Skin whitening and anti-corrugation activities of glycoprotein fractions from liquid extracts of boiled sea cucumber

Objective: To determine skin whitening and wrinkle improvement efficacy, glycoprotein fractions were extracted from liquid extracts of boiled sea cucumber and their effects on tyrosine and elastase inhibitory activities were assayed. Methods: Fractions above and below 50 k Da(>50 kD a and <50 kD a) were extracted via a series of steps invovling: boiling, filtering, desalting and freeze drying. Cytotoxicity, skin whitening and wrinkle-removing effects of boiled liquid were determined. Results: Our MTT data showed that neither glycoprotein fraction of boiled liquid induces cellular cytotoxicity up to a concentration of 10 mg/m L treatment of the mouse melanoma cell line, B16F10, with 10 mg/m L >50 kDa enhanced tyrosinase and elastase inhibitory activities by 50.84% and 28.78%, respectively. Correlations of the >50 kD a concentration with tyrosinase inhibitory(R^2=0.968) and elastase inhibitory(R2=0.983) efficacy were significant. Conclusions: >50 kD a glycoprotein fraction isolated from liquid extracts of boiled sea cucumber, which can serve as a functional cosmetic ingredient for whitening and wrinkle improvement of skin.

Fatty acid methyl ester profiles and nutritive values of 20 marine microalgae in Korea

Objecive:To screen the fatty acid(FA) composition of 20 marine microalgae species,including seven Diophyceae,six Bacillariophyeae four Chlorophyceae,two Haptophyceae and one Raphidophyceae species.Methods:Microalgal cells cultured at the Korea Institute of Ocean Science & Technology were harvested during the late exponential growth phase and the FA composition analyzed.Results:The FA composition of microalgae was speciesspecific.For example,seven different species of Dinophyceae were composed primarily of C14:0,C16:0.C18:0.C20:4n-6.C20:5n-3 and C22:6n-3.while C14:0.C16:0,C16:1.C18:0.C20:5n-3 and C22:6n-3 were abundant FAs in six species of Bacillariophyceae.In addition,four Chlurophyceae,two Haptopkyeeae and one Raphidophyceae species all contained a high degree of C16:1 n-7[(9.2R-34.91)%and(34.48-35.04)%].C14:0[(13.34-25.96)%]and[(26.69-Z8.24)%],and C16:0[(5.89-29.15)%]and[(5.70-16.81)%].Several factors contribute to the nutritional value of microalgae.including the polyunsaturated FA content and n-3 to n-6 FA ratio,which could be used to assess the nutritional quality of microalgae.Conclusions:This study is the first comprehensive assessment of the FA composition and nutritional value of microalgae species in South Korea,and identifies the potential utility of FAs as species-specific biomarkers.

Detection of coat protein gene of nervous necrosis virus using loopmediated isothermal amplification

Objective:To establish a novel and highly specific loop-mediated isothermal amplification(LAMP) assay for the identification of nervous necrosis virus(NNV) infection.Methods:A set of synthesized primers was used to match the sequences of a specific region of the nnr gene from the National Center for Biotechnology Information database,not originating from NNV-infected fish,the efficiency and specificity of LAMP were measured dependent on the concentration of DNA polymerase and the reaction temperature and time.In addition,to determine species-specific LAMP primers,cross reactivity testing was applied to the reaction between NVV and other virus families including viral hemorrhagic septicemia virus and marine birnavirus.Results:The optimized LAMP reaction carried out at 64 ℃ for 60 min,and above 4 U Bst DNA polymerase.The sensitivity of LAMP for the detection of nnv was thus about 10 times greater than the sensitivity of polymerase chain reaction.The LAMP assay primers were specific for the detection NNV infection in Epinephelus septemfasciatus.Conclusions:The development of LAMP primers based on genetic information from a public database,not virus-infected samples,may provide a very simple and convenient method to identify viral infection in aquatic organisms.

Phenol content,antioxidant and tyrosinase inhibitory activity of mangrove plants in Micronesia

Objective:To find out and compare the in vitro antioxidant and tyrosinase inhibitory activities of two species of mangrove plants.Methods:Mangrove samples were harvested at the shoreline on the island of Weno,Chuuk State in Micronesia.The phenol content,antioxidant activity(based on DPPH-free radical scavenging)and tyrosinase inhibitory activity in different tissues(leaves,barks and roots)of Rhizophora stylosa(R.stylosa)and Sonneratia alba(S.alba),collected from the island of Weno.Results:Total phenol content ranged from 4.87 to 11.96 mg per g of freeze dried samples.The highest antioxidant activity was observed in R.styiosa bark(85.5%).The highest tyrosinase inhibitory activity was found in S.alba bark.Also,total phenol content and antioxidant activity were higher in methanol extracts than in aqueous extracts.Conclusions:Taken together,the results of this study proved that mangroves can be excellent sources of antioxidant compounds.

Efficient detection of pathogen virus in sand dabs,Paralichthys olivaceus using loop-mediated isothermal amplification(LAMP)

Viral hemorrhagic septicemia virus（VHSV） and marine birnavirus（MABV） are the causative pathogens for some of the most explosive epidemics of emerging viral diseases in many Asian countries, leading to huge economic losses in aquaculture. Rapid molecular detection for surveillance or diagnosis has been a critical component in reducing the prevalence of pathogen infection. The loop-mediated isothermal amplification（LAMP） of DNA is currently one of the most commonly used molecular diagnostic tools, as it is simple, quick, and easy to amplify target DNA under isothermal conditions. In the present study, a novel and highly specific LAMP assay for the sensitive and rapid detection of VHSV and MABV infection in fish was developed. Using a set of synthesized primers matching a specific region of the genome, the efficiency and specificity of the LAMP assay were optimized in terms of the reaction temperature and DNA polymerase concentration, as they are the main determinants of the sensitivity and specificity of the LAMP assay. In particular, we demonstrated that our assay could be applied to efficient detection of VHSV and MABV infection in the wild fish, Paralichthys olivaceus. Our results demonstrate the simplicity and convenience of this method for the detection of viral infection in aquatic organisms.

Integration of the nuclease protection assay with sandwich hybridization (NPA-SH) for sensitive detection of Heterocapsa triquetra

Microalgae are photosynthetic microorganisms that function as primary producers in aquatic ecosystems. Some species of microalgae undergo rapid growth and cause harmful blooms in marine ecosystems. Heterocapsa triquetra is one of the most common bloom-forming species in estuarine and coastal waters worldwide. Although this species does not produce toxins, unlike some other Heterocapsa species, the high density of its blooms can cause significant ecological damage. We developed a H. triquetra species-specific nuclease protection assay sandwich hybridization（NPA-SH） probe that targets the large subunit of ribosomal RNA（LSU r RNA）. We tested probe specificity and sensitivity with five other dinoflagellates that also cause red tides. Our assay detected H.triquetra at a concentration of 1.5×10^4 cells/m L, more sensitive than required for a red-tide guidance warning by the Korea Ministry of Oceans and Fisheries in 2015（3.0×10^4 cells/m L）. We also used the NPA-SH assay to monitor H. triquetra in the Tongyeong region of the southern sea area of Korea during 2014. This method could detect H.triquetra cells within 3 h. Our assay is useful for monitoring H. triquetra under field conditions.

Biochemical Characterization of Soluble Acid and Alkaline Invertases from Shoots of Etiolated Pea Seedlings

Soluble invertase was purified from pea （Pisum sativum L.） by sequential procedures entailing ammonium sulfate precipitation, DEAE-Sepharose column, Con-A- and Green 19-Sepharose affinity columns, hydroxyapatite column, ultra-filtration, and Sephacryl 300 gel filtration. The purified soluble acid （SAC） and alkaline （SALK） invertases had a pH optimum of 5.3 and 7.3, respectively. The temperature optimum of two invertases was 37 ℃. The effects of various concentrations of Tris-HCI, HgCI2, and CuSO4 on the activities of the two purified enzymes were examined. Tris-HCI and HgCI2 did not affect SAC activity, whereas 10 mM Tris-HCI and 0.05 mM HgCI2 inhibited SALK activity by about 50%. SAC and SALK were inhibited by 4.8 mM and 0.6 mM CuSO4 by 50%, respectively. The enzymes display typical hyperbolic saturation kinetics for sucrose hydrolysis. The Kms of SAC and SALK were determined to be 1.8 and 38.6 mM, respectively. The molecular masses of SAC shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting were 22 kDa and 45 kDa. The molecular mass of SALK was 30 kDa. Iso-electric points of the SAC and SALK were estimated to be about pH 7.0 and pH 5.7, respectively.