes. El contenido de pigmentos, fenoles totales y flavonoides en los extractos, osciló entre 17.7-24.3 mg/g, 28.7-28.4 mg/g, y 39.3-36.7 mg/g, respectivamente. La variedad "Criolla" exhibió los mayores contenidos de pigmentos y flavonoides. La capacidad antioxidante equivalente de Trolox osciló entre 618.5-623.7 mM/mg y el ensayo de decoloración del radical DPPH osciló entre 86.4-84.3%, no observándose diferencias significativas entre ambas variedades. La inhibición de la decoloración del -caroteno osciló entre 62.3-77.9%, siendo mayor en la variedad "Criolla". El poder reductor osciló entre 85.2-86%, las capacidades quelantes de cobre y hierro oscilaron entre 57.3-59.4% y 52.2-54.4%, respectivamente, no observándose diferencias significativas entre ambas variedades. En conclusión, los resultados de este estudio demuestran que los compuestos de naturaleza polar obtenidos durante la extracción, tales como pigmentos clorofílicos, carotenoides, compuestos fenólicos y flavonoides contribuyen a la actividad antioxidante. Las hojas de S. rebaudiana Bertoni podrían ser empleadas no solo como fuente de edulcorantes no calóricos, sino también como fuente de antioxidantes de origen natural.

In silico prediction of the cosmetic whitening effects of naturally occurring lead compounds.

The identification of tyrosinase inhibitors is important, not only for the treatment of skin hyperpigmentation disorders, such as melasma, but also for the production of cosmetic whitening effects. The aim of this study was the in silico prediction of the naturally occurring lead compounds in three commonly used skin-whitening herbs: Ampelopsis japonica, Lindera aggregata, and Ginkgo biloba. The active ingredients responsible for the whitening effect of these herbs remain largely unknown. The tyrosinase binding affinities and skin permeation, skin irritancy, and corrosive properties of43 natural constituents of the three herbs were predicted by docking simulations using Surflex-Dock and the QSAR-based Dermal Permeability Coefficient Program (DERMWIN) and Skin Irritation Corrosion Rules Estimation Tool (SICRET) implemented in Toxtree. Nine constituents of the three herbs were found to have more advanced binding energies than the gold standard whitening agents, arbutin and kojic acid, but 40 were indicative of at least one skin sensitization alert, and many exhibited poor skin permeability. Linderagalactone c and (+)-n-methyllaurotetanine were found to have the strongest prospects for use in topical formulations, as they achieved high predicted tyrosinase binding scores and displayed good skin permeation properties and minimal potential for skin sensitization and irritation.

Validation of Eupatorium triplinerve Vahl leaves, a skin care herb from East Kalimantan, using a melanin biosynthesis assay.

In searching for a new material made from natural resources that could be used as a whitening agent, we focused on the plants used for skin treatment by the native people of East Kalimantan. The methanol extract of the leaves of Eupatorium triplinerve Vahl showed antimelanogenesis activity in a melanin biosynthesis assay. By activity-guided fractionation, 7-methoxycoumarin (1) was isolated as an active compound. The IC50 of 1 on mushroom tyrosinase was 2360 µM (L-tyrosine was used as the substrate) and above 2840 µM (L-DOPA was used as the substrate), respectively. Regarding melanin formation inhibition in B16 melanoma cells, the IC50 of 1 was 1780 µM with 83% cell viability at IC50. Based on these results, we validated that the leaf extract is in line with the traditional use of the Dayak tribe in East Kalimantan.

Inhibition of the p38 and PKA signaling pathways is associated with the anti-melanogenic activity of Qian-wang-hong-bai-san, a Chinese herbal formula, in B16 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Qian-wang-hong-bai-san (QW), a Chinese herbal formula, is traditionally used as a skin whitening agent in China. AIM OF STUDY: In our previous screening assays, QW was identified as an effective tyrosinase inhibitor. In this study, we aim to investigate the underlying mechanism of the anti-melanogenic effect of QW in B16 cells. MATERIALS AND METHODS: Cytotoxicity of QW in B16 cell line was examined by MTT assay. Cellular tyrosinase activity was determined based on the melanin content measured at 475 nm with a microplate spectrophotometer. Protein expression was analyzed by Western blotting and quantified by Quantity One. RESULTS: QW dose-dependently inhibited tyrosinase activity and decreased melanin content at 48 h without significant cytotoxicity in B16 cells. Western blot analysis showed that QW treatment down-regulated the expression levels of phospho-p38, phospho-CREB, MITF, tyrosinase, TRP-1 and TRP-2 in a dose-dependent manner. At the same time, QW treatment for 48 h inhibited IBMX-induced elevation of cellular melanin content and tyrosinase activity. However, the attenuation of IBMX-mediated up-regulations of phospho-CREB and phospho-PKA was readily observed with 60 min of QW treatment. CONCLUSIONS: The anti-melanogenic activity of QW in B16 melanoma cells can be attributed, at least in part, to the inhibition of the p38 MAPK and PKA signaling pathways. These findings shed new light on the molecular mechanisms of the skin-whitening property of QW.

Partially purified components of Nardostachys chinensis suppress melanin synthesis through ERK and Akt signaling pathway with cAMP down-regulation in B16F10 cells.

UNLABELLED: Ethnopharmacological relevance Nardostachys chinensis has been used in folk medicine to treat melasma and lentigines in Korea. We investigated the inhibitory activities of Nardostachys chinensis in melanogenesis and its related signaling pathway. MATERIALS AND METHODS: Bioassay-guided fractionation of Nardostachys chinensis using solvent partitioning and purification with octadecylsilane open-column chromatography resulted in partial purification. The active 20% methanol chromatographic fraction from the ethyl acetate layer (PPNC) was used to investigate melanogenesis by melanin synthesis, tyrosinase activity assay, cAMP assay, Western blot and flow cytometric analyses in B16F10 mouse melanoma cells. RESULTS: PPNC markedly inhibits melanin synthesis and tyrosinase activity in a concentration-dependent manner. We also found that PPNC decreases microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein (TRP)-1, and dopachrome tautomerase (Dct) protein expressions and MITF and tyrosinase mRNA levels. Moreover, PPNC reduces intracellular cAMP levels and activates mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3-kinase (PI3K)/Akt expression in B16F10 cells. The specific MEK/ERK inhibitor PD98059 and PI3K/Akt inhibitor LY294002, block the PPNC-induced hypopigmentation effect, and abrogate the PPNC-suppressed expression of melanogenic proteins such as MITF, tyrosinase, TRP-1, and Dct. Using flow cytometry, we elucidated whether PPNC directly induces ERK phosphorylation at the level of an intact single cell. PPNC shows marked expression of phosphorylated ERK in live B16F10 cells and abrogates PPNC-induced phosphorylated ERK by PD98059 treatment. CONCLUSIONS: PPNC stimulates MEK/ERK phosphorylation and PI3K/Akt signaling with suppressing cAMP levels and subsequently stimulating MITF and TRPs down-regulation, resulting in melanin synthesis suppression.

Inhibitory effects of Sargassum polycystum on tyrosinase activity and melanin formation in B16F10 murine melanoma cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Sargassum polycystum, a type of brown seaweed, has been used for the treatment of skin-related disorders in traditional medicine. AIM OF THE STUDY: The aim of the present study is to investigate the antimelanogenesis effect of Sargassum polycystum extracts by cell-free mushroom tyrosinase assay followed by cell viability assay, cellular tyrosinase assay and melanin content assay using B16F10 murine melanoma cells. MATERIALS AND METHODS: Sargassum polycystum was extracted with 95% ethanol and further fractionated with hexane, ethyl acetate and water. The ethanolic crude extract and its fractionated extracts were tested for their potential to act as antimelanogenesis or skin-whitening agents by their abilities to inhibit tyrosinase activity in the cell-free mushroom tyrosinase assay and cellular tyrosinase derived from melanin-forming B16F10 murine melanoma cells. The tyrosinase inhibitory activity was correlated to the inhibition of melanin production in α-MSH-stimulated and unstimulated B16F10 cells. RESULTS: Sargassum polycystum ethanolic extract and its fractions had little or no inhibitory effect on mushroom tyrosinase activity. However, when tested on cellular tyrosinase, the ethanolic extract and its non-polar fraction, hexane fraction (SPHF), showed significant inhibition of cellular tyrosinase activity. In parallel to its cellular tyrosinase inhibitory activity, SPHF was also able to inhibit basal and α-MSH-stimulated melanin production in B16F10 cells. CONCLUSIONS: Our findings showed that (i) cellular tyrosinase assay is more reliable than mushroom tyrosinase assay in the initial testing of potential antimelanogenesis agents and, (ii) SPHF inhibited melanogenesis by inhibiting cellular tyrosinase activity. SPHF may be useful for treating hyperpigmentation and as a skin-whitening agent in cosmetics industry.

Screening of marine algae for potential tyrosinase inhibitor: those inhibitors reduced tyrosinase activity and melanin synthesis in zebrafish.

In order to find new anti-browning and whitening agents in this study, we investigated 43 indigenous marine algae for tyrosinase inhibitory activity. The extracts from Endarachne binghamiae, Schizymenia dubyi, Ecklonia cava (EC) and Sargassum silquastrum (SS) evidenced potent tyrosinase inhibitory activity similar to that of positive control, kojic acid. Among those marine algae, EC and SS are distributed abundantly on Jeju Island. Therefore, we selected those two species for further studies. Our results evidenced that both species reduced cellular melanin synthesis and tyrosinase activity. On the other hand, we utilized zebrafish as an alternative in vivo model. All the tested samples evidenced excellent inhibitory effects on the pigmentation of zebrafish, most likely due to their potential tyrosinase inhibitory activity. In simultaneous in vivo toxicity tests, no toxicity was observed in either algal species, on the other hand, toxicity was observed in positive controls. These results provided that EC and SS extract could be used as an ingredient for whiting cosmetics and that zebrafish is an alternative in vivo model.

Invited review: Annatto usage and bleaching in dairy foods.

Annatto is a yellow/orange colorant that is widely used in the food industry, particularly in the dairy industry. Annatto, consisting of the carotenoids bixin and norbixin, is most commonly added to produce orange cheese, such as Cheddar, to achieve a consistent color over seasonal changes. This colorant is not all retained in the cheese, and thus a percentage remains in the whey, which is highly undesirable. As a result, whey is often bleached. Hydrogen peroxide and benzoyl peroxide are the 2 bleaching agents currently approved for bleaching whey in the United States. Recent studies have highlighted the negative effect of bleaching on whey flavor while concurrently there is a dearth of current studies on bleaching conditions and efficacy. Recent international mandates have placed additional concern on the use of benzoyl peroxide as a bleaching agent. This review discusses the advantages, disadvantages, regulatory concerns, flavor implications, and optimal usage conditions of 2 widely used bleaching agents, hydrogen peroxide and benzoyl peroxide, as well as a few alternative methods including lipoxygenase, peroxidase, and lactoperoxidase systems.