Extraction and Concentration of Waste Pueraria lobata Stems with Antioxidants and Anti-Melanogenesis Activity as a Novel Skin Whitening Agent for Natural Cosmetic Prototypes.

The root of Pueraria lobata (Willd.) is used commercially in different products, including dietary supplements, cosmetics, and teas, but its stem part is rarely used and studied. Therefore, this study evaluated the antioxidant and anti-melanogenesis activities of the bioactive fraction of P. lobata stem and investigated whether the activated carbon decolorization technique would have an impact on its activity and chemical composition. We observed that the dichloromethane fraction of P. lobata stem (DCM-PLS) has excellent antioxidant and anti-melanin synthesis activity at a concentration of 50 µg/mL. For the investigation of the anti-melanogenesis mechanism, we evaluated the mRNA expression of tyrosinase, which was depressed by the DCM-PLS. Daidzin was identified as the main active ingredient in DCM-PLS by using a high-performance liquid chromatography-diode array detector-hyphenated with tandem mass spectrometry. In addition, the activated carbon decolorization technology has no negative impact on the main components and bioactivity of DCM-PLS. DCM-PLS also did not induce any skin response in the human skin safety test. Collectively, DCM-PLS could be used as a natural type of skin-whitening agent in skin care products.

Anti-Wrinkle Effect of Isatis indigotica Leaf Extract: Evaluation of Antioxidant, Anti-Inflammation, and Clinical Activity.

Isatis indigotica leaf is an oriental herbal medicine that has been known for various pharmacological effects. However, its anti-wrinkle activity has not been fully evaluated. Therefore, we evaluated the anti-wrinkle effect of I. indigotica leaf extract on human skin. The purified extract inhibited 85.4% of 2,2-diphenyl-1-1picrylhydrazyl and 72.2% of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt radicals at a concentration of 1 mg/mL. Nitrite production was reduced by 30% after treatment with 50 µg/mL of extract. Three fractions from the extract downregulated the mRNA expression of matrix metalloproteinase-1 and -3 and upregulated the expression of interleukin 4. Among the three fractions, fraction 2 exhibited the highest activity. The major component of the extract was identified as 3,4,5-trimethoxycinnamic acid by liquid chromatography coupled with mass spectrometry. Molecular docking was conducted to predict the binding mechanism of 3,4,5-trimethoxycinnamic with matrix metalloproteinase-1 and -3, and their binding energies were -5.20 and -4.89 kcal/mol, respectively. In a clinical trial, five roughness values of visiometer and visual score were significantly reduced in treated groups compared with the placebo group after 8 weeks. I. indigotica leaf extract inhibits wrinkle formation, and could be a potential anti-wrinkle agent. This is the first clinical trial demonstrating its anti-wrinkle activity.

Evaluation of Anti-Melanogenesis Activity of Enriched Pueraria lobata Stem Extracts and Characterization of Its Phytochemical Components Using HPLC-PDA-ESI-MS/MS.

The root of Pueraria lobata (Willd.) is a widely used herbal medicine worldwide, whereas the stem of the plant is discarded or used as feed for livestock. To reuse and exploit the stem of P. lobata as a resource, we investigated its potential as a skin-whitening agent. We found that the developed, enriched P. lobata stem (PLS) extract significantly inhibited melanin production in the 3-isobutyl-1-methylxanthine-induced B16/F10 cells at a concentration of 50 µg/mL. To further confirm the mechanism of the antimelanogenic effect of the enriched PLS extracts, we examined the mRNA expression of tyrosinase, which was suppressed by the extracts. To standardize and implement effective quality control of the enriched PLS extracts, its major chemical constituents were identified by high-performance liquid chromatography-photodiode array-electrospray ionization-mass spectrometry. In total, 12 constituents were identified. In silico analysis showed that the main constituents, puerarin and daidzin, had excellent binding affinities for human tyrosinase. Collectively, our results suggest that the PLS extracts could be used as anti-pigmentation agents.

Evaluation of the Antiwrinkle Activity of Enriched Isatidis Folium Extract and an HPLC-UV Method for the Quality Control of Its Cream Products.

Currently, many extracts from natural sources are added to cosmetic products for reducing facial aging and wrinkles. This study investigated the antiwrinkle activity of enriched extract of Isatidis Folium used for a novel antiwrinkle cream product. The result demonstrated that this enriched extract has excellent antiwrinkle activity by significantly inhibiting mRNA expression of matrix metalloproteinase-1, matrix metalloproteinase-3, and pro-inflammatory cytokines IL-1ß and upregulating the mRNA expression of IL-4 and procollagen. Additionally, to implement effective quality control of the entire manufacturing process of antiwrinkle cream products based on the enriched extract of Isatidis Folium, the main chemical constituents of the enriched extract of Isatidis Folium was evaluated by high-performance liquid chromatography-photodiode array-tandem mass spectrometry (HPLC-PDA-ESI-MS/MS), five constituents were undisputedly confirmed. An HPLC-UV method in 15-min analysis time for quality assessment of the entire manufacturing process of antiwrinkle cream products was proposed and validated. The optimal conditions for extracting TMCA (3,4,5-trimethoxycinnamic acid) from the developed antiwrinkle cream products were determined using response surface methodology based on central composite design. The established HPLC method and optimal extract condition are suitable for routinely analyzing this novel antiwrinkle cream product.

The Effects of Non-Thermal Atmospheric Pressure Plasma treated Titanium Surface on Behaviors of Oral Soft Tissue Cells.

Here, we investigated the possible use of the technology known as non-thermal atmospheric pressure plasma on integration and control of cytokine release of soft tissue on titanium surface. After NTAPP was applied to titanium samples, changes of surface characteristics were measured as topographical features, contact angle, surface tension, and with X-ray photoelectron spectroscopy (XPS). Protein absorption was evaluated using a bovine serum albumin absorption assay. The attachment, viability, morphology, proliferation, and cytokine release of soft tissue on titanium were assessed. No change in topographical features was observed between control and NTAPP-treated groups. However, NTAPP treatment resulted in significant lowering of the contact angle for polar and non-polar liquids and increase of surface tension. Protein absorption was significantly enhanced on the NTAPP-treated samples. Normal soft tissue attachment was improved on the NTAPP-treated groups with good viability. Cellular morphology was improved in NTAPP-treated groups whereas cellular proliferation was not enhanced. There was a significant reduction in the amounts of cytokine release for inflamed IHOK and hTERT-hNOF on the NTAPP-treated groups; except for IL-8 for IHOKs. This study demonstrates that surface functional consequences by NTAPP exposure enhanced behavior of oral soft tissue cells without topographical change.

Effect of the ultraviolet light treatment and storage methods on the biological activity of a titanium implant surface.

OBJECTIVE: We evaluated whether the biological activity of the surface of titanium, when stored in an aqueous solution, in low vacuum, and under ambient conditions after ultraviolet light (UV) treatment is comparable to that of the surface immediately after UV treatment for 15min and that after dielectric barrier discharge (DBD) plasma treatment for 15min. METHODS: Grade IV titanium discs with machined surfaces were irradiated with UV and their surface properties were evaluated immediately and after storage for 28days in distilled H2O (dH2O), a vacuum desiccator (31.325kPa), and a sealed container under air. Their surface characteristics were evaluated by atomic force microscopy, X-ray diffraction, contact angle analysis, and X-ray photoelectron spectroscopy. Biological activities were determined by analyzing the albumin adsorption, MC3T3-E1 cell adhesion, and cytoskeleton development. RESULTS: Hydrophilicity of titanium surfaces stored in dH2O was comparable to that immediately after UV treatment and higher than that immediately after DBD plasma treatment (P<0.001). Storage in dH2O and in low vacuum immediately after UV treatment prevented hydrocarbon contamination and maintained elevated amounts of titanium and oxygen. After 28 days, protein adsorption, cellular adhesion, and cytoskeletal development of MC3T3-E1 cells on the titanium surfaces stored in dH2O were significantly enhanced compared to those stored in low vacuum and under ambient conditions while being comparable to those immediately after UV and DBD plasma treatments. SIGNIFICANCE: UV treatment of the titanium implants followed by wet storage is useful for maintaining enhanced biological activity and overcoming biological aging during shelf storage.

Bacterial attachment on titanium surfaces is dependent on topography and chemical changes induced by nonthermal atmospheric pressure plasma.

Here, we investigated the antibacterial effects of chemical changes induced by nonthermal atmospheric pressure plasma (NTAPP) on smooth and rough Ti. The morphologies of smooth and rough surfaces of Ti were examined using scanning electron microscopy (SEM). Both Ti specimens were then treated for 10 min by NTAPP with nitrogen gas. The surface roughness, chemistry, and wettability were examined by optical profilometry, x-ray photoelectron spectroscopy, and water contact angle analysis, respectively. Bacterial attachment was measured by determining the number of colony forming units and by SEM analysis. The rough Ti showed irregular micropits, whereas smooth Ti had a relatively regular pattern on the surface. There were no differences in morphology between samples before and after NTAPP treatment. NTAPP treatment resulted in changes from hydrophobic to hydrophilic properties on rough and smooth Ti; rough Ti showed relatively higher hydrophilicity. Before NTAPP treatment, Streptococcus sanguinis (S. sanguinis) showed greater attachment on rough Ti, and after NTAPP treatment, there was a significant reduction in bacterial attachment. Moreover, the bacterial attachment rate was significantly lower on rough Ti, and the structure of S. sanguinis colonies were significantly changed on NTAPP-treated Ti. NTAPP treatment inhibited bacterial attachment surrounding titanium implants, regardless of surface topography. Therefore, NTAPP treatment on Ti is a next-generation tool for antibacterial applications in the orthopaedic and dental fields.

Overcoming the biological aging of titanium using a wet storage method after ultraviolet treatment.

We evaluated whether the biological activity of the surface of titanium, when stored in an aqueous solution after ultraviolet (UV) treatment, is comparable to that of the surface immediately after UV treatment. We subjected Grade IV titanium discs with machined surfaces to UV radiation for 15 min and then tested them immediately and after storage for 28 days, with and without distilled H2O (dH2O). We evaluated the surface characteristics using surface profiling, contact angle analysis, X-ray photoelectron spectroscopy, and in terms of the surface zeta-potential. We determined the level of biological activity by analysing albumin adsorption, MC3T3-E1 and human mesenchymal cell adhesion and cytoskeleton development, as well as the production of intracellular reactive oxygen species between groups. The surface characteristics produced by the UV irradiation were maintained in dH2O for 28 days. We found that titanium stored in dH2O for 28 days after UV treatment exhibited enhanced protein adsorption, cell attachment, and cytoskeleton development. Titanium stored in dH2O for 28 days after UV irradiation exhibited a lower level of oxidative stress, comparable to that of the titanium immediately after UV treatment. UV treatment combined with wet storage can be used as a means of overcoming the biological aging of titanium.

Non-thermal atmospheric pressure plasma functionalized dental implant for enhancement of bacterial resistance and osseointegration.

OBJECTIVE: Even though roughened titanium (Ti) and Ti alloys have been clinically used as dental implant, they encourage bacterial adhesion, leading to failure of the initial stability. Here, the non-thermal atmospheric pressure plasma jet (NTAPPJ) functionalized Ti and Ti alloy were investigated to promote cellular activities but inhibit the initial attachment of the adherent pioneer bacterium, Streptococcus sanguinis, without topographical changes. METHODS: After the produced radicals from NTAPPJ were characterized, bacterial adhesion to specimens was assessed by PrestoBlue assay and live-dead staining with or without the NTAPPJ functionalizing. After the surface was characterized using optical profilometry, X-ray photoelectron spectroscopy and contact angle analysis, the ions released from the specimens were investigated. In vitro initial cell attachment (4h or 24h) with adhesion images and alkaline phosphatase activity (ALP, 14 days) measurements were performed using rat bone marrow-derived mesenchymal stem cells. RESULTS: The initial bacterial adhesion to the Ti and Ti alloy was significantly inhibited after NTAPPJ functionalizing (p<0.05) compared to those without NTAPPJ functionalizing. The bacterial adhesion-resistance effect was induced by carbon cleaning, which was dependent on the working gas used on the Ti specimens (nitrogen>ammonia and air, p<0.05). The initial cell adhesion with well-developed vinculin localization and consequent ALP activity at 14days to the NTAPPJ-functionalized specimens were superior to the non-treated specimens. SIGNIFICANCE: For the promising success of dental implants, NTAPPJ functionalizing is suggested as a novel surface modification technique; this technique can help ensure the success of integration between the dental implants and bone tissues with less concern of inflammation.

Time-dependent effects of ultraviolet and nonthermal atmospheric pressure plasma on the biological activity of titanium.

Here, we evaluated time-dependent changes in the effects of ultraviolet (UV) and nonthermal atmospheric pressure plasma (NTAPPJ) on the biological activity of titanium compared with that of untreated titanium. Grade IV machined surface titanium discs (12-mm diameter) were used immediately and stored up to 28 days after 15-min UV or 10-min NTAPPJ treatment. Changes of surface characteristics over time were evaluated using scanning electron microscopy, surface profiling, contact angle analysis, X-ray photoelectron spectroscopy, and surface zeta-potential. Changes in biological activity over time were as determined by analysing bovine serum albumin adsorption, MC3T3-E1 early adhesion and morphometry, and alkaline phosphatase (ALP) activity between groups. We found no differences in the effects of treatment on titanium between UV or NTAPPJ over time; both treatments resulted in changes from negatively charged hydrophobic (bioinert) to positively charged hydrophilic (bioactive) surfaces, allowing enhancement of albumin adsorption, osteoblastic cell attachment, and cytoskeleton development. Although this effect may not be prolonged for promotion of cell adhesion until 4 weeks, the effects were sufficient to maintain ALP activity after 7 days of incubation. This positive effect of UV and NTAPPJ treatment can enhance the biological activity of titanium over time.

Long-Term Antibacterial Performance and Bioactivity of Plasma-Engineered Ag-NPs/TiO2

We prepared TiO2 nanotubes (NT) on commercially pure titanium (cp-Ti) substrate by plasma electrolyte oxidation and adapted magnetron sputtering for incorporation of Ag-nanoparticles (Ag-NPs) onto the nanotubes (Ag-NPs/TiO2 nanotube). Power input to the Ag target per unit time was varied (5, 10, 15 W/cm2) to fabricate different shapes of Agnanoparticles onto the nanotubes while net energy input was fixed by maintaining a constant total sputter time (30, 15, 10 s, respectively). For investigation of experimental samples' characteristics, FE-SEM, TEM, EDS, XRD, XPS, SPM analysis and contact angles measurement was carried out. Through these characterization, plasma engineered Ag-NPs was successfully formed on/in the entire nanotube structure. In terms of antibacterial ability, plasma engineered Ag-NPs/TiO2 nanotubes samples significantly reduced S. aureus colony numbers compared with control. Also, simulated body fluid immersion tests with hydroxyapatite showed ion precipitation onto the surface of all experimental groups, confirmed by XRD and EDS analysis. However, plasma engineered Ag-NPs/TiO2 nanotubes groups were not cytotoxic. Furthermore, MC3T3-E1 cells were cultured on Ag-NPs/TiO2 nanotubes groups to evaluate the effect of nanostructured surface on cell functionality such as a cell proliferation and ALP activity. Ag-NPs/TiO2 nanotubes have both biocompatible and antibacterial characteristics.

[Clinical outcome in cases of viral breakthrough during lamivudine therapy in chronic hepatitis B patients].

BACKGROUND/AIMS: Long-term lamivudine therapy can induce the emergence of lamivudine resistant hepatitis B virus (HBV) mutants. Clinically emergence of the mutant is expressed by the reappearance of disappeared HBV DNA in serum. Continued lamivudine treatment has been usually recommended in cases of viral breakthrough. However, the clinical outcome in patients with viral breakthrough is not clear. The aim of this study was to investigate the clinical course of chronic hepatitis B patients after viral breakthrough during lamivudine therapy. METHODS: A total of 74 patients with chronic hepatitis B who showed viral breakthrough after at least 6 months of lamivudine treatment were included in this study. They had positive HBeAg and HBV DNA before treatment. The median follow-up duration after breakthrough was 13 months. RESULTS: After viral breakthrough, only 8 patients (11%) maintained normal ALT levels and 66 patients (89%) showed elevation of ALT. 30 patients (41%) showed acute exacerbation of hepatitis (ALT increase over five-times upper normal limit). These acute exacerbations occurred within three months after breakthrough in 19 patients (63%). In the cases of acute exacerbation, 6 patients showed decompensated progression such as elevation of serum total bilirubin. One of them died of hepatic failure. A predictive factor for acute exacerbation was not found. HBeAg seroconversion occurred in 8 patients after viral breakthrough but their clinical course was highly variable. CONCLUSIONS: Chronic hepatitis B patients who had viral breakthrough during lamivudine therapy should be followed carefully and regularly in mind of potential clinical deterioration. New strategies are needed to manage the cases of acute exacerbation after viral breakthrough.

Endoscopically observed lower esophageal capillary patterns.

BACKGROUND: It has been reported that there are four zones of distinct venous patterns around the gastroesophageal junction (GEJ); i.e. truncal, perforating, palisade (PZ) and gastric zones. Using the distal end of PZ as a marker for GEJ, this study was done to assess the length and patterns of PZ in Koreans, and to assess the prevalence of endoscopic Barrett's esophagus (E-BE) and hiatal hernia (E-HH). METHODS: 847 consecutive patients undergoing diagnostic endoscopy were included. During endoscopy, PZ, squamocolumnar junction (SCJ) and pinchcock action (PCA) were identified. Patterns were classified according to the relationships of the distal end of PZ with SCJ and PCA; A: all three at the same level, B: SCJ proximal to the other two which are at the same level, C: PCA distal to the other two which are at the same level, D: SCJ proximal to the distal end of PZ which is proximal to PCA. Cases with patterns B and D were thought to have E-BE, and those with patterns C and D to have E-HH. RESULTS: Patterns A, B, C and D were 79.2%, 12.1%, 3.8% and 4.9%, respectively. Length of PZ was 3.0 +/- 0.1 cm. E-BE and E-HH were found in 17.0% and 8.7%, respectively. Both E-BE and E-HH were more frequently found in males and in cases with reflux esophagitis. CONCLUSION: E-BE and E-HH are not so infrequent in Koreans as previously thought, if we use the distal end of PZ as an endoscopic marker of GEJ.