龜鹿二仙膠抗氧化能力與對關節細胞活性的影響

Abstract

龜鹿二仙膠是由龜板和鹿角等重要藥材，再加上枸杞、人參等多種天然成分熬製而成的中藥製劑。它的功效在於強化骨骼和減緩關節退化的症狀，常被民間視為補筋骨的良方。然而，龜鹿二仙膠在市場上的價格差異極大，品質也參差不齊，時常發生以假亂真的情況。為此，我們希望針對龜鹿二仙膠成分中的龜板和鹿角建立中藥指紋圖譜，並且檢視龜板與鹿角溶離胜肽的抗氧化能力以及對於關節滑膜細胞(synoviocyte, HIG-82)的活性，希望藉此確立龜鹿二仙膠的成分品質，並且從龜鹿二仙膠篩選出有效緩解關節退化的成分，提供科學中藥廠家研製成有效緩解關節退化的健康食品或是藥物。我們首先與科學中藥廠家合作，建立傳統中藥龜鹿二仙膠的中藥成分指紋圖譜；然後選取科學中藥廠家提供的龜板與鹿角溶離胜肽成分，利用DPPH抗自由基實驗檢視龜板與鹿角以及溶離後胜肽成分的抗氧化能力；以及採用關節滑膜細胞，利用MTT細胞存活試驗檢視龜板與鹿角溶離胜肽對關節滑膜細胞的生長存活率，藉此篩選出有效緩解關節退化的溶離胜肽成分。本實驗結果顯示：從龜板與鹿角的13種溶離胜肽成分中，發現從鹿角萃取的7種溶離胜肽成分的抗氧化能力較佳，對於關節滑膜細胞的細胞生長存活情形也最佳，至於其他從龜板萃取的6種溶離胜肽成分的抗氧化能力則較差，對於關節滑膜細胞的細胞生長存活情形也較鹿角萃取的7種溶離胜肽來得差。從胺基酸序列的分析發現抗氧化能力佳以及對於關節滑膜細胞的細胞生長有利的溶離胜肽成分，均含有丙胺酸(Alanine，Ala)-絲胺酸(Serine，Ser)-半胱胺酸(Cysteine，Cys)的序列片段。換句話說，從鹿角溶離胜肽對於抗氧化能力以及關節滑膜細胞的細胞生長，都比龜板溶離胜肽來得效果佳。因此，本論文推論從鹿角溶離胜肽應該比較有機會研製成有效緩解關節退化的健康食品或是藥物。

The main medicinal materials of Guilu Erxian Gum (GEG) include tortoise plastron and antlers with adding wolfberry and ginseng to strengthen bone density and slow down the degeneration of joints. In view of the fact that Taiwanese folks often use GEG to replenish their muscles and bones, but the market price varies greatly, the quality is different, and the fake is often reported. To this end, we hope to establish a fingerprint of traditional Chinese medicine for the tortoise plastron and antlers in GEG, and to examine the antioxidant capacity and the activity of synovial cells (synoviocyte, HIG-82) in dissolved peptides from tortoise plastron and antler. We hoped to establish the quality of GEG, and screen out dissolved peptides from GEG that can effectively relieve joint degeneration, and provide traditional Chinese medicine (TCM) manufacturers to develop healthy food or medicine that can effectively relieve joint degeneration. We first cooperated with TCM manufacturers to establish the fingerprints of GEG. Then we selected the dissolved peptides from tortoise plastron and antler that provided by TCM manufacturer, and used the DPPH anti-free radical assay to examine the antioxidant capacity in these dissolved peptides. We used the MTT cell survival assay to examine survival rate of joint synoviocytes in these dissolved peptides. We hoped to screen out the dissolved peptides that can effectively relieve joint degeneration. The results of our experiment showed that among 13 dissolved peptides from tortoise plastron and antlers, Seven dissolved peptides components extracted from antlers have been found that have better antioxidant capacity, and better cell growth and survival rate of joint synoviocytes. As for the other six dissolved peptides extracted from tortoise plastron, the antioxidant capacity is poor, and the cell growth and survival of joint synoviocytes are also worse than dissolved peptides from antlers. From the analysis of the amino acid sequence, it was found that the dissolved peptide components with good antioxidant capacity and beneficial to the cell growth of joint synoviocytes all contain sequence of Alanine (Ala)-Serine (Ser)-Cysteine (Cys). In other words, dissolved peptides from deer antlers have a better effect on antioxidant capacity and cell growth of synoviocytes than dissolved peptides from tortoise plastron. We suggested that dissolved peptides from antlers may have a better chance to develop into healthy food or medicine that can effectively relieve joint degeneration.