臺灣博碩士論文加值系統

本研究，以抗氧化、美白、防曬測定、以及紫外線引起人類正常皮膚纖維母細胞Detroit551傷害的防禦能力等之測定，進行化粧品效能實驗，評估維生素B1、B2、B3、B5、B6、B7、B9、B12等8種維生素B在化粧品的效能。實驗包括: 清除DPPH自由基、螯合亞鐵離子、還原能力、 清除ABTS+自由基、清除超氧陰離子、B16F10細胞中超氧陰離子含量、Dopachrome 生成之抑制率、 B16F10細胞中黑色素含量、 B16F10細胞中酪胺酸酶活性和紫外線吸收測定。
維生素B1於0.05 mg/mL 時能清除84.9%的ABTS+自由基；於10 mg/mL時，能清除88.3 %的DPPH自由基；於30 mg/mL 能清除77.0 %的超氧陰離子；於50 µg/ mL 時，能顯著(p <0.05) 減少B16F10細胞中34.3 %的超氧陰離子含量，並能明顯的增加經紫外線B或紫外線A 照射後的Detroit 551 細胞的存活率。
維生素B3於10 mg/mL 時，能清除76.7 %的超氧陰離子，以及抑制形成黑色素中間產物Dopachrome生成率達96.7 %；於50 µg/ mL 時，能顯著的(p < 0.05) 減少B16F10細胞中25.5 %的超氧陰離子含量，明顯的(p < 0.05) 減少23.7 %的黑色素含量，並顯著的增加經紫外線B或紫外線A 照射後的Detroit 551 細胞的存活率。
維生素B6於0.05 mg/mL 時，能清除92.7 %的ABTS+自由基；於30 mg/mL 時能清除96.3 %的超氧陰離子；於50 µg/ mL 時，能顯著(p < 0.05) 減少B16F10細胞中25.5 %的超氧陰離子含量，並能增加經紫外線B或紫外線A 照射後的Detroit 551 細胞的存活率。
維生素B9 於0.05 mg/mL 時，能清除99.1 %的ABTS+自由基；於1 mg/mL 時能清除62.9 %的超氧陰離子；於50 µg/ mL 時，能顯著(p < 0.05) 減少B16F10細胞中17.0 %的超氧陰離子含量，並能增加經紫外線B或紫外線A 照射後的Detroit 551 細胞的存活率。
綜合綜合以上結果：維生素B1、B3、B6與B9具有良好的抗氧化作用，並具有防禦紫外線A或紫外線B對細胞傷害的作用。此外，維生素B3還有抑制黑色素生成的美白作用。

In the present work, vitamin B family, including B 1, B 2, B 3, B 5, B 6, B 7, B 9, and B 12 , for cosmetic applications were evaluated by antioxidation, whitening UV block determination and protecting tests for UV- induced damage of normal human skin fibroblasts Detroit 551 cells.Experiments include DPPH radical scavenging activity, chelating ferrous ions, Reducing power, ABTS+ radical scavenging activity, superoxide anion scavenging activity, Superoxide anion scavenging activity in B16F10 cells, inhibition of Dopachrome formation , melanin content in B16F10 cells, tyrosinase activity in B16F10 cells and UV absorption.
Vitamin B 1 at 0.05 mg / mL scavenging 84.9% of the ABTS + radicals, at 10 mg / mL scavenging 88.3% of the DPPH free radicals, in 30 mg / mL scavenging 77.0% of the superoxide anions, and at 50 μg / mL shows a significant (p <0.05) inhibition effect to reduce 34.3% of the superoxide anions in the B16F10 cells and to increase the cell viability of Detroit 551 cells damaged by UV-B or UV-A irradiation.
Vitamin B 3 at 10 mg / mL scavenging 76.7% of the superoxide anions, and inhibit 96.7% of the formation of Dopachrome the melanin intermediates, at 50 μg / mL can significantly (p <0.05) reduce 25.5 % of the amount of superoxide anions in the B16F10 cells and reduce 23.7% of the melanin content , and remarkably increase the cell viability of Detroit 551 cells damaged by UV-B or UV-A irradiation.
Vitamin B 6 at 0.05 mg / mL scavenging 92.7% of the ABTS + radicals, at 30 mg / mL scavenging 96.3% of the superoxide anions, and at 50 μg / mL shows a significant (p <0.05) inhibition effect to reduce 25.5% of the superoxide anions in the B16F10 cells and to increase the cell viability of Detroit 551 cells damaged by UV-B or UV-A irradiation.
Vitamin B 9 at 0.05 mg / mL scavenging 99.1% of the ABTS + radicals, at 1 mg / mL scavenging 62.9% of the superoxide anions, and at 50 μg / mL shows a significant (p <0.05) inhibition effect to reduce 17.0% of the superoxide anions in the B16F10 cells and to increase the cell viability of Detroit 551 cells damaged by UV-B or UV-A irradiation.
The results suggest that each of Vitamin B1, B 3, B6 or B9 can be used as a good antioxidant and can protect the cells from the injury by UV light , in addition, Vitamin B3 has a extra effect to inhibit the formation of melanin for whitening.

目錄
中文摘要................................................I
英文摘要................................................III
目 錄................................................VI
表 目 錄 ...............................................XI
圖 目 錄 .............................................. XIII
第 一 章 緒論...........................................1
第一節 八種維生素B的介紹................................4
一、維生素B1.......................................4
二、維生素B2.......................................9
三、維生素B3.......................................14
四、維生素B5.......................................18
五、維生素B6.......................................21
六、維生素B7.......................................23
七、維生素B9.......................................25
八、維生素B12......................................29
第二節 基礎皮膚生理學..................................31
一、皮膚的構造與功能................................31
(一)表皮層......................................31
(二)真皮層......................................32
(三)皮下組織....................................34
二、皮膚與黑色素....................................37
(一)黑色素的生成路徑.............................37
(二)酪胺酸酶....................................38
(三)化粧品美白成分...............................39
(四)美白的機制..................................40
三、皮膚與防曬機制..................................42
(一)皮膚與紫外線.................................42
(二)物理性與化學性防曬劑..........................45
四、皮膚與抗氧化機制.................................46
(一)自由基.......................................46
(二)抗氧化維生素系統..............................49
(三)抗氧化酵素系統................................52
第 二 章 實驗方法.........................................54
第一節 實驗器材.........................................54
一、儀器............................................54
二、實驗材料.........................................56
(一)八種維生素B...................................56
(二)藥品.........................................57
三、細胞株來源.......................................60
四、Detroit 551 細胞株之培養基的配製...................60
五、B16F10細胞株之培養基的配製.........................60
六、Detroit 與 B16F10細胞數之計算.....................61
七、Detroit 與 B16F10 細胞培養與繼代..................61
八、細胞的冷凍保存及解凍...............................62
九、實驗設計與流程....................................63
第二節 實驗方法..........................................65
一、清除DPPH自由基能力之測定...........................65
二、螯合亞鐵離子能力之測定.............................66
三、還原能力之測定....................................68
四、清除ABTS+自由基能力之測定..........................69
五、清除超氧陰離子能力之測定............................71
六、B16F10細胞存活率之測定.............................73
七、B16F10細胞中超氧陰離子之測定........................73
八、Dopachrome生成抑制率之測定.........................74
九、細胞中黑色素含量之測定..............................75
十、細胞中酪胺酸酶活性之測定............................76
十一、紫外線吸收之測定...................................76
十二、Detroit 551細胞存活率之測定........................77
十三、Detroit 551細胞經紫外線B照射後存活率之測定...........77
十四、Detroit 551細胞經紫外線A照射後存活率之測定...........78
十五、統計分析..........................................78
第 三 章 結果..............................................79
一、清除DPPH自由基能力測定.............................79
二、亞鐵離子螯合能力測定...............................79
三、還原能力之測定....................................79
四、清除ABTS+自由基能力測定............................80
五、清除超氧陰離子之測定...............................80
六、B16F10細胞存活率測試..............................81
七、B16F10細胞中超氧自由基測定.........................82
八、Dopachrome生成抑制率測定..........................82
九、細胞中黑色素含量之測定.............................83
十、細胞中酪胺酸酶活性之測定...........................83
十一、紫外線吸收之測定...................................83
十二、Detroit 551細胞存活率測試.........................84
十三、Detroit 551細胞經紫外線B照射後存活率之測定..........85
十四、Detroit 551細胞經紫外線A照射後存活率之測定..........85
第 四 章 討論.............................................87
第 五 章 結論.............................................92
第 六 章 參考文獻..........................................95
第 七 章 附錄.............................................109
表 目 錄
【表2-1】儀器目錄.........................................54
【表2-2】八種維生素B......................................56
【表2-3】藥品目錄.........................................57
【表3-1】維生素B對於DPPH自由基的清除率.....................109
【表3-1-1】維生素C清除DPPH自由基的能力的EC50...............110
【表3-2】維生素B對於螯合亞鐵離子的能力......................111
【表3-3】維生素B的還原能力................................112
【表3-4】維生素B對於ABTS+自由基的清除率....................113
【表3-5】維生素B對於超氧陰離子的清除率......................114
【表3-6】維生素B對於B16F10細胞的存活率.....................115
【表3-7】維生素B對於B16F10細胞中超氧陰離子的生成率..........116
【表3-8-1】維生素B對於Dopachrome生成之抑制率...............117
【表3-8-2】維生素B對於Dopachrome生成之抑制率...............118
【表3-9】維生素B對於B16F10細胞的黑色素含量.................119
【表3-10】維生素B對於B16F10細胞的酪胺酸酶活性..............120
【表3-11】維生素B對於UV-A和UV-B的吸收積分面積..............121
【表3-12】維生素B對於Detroit 551細胞的存活率...............122
【表3-13】維生素B對紫外線B引起Detroit 551細胞傷害的防禦.....123
【表3-14】維生素B對紫外線A引起Detroit 551細胞傷害的防禦.....124
圖 目 錄
【圖1- 1】維生素B1的結構式..................................4
【圖1- 2】Thiamine轉變成TPP的輔酶型式.......................5
【圖1- 3】糖解途徑和檸檬酸循環...............................6
【圖1- 4】維生素B2的結構式..................................9
【圖1- 5】RF, FAD, FMN的結構式.............................10
【圖1- 6】FAD參與脂肪酸之β-氧化作用 .........................11
【圖1- 7】FMN將胺基酸氧化生成α-酮酸 .........................11
【圖1- 8】維生素B3的結構式..................................14
【圖1- 9】Niacin，Nicotiamide 與Tryptophan 轉變成NAD+與.....15
NADP+的輔酶型式
【圖1-10】NADH 與 NAD+互相轉換.............................16
【圖1- 11】維生素B5的結構式.................................18
【圖1-12】泛酸合成輔酶A的途徑...............................19
【圖1-13】吡哆醛、吡哆胺與吡哆醇的結構式......................21
【圖1-14】維生素B7的結構式..................................23
【圖1-15】維生素B9的結構式..................................25
【圖1-16】葉酸與甲基化反應之關係.............................27
【圖1-17】維生素B12的結構式.................................29
【圖1-18】皮膚的構造.......................................36
【圖1-19】黑色素生成路徑圖..................................41
【圖1-20】UVA與UVB的波長...................................43
【圖1-21】二氧化鈦氧化鋅對UV的收波段.........................43
【圖1-22】粒徑大小與不透光的關性.............................44
【圖1-23】體內活性反應與抗氧化劑的互平衡......................46
【圖1-24】Fenton reaction反應..............................51
【圖1-25】體內抗氧化防禦系統................................51
【圖2-1】清除DPPH自由基的原理...............................65
【圖3-1】維生素B對於DPPH自由基的清除率......................125
【圖3-3】維生素B的還原能力.................................125
【圖3-4】維生素B對於AB+S+自由基的清除率.....................126
【圖3-5】維生素B對於超氧陰離子的清除率.......................126
【圖3-6】維生素B對於B16F10細胞的存活率......................127
【圖3-7】維生素B對於B16F10細胞中超氧陰離子的生成率...........127
【圖3-8】維生素B對於 Dopachrome生成之抑制率.................128
【圖3-9】維生素B對於B16F10細胞的黑色素含量..................128
【圖3-10】維生素B對於B16F10細胞的酪胺酸酶活性................129
【圖3-12】維生素B對於Detroit 551細胞的存活率................129
【圖3-13】維生素B對紫外線B引起Detroit 551細胞傷害的防禦......130
【圖3-14】維生素B對紫外線A引起Detroit 551細胞傷害的防禦......130

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