以蛋白質體分析反應曲面最適化之牛樟芝腺苷產量

Proteomic Analysis of Response Surface Optimization for Adenosine Production in Antrodia cinnamomea

牛樟芝（Antrodia cinnamomea）是一種特有真菌中草藥，在台灣已被廣泛作為功能性食品。腺苷（adenosine）是牛樟芝中分離出重要生物活性成分之一，並且已經被研究並證實具有抗腫瘤及神經保護活性。然而，在培養基之組成對牛樟芝代謝產生腺苷之影響仍然未知。因此本研究參考反應曲面（RSM）之設計法，探討牛樟芝最適化之代謝產生腺苷的培養基成分。相比對照組，經由反應曲面法的建構，當生長培養基在最適化條件下（3.1%麥芽萃取物，3.1%葡萄糖，4.2%蛋白腖，0.5%黃豆粉，及0.1%MgSO4·7H2O），可增加腺苷產量2.8 倍（12.16 ± 0.42 毫克／克－－菌絲細胞乾重）。其中腺苷代謝產量與蛋白腖含量呈現一次方線性正相關性（P < 0.001），且與培養基中麥芽萃取物、葡萄糖及蛋白腖含量分別呈現二次方曲線迴歸負相關性（P < 0.001）。以蛋白質體學MALDI-TOF/TOF-MS 及MS/MS分析比較對照組及最適化菌絲細胞結果發現，使用最適化培養基會降低腺苷脫氨酶表現量，同時促進磷酸核糖基氨基咪唑－－琥珀酰胺（SAICAR）合成酶及數種核苷酸合成相關蛋白表現量。此研究結果顯示，藉由最適化培養基可以提高腺苷代謝產量並且可能經由抑制腺苷脫氨酶表現，進而活化SAICAR 合成酶及數種核苷酸合成相關蛋白代謝表現量。

Antrodia cinnamomea, a unique Chinese medicinal fungus, has been used as a functionalfood in Taiwan. Adenosine is one of the major bioactive constituents isolated from A. cinnamomea,and it has been reported to have anti-tumor and neuroprotective activities. However, the influenceof growth medium composition on adenosine production in A. cinnamomea remains unknown. Inthis work, the growth medium was optimized referring to the response surface methodology(RSM) to enhance adenosine production from the fermentation of A. cinnamomea. Comparedwith the control, the optimization by RSM resulted in a 2.8-fold increase (reaching 12.16 ± 0.42mg/g dry cell weight) in adenosine production with the optimal growth medium (3.1% maltextract, 3.1% glucose, 4.2% peptone, 0.5% soybean powder, and 0.1% MgSO4·7H2O). Adenosineproduction had a positive linear correlation with peptone level (P < 0.001) and showed a curvilinearnegative relationship with the quadratic effects of the levels of malt extract, glucose, and peptonein the medium (P < 0.001). Based on the proteomic comparison by MALDI-TOF/TOF-MS andMS/MS coupled with decreased expression of adenosine deaminase, the up-regulation ofphosphoribosyl aminoimidazole-succinocarboxamide (SAICAR) synthase and several nucleotidesalvage-related proteins was seen when the optimal medium was used. The results indicated thatenhancement of adenosine production by growth in the optimal medium may potentially suppress expression of adenosine deaminase and activate SAICAR synthase and nucleotide salvage-relatedproteins in nucleotide metabolism.