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A tyrosinase inhibitor was separated from camellia pollen with the aid of solvent fraction, macroporous adsorptive resin chromatography, and high‐speed countercurrent chromatography. The inhibitor was identified to be p‐coumaric acid ethyl ester (p‐CAEE) by nuclear magnetic resonance and mass spectrum. Its inhibitory activity (IC50 = 4.89 μg/ml) was about 10‐fold stronger than arbutin (IC50 = 51.54 μg/ml). The p‐CAEE inhibited tyrosinase in a noncompetitive model with the K I and K m of 1.83 μg/ml and 0.52 mM, respectively. Fluorescence spectroscopy analysis showed the p‐CAEE quenched an intrinsic fluorescence tyrosinase. UV‐Vis spectroscopy analysis showed the p‐CAEE did not interact with copper ions of the enzyme. Docking simulation implied the p‐CAEE induced a conformational change in the catalytic region and thus changed binding forces of L‐tyrosine. Our findings suggest that p‐CAEE plays an important role in inhibiting tyrosinase and provides a reference for developing pharmaceutical, cosmetic, and fruit preservation products using pollen., p‐Coumaric acid ethyl ester was first found in camellia pollen. The inhibition is reversible and dose‐dependent. p‐Coumaric acid ethyl ester altered the structure of tyrosinase. p‐Coumaric acid ethyl ester and pollen had potential applications for pharmaceutical, cosmetic, and fruit preservation. p‐Coumaric acid ethyl ester (p‐CAEE) can inhibit the activity of tyrosinase in a noncompetitive manner and can cause the transformation of tyrosinase.

The bacterial strain of Microbulbifer sp. ALW1 has demonstrated visible ability of degrading the cell wall of Laminaria japonica, and biochemical characterization has been performed on some individual enzymes to elucidate its genetic basis. However, it still remains elusive how strain ALW1 successfully breaks down the major cell wall component alginate polysaccharide and colonizes on its marine host. In this study, a mass spectrometry-based quantitative analysis of the extracellular and intracellular proteomes was introduced to elucidate the alginate degradation pathway in ALW1 strain. Mass spectrometry and biochemical assays indicated that strain ALW1 could effectively degrade alginate polysaccharide into disaccharides and trisaccharides within 12 h. Proteome analysis identified 156 and 1,047 proteins exclusively localized in extracellular and intracellular compartments, respectively, with 1,086 protein identities of dual localization. Functional annotation of the identified proteins suggested the involvement of diverse catalytic enzymes and non-catalytic molecules for the cleavage and metabolism of alginate polysaccharide. A simplified pathway was constructed to demonstrate the extracellular digestion, active transport, and intracellular conversion of alginate polysaccharide and its fragmented oligosaccharides, casting a picture of genetic loci controlling alginate catabolism by ALW1 strain. This study aims to provide a guide for utilization and genetic manipulation of the bacterial strain ALW1 for efficient alginate oligosaccharides production by fermentation.

An effective two-dimensional liquid chromatography method has been established for the analysis of all-trans-astaxanthin and its geometric isomers from Phaffia rhodozyma employing a C18 column at the first dimension and a C30 column in the second dimension, connected by a 10-port valve using the photo-diode array detector. The regression equation of astaxanthin calibration curve was established, and the precision and accuracy values were found to be in the range of 0.32-1.14% and 98.21-106.13%, respectively. By using two-dimensional liquid chromatography, it was found that day light, ultrasonic treatment, and heat treatment have significant influence on the content of all-trans-astaxanthin in the extract from P. rhodozyma due to the transformation of all-trans-astaxanthin to cis-astaxanthin. The day light and ultrasonic treatments more likely transform all-trans-astaxanthin to 9-cis-astaxanthin, and the thermal treatment transforms all-trans-astaxanthin to 13-cis-astaxanthin. These results indicate that the two-dimensional liquid chromatography method can facilitate monitoring astaxanthin isomerization in the raw extract from P. rhodozyma. In addition, the study will provide a general reference for monitoring other medicals and bioactive chemicals with geometric isomers.

The present study aims to evaluate the deodorization of Bangia fusco-purpurea using microorganism fermentation with commercial starter cultures of Saccharomyces cerevisiae, Acetobacter pasteurianus, and Lactobacillus plantarum. The results showed the fermentation with S. cerevisiae, A. pasteurianus, and L. plantarum resulted in significantly decreases (p < .05) of the fishy malodor in B. fusco-purpurea. Among the three strains, S. cerevisiae was the best for reducing the fishy malodor. The optimal inoculum size and fermentation time were 0.2% and 4 hr, respectively. After the fermentation with the S. cerevisiae, the content of 1-octen-3-ol, (E)-2-octen-1-ol, hexanal, non-(2E)-enal, (E,E)-2,4-decadienal, 3,5-octadien-2-one, and 2-pentyl-furan were hard to be detected in the seaweed, whereas increases were observed in the concentrations of 2-butyl-1-octanol, cedrol, diisobutyl phthalate, and 2,4-di-t-butylphenol. The odor active value analysis indicated the removal of fishy odor was related to the reduction, dehydrogenation, and deformylating oxygenation of hexanal, nonanal, non-(2E)-enal, and (E,E)-2,4-decadienal and esterification of 1-octen-3-ol and (E)-2-octen-1-ol. Our findings provide a technical and scientific basis for the removal of fishy odor from B. fusco-purpurea. PRACTICAL APPLICATIONS: Bangia fusco-purpurea is a seaweed that can reduce the risks of cardiovascular and chronic metabolic diseases in human body. However, the seaweed has a strong fishy malodor, which largely declines consumer's acceptance. In this study, the commercial starters of Saccharomyces cerevisiae, Acetobacter pasteurianus, and Lactobacillus plantarum were shown to reduce the fishy malodor in B. fusco-purpurea via fermentation. After the fermentation with the microorganisms especially with the S. cerevisiae, the fishy malodor significantly reduced, and the overall aroma acceptance of B. fusco-purpurea products greatly improved. Therefore, this study provides a technical basis for the removal of fishy odor from B. fusco-purpurea and processing value-added products from it and facilitating its health benefits for human.

Astaxanthin is a natural pigment that possesses benefit potentials to prevent obesity, however, its underlying mechanism is not clarified. In the current study, Phaffia rhodozyma astaxanthin was prepared and its inhibitory effect on pancreatic lipase was evaluated. The results showed that astaxanthin significantly inhibited the activity of pancreatic lipase in a dose-dependent manner within the tested concentration ranges. The kinetic analysis demonstrated that astaxanthin noncompetitively inhibited pancreatic lipase activity. In addition, astaxanthin induced secondary structure changes which resulted in decreased enzyme activities. Furthermore, the molecular docking analysis revealed that astaxanthin blocked the channel of catalytic site to delay substrate entrance or prevent product diffusion through changing the catalytic site conformation. These findings not only shed light on the underlying inhibitory mechanism of astaxanthin on pancreatic lipase, but also provide scientific evidence for extending the application of astaxanthin in functional food industries.

Tyrosinase inhibitors are important in cosmetic, medical, and food industries due to their regulation of melanin production. A tyrosinase inhibitor was purified from Camellia pollen using high-speed countercurrent chromatography and preparative high-performance liquid chromatography and was identified as caffeine by NMR and mass spectrometry. It showed strong mushroom tyrosinase inhibitory activity with an IC50 of 18.5 ± 2.31 μg/mL in a noncompetitive model. The caffeine did not interact with copper ions in the active center of the enzyme but could quench fluorescence intensity and change the secondary conformation of this tyrosinase. A molecular dynamics simulation showed that caffeine bound this tyrosinase via Lys379, Lys 376, Asp357, Glu356, Thr308, Gln307, Asp312, and Trp358, thus changing the binding sites of l-tyrosine and the loop conformation adjacent to the active center. In vitro cell model analysis revealed that caffeine exhibited significant inhibitory effects on both intracellular tyrosinase activity and melanin production of B16-F10 melanoma cells in a concentration-dependent manner. These comprehensive results suggest that caffeine is a strong tyrosinase inhibitor that has the potential to be developed as skin-whitening agents in the cosmetics and pharmaceutical industries or as antibrowning agents in the food industry.

An effective high-speed counter-current chromatography (HSCCC) method was established for the preparative isolation and purification of astaxanthin from Phaffia rhodozyma. With a two-phase solvent system composed of n-hexane-acetone-ethanol-water (1:1:1:1, v/v/v/v), 100mg crude extract of P. rhodozyma was separated to yield 20.6mg of astaxanthin at 92.0% purity. By further one step silica gel column chromatography, the purity reached 99.0%. The chemical structure of astaxanthin was confirmed by thin layer chromatography (TLC), UV spectroscopy scanning, high performance liquid chromatography with a ZORBAX SB-C18 column and a Waters Nova-pak C18 column, and ESI/MS/MS.

The hypoglycaemic effect of all-trans astaxanthin is long known for its antioxidation for type 2 diabetes, a global health concern. To reduce glycaemic index of type 2 diabetes, inhibition of key enzymes is an efficient approach, for example to inhibit α-glucosidase. Conventional chemical or synthetic enzyme-inhibitors have adverse side effects. Therefore, we studied the inhibition of α-glucosidase by all-trans astaxanthin. To inhibit α-glucosidase, the half maximal inhibitory concentration (IC50) of all-trans astaxanthin was 67.95 ± 0.03 μmol/L. All-trans astaxanthin inhibited α-glucosidase as a competitive inhibitor, as shown by kinetic analysis. All-trans astaxanthin could induce the changes of the secondary structure to reduce α-glucosidase activity. Molecular-docking analysis reveals that all-trans astaxanthin prevented substrate from binding to α-glucosidase by occupying the space of the active pocket to cause the inhibition. Our findings suggest that all-trans astaxanthin has the hypoglycaemic effect through inhibiting α-glucosidase, in addition to its antioxidation effect reported earlier.

Astaxanthin from different sources possesses different biological activities and optical isomers. The ingredients of astaxanthin mixtures from different sources on the market have often been mislabeled. Therefore, it is important to determine the sources of astaxanthin and their respective concentrations in a mixture. To solve this problem, a quantitative analysis model was established and further verified. The results showed that the deviation between the calculated concentration and the actual concentration ranged from 0 to 7 &micro, g/mL, and the recovery rate was between 88.90% and 103.56%. This indicates that the quantitative analysis model of astaxanthin was feasible and reliable. This study not only has important applications in the astaxanthin mixture component determination but may also shed light on the quantitative analysis of other sample mixtures with stereoisomers from different sources.

Separation of analytes is an important factor affecting the effectiveness of carotenoids analysis from a complex extraction. In this study, a high performance liquid chromatography procedure with improved separation was developed to determine carotenoids extracted from P. rhodozyma. Mixture design experiment was employed to optimize the organic solvent constitution, which showed that the optimal organic phase was composed (v/v) of 61% of methanol, 32% of tetrahydrofuran, and 7% of acetonitrile. Furthermore, the gradient elution procedure optimization demonstrated that the gradient elution started at the optimal organic phase concentration of 60% lead to the best separation effect. Meanwhile, 18 compounds in total 25 detected peaks were separated. Based on their absorption spectra, astaxanthin, s-carotene, zeaxanthin, β-cryptoxanthin and lycopene were identified by matching their retention times with standards. Interestingly, our improved HPLC method was effective to classify P. rhodozyma strains and monitor the degradation of astaxanthin in accordance with their carotenoids composition.