Use of active extracts to lighten skin, lips, hair and/or nails
A technology of extracts, skins, applied in the field of topical compositions
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Embodiment 1
[0049] The melanosome uptake test (Swollen fern and Limonium) was carried out in the following manner. Confluent cultured B16 melanocytes produce moderate amounts of melanosomes. However, to induce increased melanosome production in this cell line, half-confluent (60%) cultured B16 cells were treated for approximately 36 hours with normal growth medium containing 10 mM ammonium chloride (final concentration). The medium was then aspirated and the hypermelanized cells (2 x 2 ml) washed with distilled water to provide a hypotonic pressure to the cells. An aliquot (2 ml) of hypotonic lysis solution (0.02% NP-40 in water) was added to each plate and incubated for approximately 5 minutes at room temperature. Cell lysates were then examined by light microscopy and cell material from the 3 culture plates was pooled into a 15 ml conical tube and centrifuged (200 xg) for 5 min to remove cell debris. The resulting melanosome-containing supernatant was transferred to a clean 15 ml coni...
Embodiment 2
[0056] In the B16 assay (Limonium), the active ingredient is tested in B16 murine melanoma cells cultured in a monolayer. These cells constitutively produce melanin and are a model system for testing inhibition of melanin synthesis. cells in 5 x 10 3 Cells / well were inserted into 96-well plates and cultured for 24 hours. The original medium was then replaced with fresh medium containing plant extracts. Each active ingredient was added to 6 wells of B16 cells on a 96-well plate for statistical analysis. Cells were treated with medium alone as a negative control, or with the substance to be tested for 7 days. Plates were read at 540 nM to detect melanin formation. The increase in absorbance at 540 nM reflects higher melanin content in the wells.
[0057] At 0.05% (weight / volume), Limonium Limonium showed a 61% reduction in pigmentation compared to the positive control.
Embodiment 3
[0059] exist 14 In the C-dopa incorporation experiment (buta rivet), when 14 Melanogenic activity was determined by measuring the radioactive melanin formed when C-dopa was converted to the acid-insoluble melanin biopolymer in B16F10 melanoma cells. cells in 2 x 10 4 The density of cells / well was inserted into 24-well plates and cultured for 48 hours. Then with plant extracts and 0.2 μCi 14 Fresh medium of C-dopa was used instead of the original medium. Cells were further incubated for 24 hours. After incubation, the medium was discarded and the cells were rinsed with PBS, lysed by adding 0.125 ml 1N NaOH and incubated at 37°C for 30 minutes, then neutralized with 0.025 ml 5N HCl. The resulting cell lysate was transferred to a liquid scintillation vial and mixed with a scintillation mixture, and the radioactivity was measured by a scintillation counter. A portion of the cell lysate was saved and protein content was measured by Lowry's method. Results were normalized to ...
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