Conference Abstract 95

專題演講 

Ecological significance of mycorrhizal symbiosis in wild orchids. ( Dr. Iwase)

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疫病菌 MAPK 與 MEK 基因分子選殖與功能性分析 (顏豪志、黃千育、劉瑞芬)

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利用固態發酵方式開發真菌之保健食品 (李煜玲、郭惠菁、簡韶妤、吳彩平、 毛正倫)

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寄生於豆科植物日本産 3 種 Uromyces 屬銹菌之形態學與分子系統發生學相關 研 究 (鍾文鑫、柿嶌真)

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臺灣靈芝屬之初步調查 (王冬梅、吳聲華)

論文宣讀 

柳杉林大型真菌的調查與監測 (汪碧涵)

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七種野生蘭科植物蘭菌之分離 (鄭淑芬、陳效儀、陳城箖、徐源泰、張喜寧)

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台灣水稻紋枯病遺傳多樣性之探討 (陳思豪、陳隆鐘)

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台灣產巴西 蘑 菇之重金屬含量研究 (石信德.、李宜欣、林彣郁、林俊義、郭鴻裕 、吳寬澤、朱戩良)

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利用 ITS/5.8S rDNA 及 GPD 基因序列，探討尾柄孢殼菌屬和柄孢殼菌屬間的 親緣 關係 (張仲豪、王也珍)

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以紫錐菊為材料，利用微生物發酵萃取 cichoric acid (曾朝豐、吳繼光)

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蘭菌之鑑定及其接種原組成對台灣金線連幼苗生長之影響 (李明治、李國基、張 喜寧)

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蘭菌促進蝴蝶蘭出瓶苗存活率及生長之機制探討 (吳柏宏、張喜寧、黃定鼎)

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朱紅栓菌菌種生物活性探討 (潘威仁、陳啟楨、潘敏雄)

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固態培養樟芝的酒精淬取物會增進 amphotericin B 所造成人類骨肉瘤細胞死 亡 (陳玲儀、廖俊凱、劉苓佑、蘇慶華)

壁報摘要 

不同林型與疏伐度森林中大型真菌的多樣性 (王藝 陳文政 高明脩 林宛柔 汪 碧涵)

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柳杉林中的白色擬枝瑚菌 (洪家卿、張馨尹、谷德予、汪碧涵)

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Halophytophthora 菌屬之形態分類與類緣分析 (詹馥菱、謝松源、袁國芳)

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臺灣黏菌(十九)：刺絲黏菌目 (劉錦惠、張仲豪、陳雅芬)

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Pseudozyma 屬本土菌株之收集與歸群 (魏育慧、劉桂郁、李福臨)

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台灣水稻紋枯病菌菌株間差異性 (陳思豪、楊相哲、林儀真、張建華、陳隆鐘)

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台北地區大氣中真菌孢子特性與分 佈 之探討 (方盈禎、趙馨、詹長權、李崇德)

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建立茄科疫病菌分子診斷技術及探討疫病菌種內和種間之遺傳差異性 (鍾依紋、 劉文禎、潘時玫、安寶貞、陳隆鐘)

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進口木材中長喙殼菌之鑑定 (李涵荺、高孝偉、陳啟予)

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臺灣地區外擔子菌科之研究 (施欣慧、謝煥儒、傅春旭)

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分子診斷技術應用於偵測及定量檢測種媒和土壤傳播真菌 (劉文禎、張裕民、潘 時玫、陳隆鐘)

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兩種台灣產集珠黴屬新種 (何小曼、莊淑錚)

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Sacchachitosan 對於兔子眼角膜遭受鹼腐蝕傷口癒合之研究 (劉曉娟、蘇慶華

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添加中草藥探討樟芝生物轉換之作用 (謝啟弘、張家祥、何偉真、謝昌衛)

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樟芝生長因子之初步研究 (曾朝豐、王朝儀、吳繼光)

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自加馬射線照射之靈芝殘渣製備鹼多醣及其抗氧化特性 (黃婉莉、連佩盈、毛正 倫) 肉絲菇小麥之非揮發性呈味成分 (徐佳莉、簡韶妤、毛正倫)

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固態發酵

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固態 醱 酵樟芝米之抗 氧 化性質 (林欣儀，吳彩平，毛正倫)

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液態發酵繡球菌之胞內與胞外多醣活化巨噬細胞 RAW 264.7 產生抗腫瘤效應 之 研究 (張春生、湯凱宇、陳啟楨、陳健褀)

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利用紅 晶片分析 Monascus pilosus 與 Monascus purpureus 之基因表現 差異 (凃景瑜、黃英娥、廖麗玲)

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紅 麴 大豆甲醇與乙醇萃取物之抗 氧 化性質及其抗 氧 化成分 (李煜玲、毛正倫)

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機能性紅 黑豆之培養 (李惠倫、李煜玲、毛正倫)

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台灣栽培香菇 SCAR 分子標記建立 (蘇秋建、陳隆鐘)

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應用逢機增幅多型性聚合酶連鎖反應反轉錄 mRNA 探討香菇與側耳屬子實體發 育 過程基因表現之差異 (蘇秋建、陳隆鐘)

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白木耳屬菌株液體發酵液應用於化妝保養品之研究 (謝詠筌、陳啟楨)

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低溫及機械刺激對阿魏菇(Pleurotus ferulae)原基誘導之影響 (洪進雄、李欣樺)

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雞 腿菇多醣組成及其對癌細胞之毒性 (蔡淑瑤、毛正倫)

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探討營養對松茸菌絲體生長特性之影響 (楊勝傑、李世豪、廖宇賡)

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不同真菌材料之幾丁聚醣對 痤 瘡丙酸桿菌脂酶活性抑制之探討 (方君仲、蘇慶華)

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利用 IGS-RFLP 分析病原性及非病原性之唐菖蒲萎凋病菌 (紀廷霖、陳瑞祥)

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胞幹抑制劑對 Achlya 菌絲成長、形態及肌動蛋白分 佈 之影響 (陳珮君、于育萍)

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真菌幾丁聚醣抗致突變性之評估 (顏名聰、曾裕琇、毛正倫)

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以納豆菌生產 γ-聚麩胺酸之研究 (李順來、鄭明福、許孟博)

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蜜環菌醱酵物增長戊巴比妥塩誘導小鼠睡眠效果 (李順來、黃雅芬、許孟博)

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美白化妝品資材的開發與研究 (許菱晏、吳繼光)

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Ecological significance of mycorrhizal symbiosis in wild orchids Koji Iwase Fungus/Mushroom Resource and Research Center, Faculty of Agriculture, Tottori University

Orchid is a group of plants, which has specifically evolved characteristics such as very small seeds containing very small amount of energy source (sometimes called as dust seeds), flowers with fantastic morphology, and symbiosis with fungi in root cells. Orchid seeds swell to germinate when the appropriate moisture is available. If the undifferentiated embryo is colonized by suitable and compatible fungi, it could grow to form a clump of cells called protocorm followed by the differentiation of organs to grow into adult plant in nature. Orchid is known to be mycoheterotrophic in germination and protocorm stage in natural habitat because of non-photosynthetic ability. In most photosynthetic orchids, their mycorrhizal fungi have been known to belong to the form-genus Rhizoctonia. On the other hand, other basidiomycete species have been described in many achlorophyllous orchids as the mycorrhizal fungi. Fungal isolation and inoculation experiments have revealed the association of Armillaria spp. in Galeola septentrionalis and in Gastrodia elata, and the association of Erithromyces crocicreas in Galeola altissima. In this study, we conducted to identify the mycorrhizal fungi in acholophyllous species, Epipogium roseum and chlorophyllous species, Cepharanthera falcata. In Epipogium roseum, the several isolates were obtained from peloton, and the DNA analysis

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indicated some of them were classified to gunus Coprinus, and other to genus Psathyrella, both in Coprinaceae. When compatible mycorrhizal fungi is inoculated, seeds could germinate to grow into protocorm, and grew further to form inflorescence under controlled cultivation condition. This is the first study to achieve the whole life cycle from seed to flowering under controlled condition in wild orchid. In a surrounding of the colony of this orchid, many wood logs or fallen leaves are always found, which indicates that those could be used as an energy source for the orchid through the hyphae of mycorrhizal fungi. On the other hand, in C. falcata, the fungi isolated from peloton were clustered with Tomentella in Thelephoraceae or Russula in Russulaceae. Those fungi in Thelephoraceae or Russulaceae are known to from ectomycorrhiza with woody plants. Therefore, in this association among orchid C. falcata, mycorrhizal fungi, and woody plants, tri-partnership symbiosis is suggested.

疫病菌 MAPK 與 MEK 基因分子選殖與功能性分析 顏豪志、 千育、劉瑞芬 國立台灣大學植物病理與微生物學暨研究所

Molecular cloning and functional analysis of MAPKs and MEKs in Phytophthora parasitica-Hao-Zhi Yan, Chien-Yu Huang, and Ruey-Fen Liou (Department of Plant Pathology and Microbiology, National Taiwan University, Taipei). MAPK 訊息傳遞途徑在真菌生理以及病原性上扮演一個重要的角色。然而在卵菌綱方面 有關 MAPK 訊息傳遞途徑的研究仍是不清楚。在本篇研究中，利用簡併式引子對進行聚合酶 連鎖反應並配合 RACE 成功從疫病菌中選殖出 4 個 MAPK 以及 5 個 MEK 基因。分析疫病菌 的 MAPK 與 MEK 胺基酸序列發現，4 個 MAPK 皆具有保守性序列；但在 MEK 方面，除了 其中一個以外，其他皆在被活化區具有保守性。親緣性分析發現疫病菌與植物及真菌的 MAPK 與 MEK 並不屬同ㄧ個分支群。此外，利用即時定量反轉錄聚合酶連鎖反應偵測 MAPK 在疫病菌生長各時期的表現情況以及利用大腸桿菌表現 MAPK 蛋白質所進行功能性 分析的結果將在會中進行討論。 The Mitogen-activated protein kinase (MAPK) pathway, with a prototype of the cascade consisting of MAPK kinase kinase, MAPK kinase (MEK), and MAPK, is known to play essential roles in the signaling of stress response, mating, cell integrity, and pathogenicity in a variety of fungi. Not much is known, however, about the structural characteristics and possible roles of genes

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involved in the MAPK signaling pathway in Oomycetes including Phytophthora. In this study, genes encoding MAPK (ppmk) and MEK (ppmek) in P. parasitica, an important plant pathogen, were cloned by PCR using degenerate primers, followed by 5’- and 3’-RACE. Analysis of the deduced amino acid sequences indicated that ppmks possess the MAPK signature as well as the eleven conserved catalytic subdomains that are typical of Ser/Thr protein kinases and, with the exception of one gene, all ppmeks have the conserved MEK activation site. Phylogenetic analysis indicated that, with both MAPKs and MEKs, genes of P. parasitica form a clade distinct from those of fungi and plants. To further characterize the ppmk genes, their expression in response to a variety of experimental conditions was analyzed by real-time quantitative reverse transcriptasePCR. Besides, recombinant proteins of ppmks were obtained from E. coli and analyzed by in vitro kinase assay. Results obtained from these experiments will be discussed.

利用固態發酵方式開發真菌之保健食品 李煜玲、郭惠菁、簡韶妤、

彩平、毛正倫

國立中興大學食品暨應用生物科技學系

Development of fungi-fermented health food using solid state fermentation--Yu-Ling Lee, Hui-Ching Kuo, Shao-Yu Jian, Tsai-Ping Wu and Jeng-Leun Mau（Department of Food Science and Biotechnology, National Chung-Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan） 近年來，由於人們健康意識的重視，高等真菌菇類成為食物的來源已經從個人的喜好轉 換到其所具有的生理活性。目前有關市面上菇類產品，主要以提供子實體以及菌絲體等原料。 在栽培技術上，有些菇類之子實體本身栽培不容易，而以菌絲體方式提供，如冬蟲夏草 （Cordyceps sinensis）、雞肉絲菇（Termitomyces albuminosus）與樟芝 （Taiwanofungus camphoratus）等，加上近幾年來，稻米等農產品生產過剩，造成農 民生計上受影響。另外，在現代生物製品上，固態發酵的產率比液態發酵高出許多，亦有栽 培上不易受污染、使用萃取溶劑少、發酵殘餘物處理簡單、減少通氣、濃縮與乾燥成本等優點。 因此，本報告主要介紹冬蟲夏草與樟芝以白米作為固態發酵基質，以及雞肉絲菇以小麥作 為固態發酵基質，探討其不同生長條件下，利用麥角固醇含量與pH值變化，選出其最佳固

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態發酵培養條件。 Recently, the use of mushroom as a food source has changed from personal preference to its physiological activities due to the concern of health. Currently, mushrooms are obtained in the form of fruit bodies and mycelia. Some species of fruit bodies are hardly cultivated, such as Cordyceps sinensis, Termitomyces albuminosus and Taiwanofungus camphoratus. In addition, an excess production of rice became a major issue for farmers. Then, many reports showed that solidstate fermentation may offer numerous advantages for the production of microbial chemicals and enzymes. Therefore, this report was to evaluate the optimal conditions of Cordyceps-fermented rice, Taiwanofungus-fermented rice and Termitomyces-fermented wheat as evidenced by their ergosterol contents and pH values.

寄生於豆科植物日本産 3 種 Uromyces 屬銹菌之形態學 與分子系統發生學相關研究 鍾文鑫 1、柿嶌真 2 1

國立中興大學植物病理學系，2 日本筑波大學生命環境科學研究科

Morphological and molecular phylogenetic studies of three Uromyces species on legumes in Japan －Wen-Hsin Chung1, Makoto Kakishima2 (Department of Plant Pathology, National Chung Hsing University, Taiwan，Life and Environmental Sciences, University of Tsukuba, Japan) 寄生於豌豆之 Uromyces viciae-fabae，基於夏孢子壁的厚度與寄主專一性可將其分 成 var. viciae-fabae 與 var. orobi 兩個變種。從日本國内各地所收集到寄生於豆科 Vicia 屬、Lathyrus 屬以及 Pisum 屬之 U. viciae-fabae 乾燥標本 94 份，依夏孢子與冬孢子的 形態學特徵狀，並不能將 U. viciae-fabae 之標本區分成不同的變種。另選取 23 個標本進 行 D1/D2 與 ITS 分子親緣性解析，所有被分析的標本形成單一分子系統。顯示 U. viciae-

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fabae 並不能被分類成不同的變種。此外，在日本 Uromyces appendiculatus 被記載有 三個變種，且主要以夏孢子發芽孔之位置及寄主專一性當作與 U. vignae 區別的分類學特 徵，但由於這兩個種在形態學及生物學的變異太大，常造成混淆。本研究從日本國内各地收 集寄生於 Phaseolus 屬、Vigna 屬、Apios 屬、Lablab 屬以及 Dunbaria 屬之 U. appendiculatus 及 U. vignae 計 225 份之乾燥標本，進行夏孢子及冬孢子之形態學的觀 察。結果顯示，夏孢子發芽孔之位置和冬孢子壁之厚薄為重要之分類特徵，且所觀察之標本 可被區分成三個分類群。自形態學觀察之標本中選取 45 份標本進行分子親緣性的分析，依 D1/D2 與 ITS 核苷酸序列可分別將被分析的標本分成 2 個及 3 個分子系統。本研究中得知 寄主範圍並不與形態學及分子生學吻合，並建議與 ITS 分子系統呈對應關係之形態學標本 應分屬不同的種。 Pea rust fungus, Uromyces viciae-fabae, has been classified into two varieties, var. viciaefabae and var. orobi, based on difference in urediniospore wall thickness and putative host specificity in Japan. Morphological features of urediniospores and teliospores of 94 rust specimens from Vicia, Lathyrus and Pisum did not show definite host specific morphological groups. While 23 Uromyces specimens from Vicia, Lathyrus and Pisum formed a single genetic clade based on D1/D2 and ITS regions. These results suggest that U. viciae-fabae populations on different host plants are not differentiated to groups that can be recognized as varieties. Uromyces appendiculatus, inclusive of three varieties, is distinguished from U. vignae primarily by the position of urediniospore germ pores and putative host specificity. However, opinions over these morphological and physiological features as a taxonomic character have varied greatly and distinction of these species has often been confused. Morphological features of urediniospores and teliospores of 225 rust fungus specimens on species of Phaseolus, Vigna, Apios, Lablab and Dunbaria were examined. The position of germ pores in urediniospores and the teliospore wall thickness were considered as good characters to separate three morphological groups. While 45

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specimens fell into two and three clades based on the nucleotide sequence at D1/D2 and ITS regions, respectively. Neither morphological groups nor molecular clades were host-limited. It is suggested that three morphological groups that corresponded to three distinct ITS clades constitute distinct species.

臺灣靈芝屬之初步調查 王冬梅 1,2 丶吳聲華 1 1

國立自然科學博物館台中自然科學博物館植物學組， 2

中國科學院微生物研究所真菌地衣系統學實驗室

Preliminary survey of Ganoderma in Taiwan--Dong Mei Wang1,2, Sheng Hua Wu1 (1Department of Botany, National Museum of Natural Science, Taichung, Taiwan 404, ROC, 2Systematic Mycology and Lichenology Laboratory, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100080, China)

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靈芝屬是一類多孔菌，以經濟和生態上的重要性著稱。基於文獻調查，臺灣靈芝屬的調 查始於上個世紀初，迄今臺灣已報導 25 個名稱。其中有些名稱已被處理爲靈芝屬其他種的 異名，或已被轉移至其他屬。我們目前正在撰寫臺灣靈芝屬的名錄。在形態研究方面，已觀 察靈芝屬 25 個名稱和部分未鑒定的標本，共計 233 號。在分子實驗方面，已成功測定 79 條 ITS 序列和 21 條 mitSSU 序列。基於形態和分子的研究結果，我們目前暫時承認 19 個 靈芝屬的分類單元，其中臺灣的有 14 個。在這些分類單元中，有些是臺灣的新記錄種，如 G. flexipes Pat. 和 G. multiplicatum (Mont.) Pat.。臺灣稱爲 G. australe (Fr.) Pat.和 G. lucidum (Curtis) P. Karst.的菌物則被證實包括幾個種。 Ganoderma P. Karst., a polypore genus, is well known for its economical and ecological importance. Survey of Taiwanian Ganoderma was initiated in early of the 20th century and 25 names, so far, have been recorded from Taiwan by a literature search. Some names were treated as synonyms, or transferred to other genera. The checklist of Taiwanian Ganoderma is being prepared. In our study, 233 Ganoderma specimens have been subjected to morphological examination, including 25 names except for those unidentified materials. Seventy-nine ITS sequences and 21 mitSSU sequences have been obtained from our study. Currently, 19 Ganoderma taxa including 14 from Taiwan have been tentatively recognized from these specimens based on morphological and molecular data. Among these recognized taxa, some are new to Taiwan, e.g. G. flexipes Pat. and G. multiplicatum (Mont.) Pat.. The taxa generally treated as G. australe (Fr.) Pat. and G. lucidum (Curtis) P. Karst. in Taiwan have been proved to comprise several species.

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論文宣讀摘要

柳杉林大型真菌的調查與監測 汪碧涵 東海大學生命科學系

Investigation and Monitoring of Macrofungal Diversity in a Cryptomeria japonica plantation--PiHan Wang（Department of Life Science, Tunghai University） Pi-Han Wang (Department of Life Science, Tunghai University) 森林生態系是臺灣最重要的生態系之一，其中，14%是大面積單純林相人工造林。為使 人工林得以永續發展經營，有必要實施疏伐作業，以增加杉木人工林結構之異質度和生物 多樣性。95 年起，我們調查與監測巒大事業區人倫林道內 40 年生柳杉人工林中的大型真菌 多樣性，收集疏伐前基本資料：出菇種類以秋季最高，夏季、春季次之，冬季最低；出菇數 量亦然。人工林十二個樣區調查計 392 種 15,846 真菌，以口蘑科、環柄菇科為優勢；天然闊 葉林中含有豐富的蕈菇類，單一樣區調查計 70 種 1,228 個大型真菌，口蘑科為優勢。人工林 中 99% 為腐生菌，1%為共生菌，包括紅絨蓋牛肝菌、乳牛肝菌及混淆松塔牛肝菌；天然林

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中 87%為腐生菌，13%為紅絨蓋牛肝菌及紅菇科共生菌。紅菇科共生菌只在天然林出現。明 年年初將進行疏伐，對照未疏伐林，監測 25% 和 50% 二個不同疏伐程度的人工林棲地中真 菌的變化，期能瞭解不同疏伐程度林相與生態的影響。 Forest ecosystem is one of the most important ecosystems in Taiwan. Among these area, 14% are plantation forests. The new goal is timber production with consideration of its impacts on climate changes, biodiversity losses, ecosystem functions and public acceptance. We propose to monitor and quantify macrofungal community dynamics after 0%, 25% and 50% thinning practices from 2007. We have established 12 one hectare permanent plots in Cryptomerioid japonica plantation forest in central Taiwan. Within this plot, macrofungi are systematically survey and collect as baseline data in 2006. In this year, we collected 392 mushroom species, and 15,846 samples from the plantation plots. Tricholomataceae and Lepiotaceae are dominant in the plantation plots. We collected 70 mushroom species, and 1,228 samples from natural forest plot. Tricholomataceae is dominant in the natural forest plot. There are 99% saprophytic fungi and 1% paragenesis fungi in plantation plots. There are 87% saprophytic fungi and 13% paragenesis fungi in the natural plot.

七種野生蘭科植物蘭菌之分離 鄭淑芬 1、陳效儀 1、陳城箖 2、徐源泰 1、張喜寧 1 1

國立臺灣大學園藝學系，２嘉南藥理科技大學醫務管理系

Orchid mycorrhizal fungi were isolated from the roots of seven wild grown orchids--Shu-Fen Cheng1, Hsiao-Yi Chen1, Jin-Liang Chen2 , Yuan-Tay Shyu1, and C.N. Chang1(1Department of Horticulture, National Taiwan University, ２ Dept. of Hospital and Health Care Administration) 由台灣北、中、南四處地方採集五種原生根節蘭及二種原生鶴頂蘭，進行此等原生蘭花 根部菌類分離，共分離出 104 株代碼真菌菌種，分別再以菌落、菌絲和孢子等傳統形態特徵 鑑定菌種，結果發現以木黴菌（Trichoderma）及毛黴菌(Mucor)屬內之菌株佔優勢，而利用 核酸序列比對之鑑定結果則以炭角菌(Xylaria)屬內之菌株佔多數。其中 57 株尚未鑑定，而有 3 株同時以傳統及核酸序列比對進行鑑定，結果均不相同，需要進一步探討。

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Roots of five species of Calanthe and two species of Phaius from north, midland and south of Taiwan were collected for the isolation of orchid mycorrhizal fungi. Total for 104 isolates of fungi were isolated. They are identified respectively by the traditional method or DNA sequence. Results showed that Trichoderma and Mucor were most often isolated fungi. The DNA sequence results showed that only Xylaris was most common fungus. Fifty seven isolate species are not identified yet and three isolates were identified with both traditional and DNA sequence of cultured pellets at the same time, the result are not corresponding and needed further investigation.

水稻紋枯病菌在台灣之多樣性 陳思豪 1、楊相哲 2、林儀真 3、張建華 4、陳隆鐘 5 1,2,3,4,5

國立中興大學植物病理學系碩士班

The diversity of the rice sheath blight pathogens in Taiwan--Sz-Hau Chen1, Siang-Jhe Yang2, Yi-jen Lin3, C.H. Chang4, Lung-Chung Chen5 (1,2,3,4,5Department of Plant Pathology National Chung Hsing University) 水稻是許多國家的主要糧食。許多病原菌皆會危害水稻並降低其產量，其中以稻熱病、 紋枯病為主要病害。水稻紋枯病是由立枯絲核菌第一融合群亞群(Rhizoctonia solani AG1IA)所引起，紋枯病菌主要發生在高溫高濕氣候的地區，由於台灣依年四季多為高溫高濕氣

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候所以病害發生快速而嚴重。從台灣各地水稻紋枯病病株上所分離到的病原菌中，除了分離 到 Rhizoctonia solani AG1-IA 外還可分離到生小菌核或是淡褐色菌核的立枯絲核菌。此外 在於其生理特性觀察中發現在不同溫度、酸鹼值情形下對其菌核數量、菌核乾重等，發現菌 絲最適生長溫度為 28 度、菌落大小平均為 67.3625mm/day、菌絲生長速率為 1.198mm。 在菌絲融合群(MCGs)試驗方面，依照菌絲間的親和性將台灣水稻紋枯病菌分為四群。此外 利用溫室接水稻台梗九號上所獲得的病害指數在不同區域獲得水稻紋枯病菌之間發現並無 差異以 0 到 5 分等介於 2.8 到 3.3 之間。利用單一引子聚合酵素連鎖反應(Single primer PCR)增幅出的差異性片段經由分析軟體 NTSYS 依照 UPGMA 分群法對各菌株進行差異比 較，發現相似度在 68.8%時便明顯的區分出台灣水稻紋枯病菌第一融合群亞群 (Rhizoctonia solani AG1-IA)。而在相似度達 86%時可將台灣水稻紋枯病菌大致分成五群， 分別以台灣北區(宜蘭縣、桃園縣、新竹市)、中區(台中縣市、南投縣、彰化縣、雲林縣及嘉義市)、 東部地區(花蓮縣、台東縣)以及南部(嘉義縣、台南縣、屏東縣)等，而第五群則因為地理位置 並沒有共同性。近一步的分析差異性的結果則發現其演化樹在 60%的遺傳距離下由五個群 再細分為 11 個族群分別代表不同的區域。分別在為族群 I 為桃園 台中 彰化 雲林 屏東以及 花蓮等縣市，明顯的貫穿台灣西部。而族群 II 則是以高雄為主，族群 IV 為嘉義，族群 V 為 台南 族群 VIII 為桃園，族群 IX 為南投，族群 X 為嘉義和花蓮。而最後一個族群則是傾向以 宜蘭為主。此外以 dsRNA 作為親源性分析的結果則分為八個族群其分別為族群 I 彰化 族群 II 台中 族群 III 屏東 族群 VII 台南 族群 VIII 宜蘭以及桃源、新竹等地。並且以此了解台灣水 稻紋枯病菌菌株間差異主要依地理環境而不同，且環境上的地理區隔對菌株的遺傳差異性 影響遠比水稻品種之因素大。因此了解水稻病害發生之源頭以及病原菌之地域分布性以及其 若獨立機制將有助於管理水稻病害。 Rice is used to be the major agriculture production in lots of country. There are couple pathogens which cause the reduction of yield, especially the rice sheath blight and rice blight. The rice sheath blight is caused by Rhizoctonia solani AG1-IA, and the existing of sheath blight dominantly happened in the area with high temperature and moisture. There fore, the position of Taiwan is the reason why cultivated rice suffered because of the seasons are closed to the weather aforementioned. After isolation of the pathogens collected from each county of Taiwan, and besides the isolation of Rhizoctonia solani AG1-IA, there still have some others with smaller sclerotia or light colored one. Besides, with the inoculation of different isolates collected in Taiwan on rice TaiKing 9 , there seems to be no such difference of disease index surrounded between 2.8 to 3.3 under the circumstance of disease index between 0 and 5.0. Besides, about physiological characteristic test, isolates were cultured at various temperature and pH value, to botanize mycelial growth rate and dry weight of sclerotia, and numbers of sclerotia. The result suggested that the mycelial growth of all isolates was 28 ℃, and the average colony size was 67.3625 mm/day, and then the average mycelial growth rate was 1.198 mm/hr. About mycelial compatible groups (MCGs) test, there were four groups in Taiwan by mycelial compatible and incompatible ration. On the other hand, using single primer PCR to amplify difference fragments and to analysis

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genetic similarity by UPGMA cluster analysis with computer software Gel Compare. Dendrogram based on the analysis of 54 isolates of Rhizoctonia solani AG1-IA, and the result suggested to differentiate Rhizoctonia solani AG1-IA in Taiwan or Japan and other AGs in similarity 68.8%; and further more, isolates of Rhizoctonia solani AG1-IA from Taiwan could differentiate into five groups in similarity 86%. Group north: isolates from Yilan, Hsinchu, Taoyua. Group midland: isolates from Taichung, Hanghua, Nantou, Chiayi, Yunlin. Group east: isolates from Hualien. Group south: Pingtung, Tainan, Chiayi. Group complex: a large breadth group. In deep, the phylogenetic tree was obviously discriminated in to eleven subgroups of the dormant population of R. solani AG1-IA based on the five groups. The subgroups of phylogenetic tree are as follows: Group I includes Taoyuan County, TaiChung City, Changhua County and Yunlin county, Pintung County, HualienCounty. That seems to be a crop production street across the western field of island Taiwan. The Group II is related only with Kaohsiung County. And Group IV is closed to deposit on Chiayi County. The last are that Group V Tainan County and Group VIII is Taoyuan County, Group IX the Naotou County, Group X Chiayi County and Hualien County. The last group XI is prefer to Yilan County. Beside, the phylogenetic tree of the diversity with dsRNAs are constituted by eight subgroups separated to: Group I Changhua County Yunlin County, Chiayi County, Hualien County Group II Taichung County Group III Pingtung County Group VII Tainan County and the Group VIII Yilan County, Taoyuan County and Hsinchu County. We also have the result which is interconnected both with the phylogenetic tree with dsRNA and RAPD, which means the distribution do have specific geographical specification among the dsRNA and the RPAD. The relationship between MCGs and dendrogram based on SSR-PCR analysis showed that diversity of isolates from Taiwan had high degree with geography. This had a great effect upon the diversity of isolates from Taiwan with geography more than with rice variety (host). Therefore under the circumstance with reducing pesticide supplying, to realize the mechanism with virulence and hypovirulence would be a potential plan to manage the sheath blight of rice.

台灣產巴西 蘑 菇之重金屬含量研究 石信德 1.、李宜欣 1、林彣郁 1、林俊義 3、郭鴻裕 2、 寬澤 1、朱戩良 2 1

農委會農業試驗所植物病理組，2 農委會農業試驗所農業化學組， 3

農委會農業試驗所所長

The research of heavy metal content in Agaricus blazei Murrill in Taiwan--Hsin-Der Shih1, IHsin Lee1, Wen-Yuh Lin1, Chien-Yih Lin3, Horng-Yuh Guo2, Kuan-Tzer Wu1 and Chien-Liang Chu2 (1Divison of Plant Pathology, 2Divison of Agricultural Chemistry, 3Director-general, Taiwan Agricultural Research Institute, Wufeng 413, Taiwan)

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巴西蘑菇(Agaricus blazei Murrill；簡稱 ABM)近年來研究發現其 β-D-葡聚糖具有獨 特的抗癌功效，且在臨床上證實其具有提高免疫力、降血脂、降血壓、改善肝機能及動脈硬化 等功效。然而其易蓄積鎘的特性影響品質甚巨。針對台灣巴西蘑菇大型栽培農場於栽培空間、 栽培條件及栽培方法進行調查顯示這些農場主要分佈於中彰投地區，其餘則分散在台南及 高雄地區。目前台灣地區栽培巴西蘑菇方式大多先以木屑太空包培養菌絲後，再予脫袋覆土 刺激出菇。多數菇農利用菇寮配合季節做培養，少數栽培場則利用空調菇舍栽培。大多數菇 農的覆土土壤為就地取材，採集山區、河流或田土，少數使用購置的泥炭土。栽培方法主要 有 3 種，分別為：1.以床栽方式將太空包割包排列於地面或以塑膠框架高後覆土栽培、2.以 太空包或木屑自動瓶上直接覆土出菇及 3.利用箱栽培養。重金屬分析樣本分別為木屑太空包 （含本體、基質木屑、各種輔料）、覆土土壤、灌溉水源及子實體（新鮮或烘乾）等四大部分， 試驗結果顯示巴西蘑菇產量受品種、栽培環境及栽培方式而有很大的影響，尤其遭鎘污染之 覆土層會產生高鎘含量菇體。分析農場使用資材和產出鮮菇或乾菇之重金屬含量顯示有部份 國產巴西蘑菇樣品之鎘含量超過 2 ppm；少數樣品鉛含量超過 3 ppm。將此分析結果和栽 培資材的重金屬含量交叉比對推估不同品種巴西蘑菇子實體對鎘的積聚倍數，發現由最低 的 3.71 倍至最高的 187.38 倍皆有可能。此外，將蒐集而得的巴西蘑菇菇傘與菇柄分別測 定其重金屬含量，結果發現菇傘內大部分重金屬含量皆比菇柄高。本研究期望建立一套質優 安全的巴西蘑菇 GAP 量產體系，使台灣生產的巴西蘑菇不但能符合食品安全衛生標準，還 能應付國內外廣大的銷售市場，進而提高在國際上的競爭力。 It has been reported in recent years that Agaricus blazei Murrill (ABM) has β-D- glucan, which has anticancer effect and was verified in the clinical trials that it is efficient in improving the immunity system, reducing serum lipid, activating the third component in complement system and interferon induction, etc. However, some papers have reported that it is characteristic of ABM to accumulate heavy metals during growth period which influence its quality greatly. ABM could be manufactured into highly potential products for the market place if the heavy metal content of ABMs produced in Taiwan could be lowered. Focusing on major ABM farms in Taiwan, this study investigated cultivation conditions and methods. The farms studied located mostly in Taichung, Nantou, Changhua County in central Taiwan and Tainan, Kaohsiung County in southern Taiwan. Of the seventeen farms investigated, only four utilized air-conditioned growing houses, which produce ABM all year round. Soil used by most farmers to cover the sawdust bags is taken locally from mountain areas, river beds (which contain mostly sand) or crop fields; very few farmers purchase peat moss for the purpose. There are three main cultivation methods; one method places the sawdust bag on the bed and another method places the sawdust bag directly on the ground, while the last method places the sawdust in the tray. The investigation sampled various materials used during the cultivation process, and the materials included compost (fertilizer and raw material), casing mixtures, water, and fruiting body both fresh and dried. The analysis result showed some of the ABM produced domestically contain Cadmimum over 2 ppm, and few ABM

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sampled contain lead over 3 ppm. Based on the finding of this research, we assumed there is a difference of 3.71-187.38 folds in Cadmimum accumulation ability amongst different types of ABMs. The result also showed that the cap of the ABMs contained more Cadmimum concentration than the leg of the ABMs. Further, after boiling the ABM in two separate experiments, one with the whole fruiting body while the other with the fruiting body grounded up into powder, the results showed the Cadmimum in ABM has low solubility. Our data suggested that high Cadmimum concentration casing mixture will surely produce ABMs with high density of Cadmimum. Future research will explore further into the methods of reducing Cadmimum concentration during ABM cultivation. The project is optimistic that a GAP production system of ABM will be established in Taiwan. The production of ABM in Taiwan will not only meet the FDA standard for quality but will also satisfy the quantity required for domestic and international market.

利用 ITS/5.8S rDNA 及 GPD 基因序列， 探討尾柄孢殼菌屬和柄孢殼菌屬間的親緣關係 張仲豪、王也珍 國立自然科學博物館

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Phylogeny of Cercophora and Podospora (Lasiosphaeriaceae) inferred from nuclear internal transcribed spacers and GPD sequences--Jong-How Chang and Yei-Zeng Wang (National Museum of Natural Science, 1 Kuan-Chien Rd. Taichung 404, Taiwan) 尾柄孢殼菌屬(Cercophora)和柄孢殼菌屬(Podospora)都是生長在草食性動物糞便上 的真菌類，在形態分類上，為相當近似的兩個屬，其最主要的差別在於未成熟子囊孢子形 態特徵的不同。尾柄孢殼菌屬在台灣共紀錄有 6 種，而柄孢殼菌屬在台灣共紀錄有 22 種。尾 柄孢殼菌屬主要分類特徵為其未成熟孢子呈透明蜿蜒狀(hyaline sigmoid ascospore)， 與子囊近頂端通常具球狀體 (subapical globule)。柄孢殼菌屬主要特徵為子囊孢子頂端具 有發芽孔、底部有透明的柄狀細胞(pedicel)、以及膠質的附屬構造，所以此二屬可能在親緣 關係上非常接近。另外，有學者依據子囊殼的外部形態與子囊孢子的生長情況，將柄孢殼菌 屬再分出一個裂殼菌屬(Schizothecium)。 本研究是以細胞核 DNA 分子結構來探討尾柄孢殼菌屬和柄孢殼菌屬間的親緣關係。先 以聚合酶鏈結增幅反應(polymerase chain reaction, PCR)的方法，增幅目標菌種的 ITS/5.8S rDNA 及 GPD ( 甘 油 醛 -3- 磷 酸 脫 氫 酶 ； glyceraldehyde-3-phosphate dehydrogenase) 基 因 序 列 。 並 以 ITS/5.8S rDNA 與 GPD 序 列 ， 利 用 最 大 簡 約 法 (maximum parsimony)與貝貽理論(Bayesian Inference)找到最佳的親緣關係樹，以 驗證尾柄孢殼菌屬和柄孢殼菌屬在形態上的分類是否與分子系統一致。 由分析結果顯示，尾柄孢殼菌屬和柄孢殼菌屬皆屬於多系群(polyphyletic)，且在歸 群分枝上，包含許多在形態分類上相近的物種。其中子囊殼毛具膨大細胞的尾柄孢殼菌屬和 柄孢殼菌屬成員在親緣系統上歸為一群，所以我們的結果並不支持將柄孢殼菌屬再另外分 出一個裂殼菌屬。 Coprophilous Pyrenomycetes of Cercophora and Podospora (Lasiosphaeriaceae) are usually grown on dung of herbivores. Six species of Cercophora and twenty-two species of Podospora have been previously reported from Taiwan. The genus Podospora is characterized by ascospores with hyaline pedicel and gelatinous appendages while Cercophora Fuckel has hyaline, cylindrical ascospore with a swollen, pigmented head. Based on the morphology of perithecia and spore development, Podospora was divided into Podospora and Schizothecium by some mycologists. This study focuses on the phylogenetic relationships of Cercophora and Podospora. Crude genomic DNA from each pure strain was isolated. The ribosomal DNA internal transcribed spacers (ITS/5.8S rDNA) and the fragment of glyceraldehyde-3-phosphate dehydrogenase (GPD) sequences were amplified by polymerase chain reaction (PCR). Multiple gene sequences were analyzed using maximum parsimony and Bayesian analyses. In all analyses, Cercophora and Podospora were found to be polyphyletic, consisting of a group of morphologically heterogeneous and phylogenetically distant species. Those species characterized by perithecia adorned with swollen agglutinated hairs are grouped in one clade. Currently, there is insufficient evidence to support the transfer of P. conica group into the genus

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Schizothecium. Thus, we consider Schizothecium as a synonym under Podospora.

以紫錐菊為材料，利用微生物發酵萃取 cichoric acid 曾朝豐、 繼光 亞洲大學生物科技與生物資訊研究所-生科組

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Extraction of cichoric acid-active ingredient of coneflower by microbial fermentation--Chao（ Feng Tseng, Chi-Guang Wu Department of Biotechnology and Bioinformatics, Asia University, Taichung, Taiwan） 微生物發酵並配合以 HPLC 進行成分分析，發現發酵前後之 cichoric acid 含量有明顯 差異。本研究指出經過微生物發酵後，紫錐菊中的 cichoric acid 可伴隨著酒精的產出而被 萃取出來，濃度比發酵前增加三倍。在菌根菌接種實驗中，接種菌根菌的紫錐菊（VA 組） 總乾重比未接種菌根菌的紫錐菊（CK 組）高，實驗中也發現 VA 組葉子 cichoric acid 濃 度為 472.520 μg/mL，CK 組卻只有 245.638μg/mL。在一連串的實驗中，利用菌根菌接 種去提高紫錐菊中的 cichoric acid，再利用微生物發酵的方法，製造出紫錐菊的產品，開 發紫錐菊的應用價值與商業利益。 關鍵字：紫錐菊、菌根菌、cichoric acid In this study, fermentation was applied to extract cichoric acid from coneflower, which was monitored by HPLC analysis. Our data indicates that the concentration of cichoric acid show significant difference pre- and post- fermentation. The concentration of cichoric acid increased three times if compared with the initial background value. In the mycorrhizal inoculation test, the total dry weight of inoculated coneflower (VA) was more than control. The concentration of cichoric acid in the leaves of inoculated coneflower, 472.520 μg/mL was significantly higher than uninoculated control 245.638 μg/mL. In this study, a combination of mycorrhizal inoculation and microbial fermentation strategy to promote the use of coneflower and its commercial value was evaluated. Keyword: Coneflower ( Echinacea sp.), Mycorrhiza, Cichoric acid.

蘭菌之鑑定及其接種原組成對台灣金線連幼苗生長之影響 李明治 1 丶李國基 2 丶張喜寧 1

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1

台灣大學園藝學研究所，2 行政院農委會

Identification of orchid mycorrhizal fungi and it’s inoculum composition effects on the growth of seedlings of Anoectochilus formosanus Hayata--Ming-Chih Lee1, Kuo-Chi Lee2 and Doris C.N. Chang1(1Graduate Institute of Horticulture, Taiwan University, No. 1, Sec. 4, Roosevelt Road, 10617 Taipei, Taiwan and 2Council of Agriculture, 37 Nan-Hai Road, 10014 Taipei, Taiwan) 周(2004)已證實 R02 及 R04 菌株對金線連之生長具促進效果，故利用形態觀察、檢索表 融合群分析及 ITS rDNA 序列比對等方法，對 R01~R09 等 9 種蘭菌進行區分與鑑定，結 果 分 別 為 Rhizoctonia endophytica (R01) 、 Rhizoctonia callae (R02) 、 Fusarium oxysporum (原 R03 改為 F01)、Fusarium solani (原 R08 改為 F02)及 Rhizoctonia solani (R04、R05、R06、R07 及 R09 屬 AG-6 group)，將有助於提供研究及栽培利用時 之參考依據。台灣金線連(台東種)於低溫環境下栽培時，接種蘭菌 R09 (R. solani，多核) 可顯著增加出瓶苗之鮮重，套袋法栽培時則以 R07 (R. solani，多核)菌株具鮮重促進效果。 以 2~3 種蘭菌進行混合接種時，其對於套袋及非套袋栽培金線連之生長影響並不顯著。故 單株蘭菌接種在台灣金線連(台東種)的栽培利用上較具推薦價值，而且以 R09 菌株(即 R. solani, AG-6)對金線連之商業栽培具應用潛能。 Chou(2004) indicated that Rhizoctonia spp. of orchid mycorrhizal fungi (OMF) R02 and R04 could highly-enhance the growth of Anoectochilus formosanus Hayata. Morphology observation, keys identification, hyphal anastomosis reactions and internal transcribed spacer ribosomal DNA(ITS rDNA) were used to identify the OMF isotates R01~R09. Results showed that they were Rhizoctonia endophytica (R01), Rhizoctonia callae (R02), Fusarium oxysporum (R03, now changed to F01)、Fusarium solani (R08 now as F02) and Rhizoctonia solani (R04, R05, R06, R07 and R09 all belonged to AG-6 group), respectively. When A. formosanus Hayata (Taitung cv.) under plastic bag cultivation method(PBCM) and cultivated at low temperature condition, the fresh weights of seedlings were significantly enhanced by the inoculation of OMF isolates R09 and R07 (both were Rhizoctonia solani, multi-nucleate). To use a mixture of two or three Rhizoctonia spp. of solates on the growth of A. formosanus Hayata by both PBCM and traditional cultivation methods. Results showed that growth of seedlings were about the same. Therefore, single isolate inoculation of OMF was recommended for the cultivation of A. formosanus Hayata. It is recommended to use OMF isolate R09 (R. solani, AG-6) as inoculum for the cultivation of A. formosanus Hayata in commercial use.

蘭菌促進蝴蝶蘭出瓶苗存活率及生長之機制探討

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柏宏 1 丶張喜寧 1 丶黃定鼎 2 1

台灣大學園藝，2 成功大學生命科學系

Study of the mechanisms of orchid mycorrhizal fungi on the survival rate and the growth of Phalaenopsis--Po-Hung Wu1, Doris C.N. Chang1 and D.D Huang2(1Graduate Institute of Horticulture, Taiwan University, No. 1, Sec. 4, Roosevelt Road, 10617 Taipei, Taiwan and 2 Department of Life Science, Cheng-Kung University, No1, Ta-Hsueh Road, 701 Tainan City, Taiwan) 先期研究證實，蝴蝶蘭接種蘭菌，可提高數個蝴蝶蘭栽培種之出瓶存活率、營養生長及 開花品質。本研究於蝴蝶蘭瓶苗出瓶時接種蘭菌，經二個月及四個月之栽培後，以抑制性消 減雜交技術(Suppression Subtractive Hybridization，SSH)，比較菌根與非菌根蝴蝶 蘭 根 部 基 因 表 現 之 差 異 。 結 果 得 到 6 個 與 其 他 高 等 植 物 之 cytochrome P450 monooxygenase、GDA2 protein、pectinesterase、PVPR3、glutaredoxin、droughtinduced protein 等抗病、抗逆境蛋白具極高相似度之核酸序列，經 RT-PCR 之方法確認後， 證 實 蝴 蝶 蘭 大 白 花 栽 培 種 KC1111 之 菌 根 植 株 ， 其 cytochrome P450 monooxygenase 、 GDA2 protein 、 pectinesterase 、 PVPR3 及 drought-induced protein 等基因，確實有較高之表現，由此推測，蝴蝶蘭之菌根植株，出瓶存活率提高，可 能是由於這些抗病、抗逆境基因的表現量提高所造成。 It is known from the preliminary study that inoculation of orchid mycorrhizal fungi on Phalaenopsis can elevate the survival rate, vegetable growth and flowering quality of several Phalaenopsis cultivars. In this study, plantlets of Phalaenopsis from the culturing flask were inoculated with orchid mycorrhizal fungi and grown in growth chamber for two- and four- months individually. Then Suppression Subtractive Hybridization (SSH) were used to compare the differential expression of genes between mycorrhiz Phalaenopsis al and nonmycorrhizal roots. It is found that 6 nucleic acid sequences which showed high homology to the gene sequences of cytochrome P450 monooxygenase, GDA2 protein, pectinesterase , PVPR3 , glutaredoxin , and drought-induced protein from other vascular plants. RT-PCR analysis of the transcripts’expression pattern of these pathogenesis-related protein and stress-resistant protein confirmed that all of them were up-regulated in Phalaenopsis cultivar KC1111. It is therefore proposed that inoculation of orchid mycorrhizal fungi in Phalaenopsis could increase the expression of these pathogenesisrelated and stress-resistant genes.

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朱紅栓菌菌種生物活性探討 潘威仁 1、陳 楨 1、潘敏雄 2 南台科技大學生物科技研究所 1、高雄海洋科技大學水產食品科學系 2

Investigation of Bioactivities from culture study of Pycnoporus cinnabarinus -Wei-Ren-Pan1, Chee-Jen-Chen 1, Min-Hsiung-Pan2 (Sounth Taiwan of technology University . Department of Biotechnology. No.1, Nantai St, Yung-Kang City, Tainan and National Kaohsiung Marine University . Department of Sea Food Science. No.142, Haijhuan Rd., Nanzih District, Kaohsiung City 81143, Taiwan ) 本研究是利用朱紅栓菌(Pycnoporus cinnabarinus)菌種先探討其較適生長條件後， 進行實驗室人工栽培；利用收穫的子實體進行生物活性的分析，主要分為抗發炎及細胞凋 亡等活性。在抗發炎方面結果顯示，以酒萃子實體效果較佳，濃度越高抑制 NO 產生能力越 強，80 g/ml 為最大有效濃度；在 iNOS 與 COX-2 的表現上，酒萃子實體 20 g/ml 的濃 度下，即可抑制由 LPS 所誘發的發炎相關基因 iNOS 與 COX-2。在細胞凋亡方面，以子實 體 EtOH、EA 萃取物處理 MCF-7 細胞，顯示依濃度增加時，Sub G1 的量也隨之增加。 In this study. the culture of Pycnoporus cinnabarinus after the investigation of its growing conditions, the laboratory cultivation and shaking flask were carried out and the biological activities analyzed , with extracts of fruit bodies, Divided into anti-inflammatory activity and cell apoptosis. In anti-inflammatory, the best activity has EtOH extracts, inhibits NO activity increased with increasing concentration, best activity concentration is 80µg/ml,in iNOS/COX-2 expression, concentration 20µg/ml could inhibit iNOS/COX-2 expression in RAW264.7 cells stimulated by LPS. Furthermore, the cells apoptosis ,MCF-7 cells treated by EtOH/EA extracts of P. cinnabarinus fruit bodies, the result indicated that Sub G1 of value increased with increasing concentration.

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固態培養樟芝的酒精淬取物會增進amphotericin B所造成人 類骨肉瘤細胞死亡 陳玲儀 1、廖俊凱 12、劉苓佑 1、蘇慶華 3 台北醫學大學醫學研究所， 台北縣新店耕莘醫院家庭醫學科， 臺北醫學大學生物醫學材料研究所及微生物及免疫學科

Ethanol extract of solid state cultivated Taiwanofungus camphoratus extract enhance synergistically cell death of amphotericin B in human osteosarcoma cell line－Ling -Yi Chen1, Chuh-Kai Liao12, Ling-Yo Liu1and Ching-Hua Su3 (1Graduate Institute of Medical Sciences, Taipei Medical University，2 Department of Family Medicine, Cardinal Hospital ,3Graduate Institute of Biomedical Materials and Department of Microbiology and Immunology, Taipei Medical University, 250 Wu-Hsing Street, 110 Taipei, Taiwan) 樟芝是在台灣發現的一種獨特的菌。樟芝的酒精淬取物中含許多活性的化合物。根據以 前的研究，樟芝的酒精淬取物已經在一些實驗過程中有著治療癌細胞的影響。 在許多類型 的癌症中，骨肉瘤難以用化學療法治療，且骨肉瘤所造的病理學機轉也不清楚。 而在我們 的實驗發現固態培養樟芝的酒精淬取物會抑制骨肉瘤，不但如此,固態培養樟芝的酒精淬取 物更會增進 amphotericin B 對於治療癌症的影響。 我們發現在 MG63 細胞中用 100 g/ ml 樟芝的酒精淬取物處理， 24 小時後洗掉，再加入 amphotericin B 3ug/ml， 與單獨 加入固態培養樟芝的酒精淬取物，或者單獨只加 amphotericin B 相比較，發現有增加更 多 apoptosis 細胞且增進細胞毒性效應 。 而且用 100 g/ml 樟芝的酒精淬取物處理， 24 小時後洗掉，再加入 amphotericin B 3ug/ml，會影響在細胞週期中的 G2 期，會增進 p 53，p21，表現，減少 cyclin B1 和 CDC2 和 caspases 的表現。我們的結論發現:固態培 養樟芝的酒精淬取物會增進 amphotericin B 治療骨肉瘤。 Taiwannofungus camphorates (Antrodia camphorate) is a unique fungus found in Taiwan. T. camphoratus contains many active biological compounds which are ethanol souble. According to previous data, ethanol extract from T. camphoratus had reflected anti-tumor effect in some experiments. Among the numerous type of tumors, the osteosarcoma tumors is difficult to be treated by chemotherapy, and the etiology is still remains unclear. The present study is focus on ethanol extract of solid state cultivated (ESSC) T. camphoratus , and their synergistically effect

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with amphotericin B on osteosarcoma cell line as well as title mechanism of inhibition. MG63 osteosarcoma cell lines was used to test the inhibiory concentration of ESSC T. camphoratus, amphotericin B, and their combined effect. We find that the cell line treated with 100μg/ml ESSC T. camphoratus, amphotericin B for 24 hr, washed out with PBS and then added 3μg/ml of AmB for 24 hour induced more apoptosis and potentated the cytotoxic effect in MG63 cell line that is compare with treating of ESSC T. camphorates alone, or amphotericin B alone. Their synergistic effect causes cell cycle block at G2, upregulates p53, p21, levels ,decreases cyclin B1 and CDC2 expression ,and activation of caspases. The result provide evidences that the synergistic effect acts the apoptotic pathway. Taken together, our findings suggested that TC will be effective in the treatment of osteosarcoma tumors . Abbreviations: Taiwannofungus camphoratus ,TC； ethanol extract of solid state cultivated EESC; AmB, amphotericin B

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壁報論文摘要

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不同林型與疏伐度森林中大型真菌的多樣性 王藝 、陳文政、高明脩、林宛柔、汪碧涵 東海大學生命科學系

Macrofungi diversity in different type and thinning degree forests－Yi-Ting Wang, Wan-Cheng Chen, Ming- Hsiu Kao Wan-Rou Lin and Pi-Han Wang(Department of Life Science, Tunghai University) 台灣有近四十二萬公頃的人造林，經營管理對森林生態有決定性的影響，疏伐對生物 多樣性造成的衝擊亟待調查。本研究比較人工林與鄰近天然林的真菌相，並報告不同疏伐程 度對大型蕈菇多樣性的影響。實驗選擇位於大雪山林場約 30 年生的紅檜林與鄰近樟科與殼 斗科為主的天然林，人工紅檜林有未疏伐(1,500 株/公頃)、中度疏伐(1,000 株/公頃)及重 度疏伐(825 株/公頃)等林分密度調整區域，以穿越線法調查記錄樣區內的蕈菇種類，現地 記錄其形態、顏色，並詳細記錄其著生基質及數量。代表樣本採集回實驗室後，根據形態初 步進行科的分類，再根據顯微特徵鑑定至種。五月調查發現 11 科 52 種真菌共 1,168 個子 實體，九月發現 22 科 98 種共 6,732 個子實體，秋季多樣性與族群量皆高於春季，以口蘑 科居多。四個樣區中，天然闊葉林所發現的真菌數量及種類最多，春季 22 種，秋季 55 種 真菌營生方式有木棲腐生、土棲腐生、木棲寄生及土棲共生四類。人工林則均未發現共生菌， 以木棲腐生、土棲腐生及木棲寄生三類為主。疏伐處理可以增加真菌多樣性：重度疏伐林多 樣性高於中度疏伐林，未疏伐人工林族群數量及種類最少。 There are about 420,000 ha plantation forests in Taiwan. Different forest management practices will inevitably affect the forest ecosystem. In this study, we compare the macrofungi diversity of nature and plantation forests, and evaluate the effects of different thinning practices on macrofungi biodiversity. The study site is located in Tai-Siet-San forest park. We established three plots in 30 years old Chamaecyparis formodensis plantation forest, and one plot in the natural forest which are mostly dominated by Fagaceae and Lauraceae. Among the plantation plots, one is located in the non-thinning forest (1,500 tree/ha), another is in the moderate-thinning forest (1,000 tree/ha) and still another in the heavy-thinning forest (825 tree/ha). Fungi were inventoried along 200m transects running across each site. We recorded the morphogens, color, substrates and the number of fruiting body. The sporocarps were identified to species level whenever is possible. In May, 1168 sporocarps were counted, including 11 families, 52 species. In September, 6732 sporocarps were counted, including 22 families, 98 species. Both species diversity and richness in

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the fall were higher than those in spring. Tricholomataceae was dominant in the study site. Among the four sampling plots, natural forest plot always has the highest species diversity and the number of fruiting body, in which 22 species were found in spring and 55 species were found in fall. There are four habitats of fungi, including saprobic on decaying wood, saprobic on soil, parasites on wood and paragenetic on soil. There was no paragenesis fungus occurred in the plantation forest plot. Our results suggested that thinning increased fungi diversity. In non-thinning plantation plot, both population size and number of species are the lower than other plots. In addition, fungal diversity in heavy-thinning plot is higher than those in moderate-thinning plot.

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柳杉林中的白色擬枝瑚菌 洪家卿、張馨尹、谷德予、汪碧涵 東海大學生命科學系

Ramariopsis kunzei, white coral fungi in the Cryptomeria japonica plantations－Chan-Ching Hung, Hsin-Yin Chang, Te-Yu Guu and Pi-Han Wang(Department of Life Science, Tunghai University) 白色擬枝瑚菌，為土棲腐生，子實體乳白色至象牙白色。本研究觀察柳杉林中白色擬 枝瑚菌的生長分布與出菇季節。今年度在南投縣人倫林道的七十八公頃、四十年生柳杉林內 進行調查，每月調查紀錄海拔 1,275 至 1,500 公尺之間的十個一公頃樣區的白色擬枝瑚菌， 發現其主要族群分布集中在四個樣區，兩個樣區有少量出現，另外四個樣區則沒有白色擬 枝瑚菌出菇。白色擬枝瑚菌最適生長的樣區均在海拔高度在 1,400 到 1,470 公尺之間的北 向坡柳杉林。每個樣區的出菇季節不一：第六樣區在五月開始發現白色擬枝瑚菌出菇，六到 九月每月維持近 200 個大量穩定出菇；第七樣區出菇量最大，於六、七月每月超過 200 個， 八月達 345 個，九月減至 159 個；第五樣區則僅在六月出現 20 個，其後未再長出。白色 擬枝 瑚菌 子實體，高 5.0~11.0 公分，主幹分支 2~5 個，柄長 1.0~3.0 公分，柄粗 0.1~0.6 公分，其上 4~11 回二叉分枝，小枝直立，圓柱型，頂端尖銳，子實層生於分枝 表面。擔孢子廣橢圓形，表面具疣，3.96 至 6.93 微米長，1.89 至 3.96 微米寬。正進行族 群遺傳分析。 Ramariopsis kunzei is a wood-saprobe fungi, which varied from ivory to cream white in color. This study surveyed the distribution and fruiting season of Ramariopsis kunzei within a 40 year Cryptomeria japonica plantations forest at Nantou in Taiwan. We surveyed ten 1-ha plots located between1,275m a.s.l. to 1,500m a.s.l. every month in 2006. R. kunzei mainly distributed in four plots, few fruiting bodies were found in two plots, and none in the other four plots. Plots with the presence of R. kunzei are all located at the North-facing slope between altitudes from 1,400 to 1,470. Fruiting season varied between plots: At plot 6, fruiting bodies were abundant in June to August; at plot 7, which the number of fruiting bodies is most abundant, 200 fruiting body were recorded in June and July and the number reached 345 in August, but decreased to 159 in September; at plot 5, only 20 fruiting bodies were found in June. Fruiting bodies of Ramariopsis

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kunzei are up to 5.0~11.0cm tall, 2~5 branches from the main stem. Each branch is 1.0~3.0cm in length and 0.1~0.6cm in width from which 4~11 dichotomous branching. Tip pointed branchlet on the branch upright, on the surface. Basidiospores shaped wide and oval verrucose, 3.96~6.93 in length and 1.89~3.98 in width. Now we are working on polymorphism population genetics.

Halophytophthora 菌屬之形態分類與類緣分析 詹馥菱、謝松源、袁國芳 食品工業發展研究所 生物資源保存及研究中心

Taxonomy and Phylogenetic analysis of Halophytophthora--Fu-Ling Chan, Sung-Yuan Hsieh and Gwo-Fang Yuan (Bioresources Collection and Research Center / Food Industrial Research and Development Institute, Hsinchu, Taiwan.) Halophytophthora 屬是紅樹林中常見的藻菌，多分離自紅樹林的落葉中。由 於 Halophytophthora 進入落葉的速度十分快速，因此被認為是紅樹林生態圈中的首要分解 者。目前 Halophytophthora 有 15 個種和 2 個變種，其分類是依據游動胞囊的形態及游動 孢子的釋放機制。本研究以 28S 核醣體 DNA 序列 D1D2 片段分析，並搭配其游動胞囊形態 與游動孢子之釋放機制研究其類緣關係。研究結果顯示 Halophytophthora 屬應進一步區 分為數個不同的屬。 Halophytophthora species are common zoosporic oomycetes in subtropical and tropical mangrove forest. They were mainly collected from fallen leaves of mangrove, and were thought to be important decomposers in mangrove ecosystem. There are fifteen species and two varieties in the world. Species delimitation in Halophytophthora is based mainly on the morphology of zoosporangium, the presence of vesicle or/and plug, and the mode of zoospore release. Based on 28S large subunit ribosomal RNA gene D1D2 fragment sequence analysis, and the difference in the morphology and structure of zoosporangia, most species except four should move out from Halophytophthora, and several new genus need to be erect for accommodate these fungi.

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臺灣黏菌(十九)：刺絲黏菌目 劉錦惠 1、張仲豪 2、陳雅芬 1 1. 國立臺灣大學植物科學研究所，2. 國立自然科學博物館

Myxomycetes of Taiwan XIX. The Order Echinosteliales--Chin-Hui Liu1, Jong-How Chang2 and Ya-Fen Chen1 (1. Institute of Plant Science, National Taiwan University, 2. National Museum of Natural Science) 本篇報導 6 個刺絲黏菌目的成員，其中 3 種是台灣的新紀錄種，包括頂覆刺絲黏菌 (Echinostelium apitectum Whitney)、樹形刺絲黏菌(E. arboreum Keller & Brooks)、 寡絲刺絲黏菌(E. paucifilum Whitney)，所有子實體都以濕室培養方法獲得。另外內文並 提供台灣所紀錄的刺絲黏菌目內之各科之檢索表，以及碎皮黏菌屬(Clastoderma)與刺絲 黏菌屬(Echinostelium)內物種檢索表。 Six taxa in the order Echinosteliales are reported from Taiwan. Of the six taxa, Echinostelium apitectum Whitney, E. arboreum Keller & Brooks, and E. paucifilum Whitney are newly recorded, which are all collected from moist-chamber cultures. Keys to the families of Echinosteliales, and to the species of Clastoderma and Echinostelium recorded in Taiwan are also provided.

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Pseudozyma 屬本土菌株之收集與歸群 魏育慧、劉桂郁、李福臨 食品工業發展研究所生物資源保存及研究中心

The collection and phylogenic group of genus Psuedozyma in Taiwan－Yu-Hui Wei, Guey-Yuh Liou, Fwu-Ling Lee (Bioresource Collection and Research Center (BCRC), Food Industry Research and Development Institute, Hsinchu 300, Taiwan, R.O.C.) 本研究內容主要為台灣本土之植物及昆蟲表面，類酵母真菌之分離與歸群；收集之菌 株均已寄存於食品工業發展研究所生物資源保存及研究中心。依據 rDNA 中 LSU 片段之 D1, D2 區域序列分析之結果，研究所收集到之菌株隸屬於 Pseudozyma 屬，分類上為擔 子菌門黑穗菌綱之無性世代，是稀有的類酵母真菌。目前 Pseudozyma 屬有 11 個種。 rDNA 序 列 分 析 顯 示 這 些 菌 株 與 Pseudozyma hubeiensis 、 P. tsukubaensis 、 P. aphidis、P. antarctica 等種之 D1, D2 區域序列完全相同；此外，rDNA 序列構築之樹狀 圖顯示，有部分菌株其類緣關係與 P. fusiformata 及 P. shanxiensis 較相近，但菌株與本 屬菌種之序列間，仍有相當程度之差異存在。菌株歸群之結果顯示台灣本土之 Psuedozyma 屬菌株具有豐富的多樣性。 During our study of yeast-like fungi from plants and insects in Taiwan, all strains in this study were collected and deposited in the Bioresource Collection and Research Center (BCRC) of the Food Industry Research and Development Institute. Based on the D1/D2 region of LSU rDNA sequences, these strains were clustered in the clade of unusual yeast-like fungi, genus Pseudozyma. The sequences of the D1/D2 region of LSU rDNA of fifteen strains were identical to the type strains of P. antarctica, P. aphidis, P. hubeiensis, and P. tsukubaensis. However, the phylogenetic

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analysis indicated that five strains were clustered in the clades of P. fusiformata and P. shanxiensis, but the sequences of these domestic strains were quite different from the type strains. Genus Pseudozyma is the anamorph of Ustilaginomycetes and currently contains eleven species. The survey of Pseudozyma species in Taiwan showed the presence of at least six distinct species. The phylogenic analysis shows a great biodiversity of the genus Psuedozyma in Taiwan.

台灣水稻紋枯病菌菌株間差異性 陳思豪 1、楊相哲 2、林儀真 3、張建華 4、陳隆鐘 5 1,2,3,4,5

國立中興大學植物病理學系研究所

Characterization of diversity in Rhizoctonia solani in Taiwan--Sz-Hau Chen1, Siang-Jhe Yang2, Yi-jen Lin3, C.H. Chang4, Lung-Chung Chen5 (1,2,3,4,5Department of Plant Pathology National Chung Hsing University) 水稻紋枯病在台灣一直是重要的病害之一。近 20 年來在本島只有少數對於該病害的研 究報告。對於過去和今日在病害上的差異一直是我們所感興趣的，因此為了解其菌株間差異 性，在於其生理特性觀察中發現在不同溫度、酸鹼值情形等作為差異的分類。在菌絲融合群 (MCGs)試驗方面，依照菌絲間的親和性將台灣水稻紋枯病菌分為四群。此外利用溫室接水 稻台梗九號上所獲得的病害指數在不同區域獲得水稻紋枯病菌之間發現並無差異以 0 到 5 為病害指數之下大致分布於 2.8 到 3.3 之間。與藉由單一引子聚合酵素連鎖反應(Single primer PCR)增幅出的差異性片段經由分析軟體 NTSYS 依照 UPGMA 分群法對各菌株進行 比較，發現其明顯的區分出台灣水稻紋枯病菌第一融合群亞群(Rhizoctonia solani AG1IA)。而在相似度達 86%時可將台灣水稻紋枯病菌大致分成五群，分別以台灣北區(宜蘭縣、 桃園縣、新竹市)、中區(台中縣市、南投縣、彰化縣、雲林縣及嘉義市)、東部地區(花蓮縣、台東 縣)以及南部(嘉義縣、台南縣、屏東縣)等，而第五群則因為地理位置並沒有共同性。近一步的 分析差異性的結果則發現其演化樹在 60%的遺傳距離下由五個群再細分為 11 個族群分別 代表不同的區域。而各族群則明顯的與地域性有相關性，分別以台灣北部縣市以及西部縣市

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等東部等為區分。以 dsRNA 作為親源性分析的結果則分為八個族群其分別為彰化、台中、屏 東、台南、宜蘭以及桃源、新竹等地。並且以此了解台灣水稻紋枯病菌菌株間差異主要依地理 環境而不同，且環境上的地理區隔對菌株的遺傳差異性影響遠比水稻品種之因素大。因此了 解水稻病害發生之源頭以及病原菌之地域分布性以及其若獨立機制將有助於管理水稻病害。 The sheath blight is still an important one of rice epidemic in Taiwan. There are fewer reports this disease during the last 20 years in the island. We are interested in the change of the disease and want to figure out the difference between nowadays and the past. Therefore, in order to realize the difference between isolates, we have physiological characteristic test. The isolates were cultured at various temperature and pH value, to botanize mycelial growth rate and dry weight of sclerotia, and numbers of sclerotia and base on the difference to discriminate the isolates. On anther hand, with mycelial compatible groups (MCGs) test, there are four groups in Taiwan with the difference between mycelial compatible and incompatible ration. Besides, with the inoculation of different isolates collected in Taiwan from rice TaiKing 9 , there seems to be no such difference of disease index surrounded between 2.8 to 3.3 under the circumstance of disease index between 0 and 5.0. The conclusion with this is different in comparison to the former research. On the other hand, by using single primer PCR to amplify difference fragments and to analysis genetic similarity by UPGMA cluster analysis with computer software NTSYS to have Gel Compare. Dendrogram based on the analysis of 54 isolates of Rhizoctonia solani AG1-IA, and the results suggest the difference between Rhizoctonia solani AG1-IA in Taiwan or Japan and other AGs. Further more, isolates of Rhizoctonia solani AG1-IA from Taiwan could differentiate into five groups in similarity 86%. Group north: isolates from Yilan, Hsinchu, Taoyua. Group midland: isolates from Taichung, Hanghua, Nantou, Chiayi, Yunlin. Group east: isolates from Hualien. Group south: Pingtung, Tainan, Chiayi. Group complex: a large breadth group. In deep, the phylogenetic tree was obviously discriminated in to eleven subgroups of the dormant population of R. solani AG1IA based on the five groups. The subgroups of phylogenetic tree are as follows and which includes Taoyuan County, TaiChung City, Changhua County and Yunlin county, Pintung County, HualienCounty. That seems to be a crop production street across the western field of island Taiwan. The Group two is related only with Kaohsiung County. And Group four is closed to deposit on Chiayi County. The last are that Group five Tainan County and Group eight is Taoyuan County, Group night the Naotou County, Group X Chiayi County and Hualien County. The last group eleven is prefer to Yilan County. Beside, the phylogenetic tree of the diversity with dsRNAs are constituted by eight subgroups separated to: Group one Changhua County Yunlin County, Chiayi County, Hualien County Group two Taichung County Group three Pingtung County Group seven Tainan County and the Group eight Yilan County, Taoyuan County and Hsinchu County. We also have the result which is interconnected both with the phylogenetic tree cross eachother, which means the distribution do have specific geographical specification among the dsRNA and the RPAD. The relationship between MCGs and dendrogram based on SSR-PCR analysis showed that

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diversity of isolates from Taiwan had high degree with geography. This had a great effect upon the diversity of isolates from Taiwan with geography more than with rice variety (host). Therefore under the circumstance with reducing pesticide supplying, to realize the mechanism with virulence and hypovirulence would be a potential plan to manage the sheath blight of rice.

台北地區大氣中真菌孢子特性與分 佈 之探討 方盈禎 1、趙馨 1、詹長權 2、李崇德 3 1

臺北醫學公共衛生學系，2 臺灣大學職業醫學與工業衛生研究所， 3

中央大學環境工程研究所

Characteristics and Distributions of Ambient Fungal Spores in the Taipei Area--Ying-Chen Fang1, Hsing Jasmine Chao1, Chang-Chuan Chan2 and Chung-Te Lee3 (1Graduate Institute of Public Health, Taipei Medical University, 250 Wu-Hsing Street, 110 Taipei, Taiwan， 2Institute of Occupational Medicine and Industrial Hygiene, National Taiwan University， 3Graduate Institute of Environmental Engineering, National Central University) 大氣中的生物性微粒無所不在，我們日常生活中隨時都可能暴露到這些生物性微粒， 包括花粉、真菌孢子、細菌、病毒，以及其他源自於動物或植物體的碎片。本研究於 2005 年 1–12 月利用 Burkard 連續性孢子採樣器進行真菌孢子之採樣，以瞭解台北地區大氣中真 菌孢子的種類與濃度分佈，並評估真菌孢子與其他環境因子間的相關性。研究期間總真菌孢 子 平 均 濃 度 為 1930.62 spores/m3 ， 優 勢 菌 種 包 括 ascospores 、 basidiospores 、 Cladosporium 、 unidentified spores、Aspergillus/Penicillium 及 Fusarium。在多變項迴歸模式中發現，溫度是影響真 菌孢子濃度最主要的環境因子，其他如相對濕度、風速、臭氧等也和真菌孢子的濃度有顯著

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相關。未來應進行長期研究以評估大氣過敏原對健康的影響，並瞭解真菌孢子與空氣污染物 及氣象因子間的交互作用。 Biological particulates are ubiquitous in our daily environments, including pollens, fungal spores, bacteria, viruses, or any fragments derived from microorganism, plants and animals. We used a Burkard seven-day recording volumetric spore trap to collect ambient fungal spores from January to December 2005. We evaluated the compositions and distributions of ambient fungi in the Taipei area and examined the relationships between fungi and other environmental parameters. During the study period, the average concentration of total fungal spores was 1930.62 spores/m3, and the predominant fungal taxa were ascospores, basidiospores, Cladosporium, unidentified spores, Aspergillus/Penicillium and Fusarium. In regression models, we found that temperature was the most consistent factor correlated with fungal concentrations. Other environmental parameters, including relative humidity, wind speed, and ozone, also had statistically significant relationships with fungal levels. Future studies should be implemented to evaluate the health impacts of aeroallergens, and to examine the interactions among fungal spores, meteorological factors and air pollutants.

建立茄科疫病菌分子診斷技術 及探討疫病菌種內和種間之遺傳差異性 鍾依紋 1、劉文禎 1、潘時 1

2

、安寶貞 3、陳隆鐘 1

中興大學植物病理系，2 中興大學生命科學系， 3

行政院農業委員會農業試驗所

Establishment of molecular diagnosis for detecting Phytophthora capsici and genetic diversity of Phytophthora inter- and intra-species--Yi-Wen Chung 1, Wen-Chen Liu 1, Shih-Mei Pan2, PaoJhen Ann3, Lung-Chung Chen1(1Department of Plant Pathology, National Chung Hsing University ， 2Department of Life Sciences, National Chung Hsing University ， 3Agricultural Research Institute, Council of Agriculture, Executive Yuan, R.O.C.) 茄科疫病菌為一重要病原真菌，其在許多茄科作物(如辣椒、茄子、蕃茄) 與葫蘆科作物 (如哈密瓜、胡瓜、櫛瓜、南瓜、西瓜)中會引起種腐病及種苗的猝倒、根腐及冠腐、葉斑、莖部病 灶、葉枯和果腐病，而幼苗前猝倒病與後猝倒病亦皆會發生。植株的根部和基部會褪色且受 感染的種苗常會倒伏。冠腐病會在很短時間內使植株全株枯萎死亡。葉斑為黑褐色，直徑 1

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至數英吋。植株藤蔓的任何部位皆可被侵染。莖部病灶為黑褐色、水浸狀且環繞莖部時會引起 莖部枯萎死亡。在潮濕的情況下，有疫病的種苗其受感染的區域會被生長的白色菌絲所覆蓋。 茄科疫病菌，正如其他疫病菌一樣，也會在不同寄主的成熟植株上產生非常多樣化的病徵 ， 且其對不同寄主之病原性具有歧異性現象。目前在台灣所分離到之茄科疫病菌菌株均為 A1 配對型，且尚未發現其具有厚膜孢子，故在自然界中此病原菌是以何種構造或方式殘存仍 屬未知，故分子標記之建立可協助該菌在自然界中之生態研究。因此，本研究以隨機引子增 幅核酸多形性分析篩選茄科疫病菌檢測用分子標記。由供試的引子中篩選出引子 OPB07 可 增幅約 800 bp 的專一性核酸片段；另經南方氏轉漬分析此 800 bp 核酸片段均存在所有 P. capsici 菌株的總量基因體核酸中。將此片段選殖，並經核甘酸定序及設計 3 組 PCR 引子 對，其中以 B07F3/B07R3 引子對所有 P. capsici 菌株經聚合酵素連鎖反應後皆能增幅 381 bp 的核酸片段，但是對於其他 7 種疫病菌( Phytophthora spp. )、4 種腐霉病菌( Pythium spp. )、及健康甜椒、辣椒、番茄、茄子植物組織的基因體核酸皆無法增幅出片段。人 工接種 P. capsici 的甜椒與田間罹病之辣椒植株經初步萃取後的 DNA 亦可由 PCR 反應中 增幅出 381 bp 的專一性片段。另外由台灣各地不同寄主所分離到的茄科疫病菌，其菌落形 態具有玫瑰花瓣狀、星狀、幅射狀、絨毛狀等；而胞囊形態呈橢圓形、梨形或紡錘形，每個胞 囊具有一至二個乳突( papillate )。為瞭解各菌株之遺傳差異，由 18 個茄科疫病菌菌株進 行 RAPD 分析，選取 13 個引子增幅結果應用電腦軟體 NTSYS-pc 依 UPGMA 分群法進行 各菌株遺傳差異之比較，發現茄科疫病菌依不同寄主種類其相似度亦不同，其中由甜椒、辣 椒、蕃茄等植物分離之菌株其相似度較高，相似值為 0.73~0.86；另外由 8 種疫病菌菌株 進行 RAPD 分析，選取 8 個引子增幅結果分析疫病菌種間遺傳親緣差異，由相似值所繪出 之 親 緣 樹 狀 圖 發 現 P. capsici 與 P. cryptogea 其 親 緣 關 係 較 接 近 ， 相 似 值 為 0.18~0.26 ； P. colocasiae 與 P. citrophthora 之 親 緣 關 係 較 接 近 ， 相 似 值 為 0.35~0.39；P. parasitica 與 P. drechsleri 之相似值為 0.18~0.35。綜合以上之結果， 由茄科疫病菌專一性分子標記配合 PCR 技術應用於田間茄科疫病菌的偵測系統建立及利用 RAPD 探討疫病菌之種內種間遺傳差異是否與病原性具有相關性均值得進一步探討研究。 關鍵詞：茄科疫病菌、分子診斷、親緣分析 Phytophthora capsici is an important fungal pathogen that causes seed rot, seedling damping-off, root and crown rot, leaf spots, stem lesions, foliar blight, and fruit rot in many solanaceous crops (pepper, eggplant, tomato) and cucurbits (cantaloupe, cucumber, summer squash, pumpkin, watermelon). Rotting of seedlings prior to emergence (preemergence damping-off) and blighting of recently emerged seedlings (postemergence damping-off) can occur. The roots and plant base may be discolored and infected seedlings often fall over. Crown rot causes the entire plant to collapse and die in a short period of time. Leaf spots are dark brown, one to several centimeters inches in diameter. Vines can be affected at any part. The lesions are dark brown, water-soaked, and girdle the stem, causing the stem to collapse and die. White fungal growth may cover infected areas of blighted seedlings under moist

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conditions. Phytophthora capsici, as well as other Phytophthora spp., can also produce a wide variety of symptoms on mature plants that vary by host. Since the species P. capsici has been broadened, there is considerable diversity in host-specific pathogenicity. The isolates of P. capsici in Taiwan belonged to A1 mating type , and no chlamydospore was found. The structure or survival of P. capsici in the field is unkown. The molecular marker used to understand the pathogen ecological approach in the nature should be established and developed. Therefore, we searched the molecular marker for detection of P. capsici using by RAPD. The primer OPB-07 can amplify a 800 bp species-specific fragment through RAPD and Southern’s hybridization. The species-specific fragment was cloned and designed these PCR primer pairs from 800bp nucleotide sequences. The primer B07F3/B07R3 could amplify a 381 bp fragment to isolates of P. capsici by polymerase chain reaction, but 7 Phytophthora spp., 4 Pythium spp., and health tomato, red pepper, sweet pepper, and eggplant tissues could not amplify any fragment. DNA extracts from pepper after inoculating and infected pepper in the field could be amplified a 381 bp fragment by primer B07F3/B07R3. And the colony morphology of P. capsici isolates from different hosts could be distinguished into unifom, radiate, chrysanthemum, rosette and stellate. The sporangial shapes include ellipsoid, spherical, broadly ovoid, obturbinate, obovoid, fusiform, and pyriform. Each sporangium has one or two papilla. To approach the genetic diversity among isolates of Phytophthora capsici intra-species and Phytophthora inter-species, 13 primer results from 25 primer RAPD profiles with amplification of 18 P. capsici isolates were selected to analysis genetic similarity by UPGMA cluster analysis with computer software NTSYS-pc, and 8 primer results from 25 primer RAPD profiles with amplification of Phytophthora spp. isolates were selected . We found that isolates of P. capsici from different hosts have different genetic diversity. The P. capsici isolates from pepper and tomato have relatively high degree in genetic similarity from 0.73 ~ 0.86. Dendrogram based on the analysis of 27 isolates of P capsici, P. citrophthora, P. parasitica, P. palmivora, P. colocasiae, P. cryptogea, and P. dreschsleri using RAPD showed the relatively higher similarity coefficients for P. capsici and P. cryptogea, ranging from 0.18 ~ 0.26. The high level of similarity between isolates of P. colocasiae and P. citrophthora ranged from 0.35 ~ 0.39, and similarity coefficients of the isolates of P. parasitica and P. dreschsleri ranged from 0.18 ~ 0.35. By the speies-specific molecular marker, we can develope a complete system for detecting P. capsici in the field, and explore the genetic varation of Phytophthora inter-species and intra-species in relation to pathogenicity by RAPD analysis. The establishment of molecular markers and the posses of genetic diversity of Phytophthora capsici intra-species and Phytophthora inter-species are further studied. Keywords：Phytophthora capsici、Molecular diagnosis、Phylogenetic analysis

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進口木材中長喙殼菌之鑑定 李涵荺 1、高孝偉 2、陳 予 1 1

國立中興大學植物病理學系，2國立中興大學生命科學系

Identification of Ophiostoma species in imported wood—H.S. Li1, H.W. Kao2 and C.Y. Chen1 (1Department of Plant Pathology, National Chung-Hsing University ， 2Department of Life Sciences, National Chung-Hsing University) 長喙殼菌普遍存在於林木之中，也常會影響木材之品質。利用MSC選擇性培養基由進口木 材中總共分離獲得88個長喙殼菌菌株，從形態上可以依無性時期之分生孢子將其區分為三 個類群，其中以Ophiostoma piceae complex類群之Ophiostoma quercus最多，進一 步結合了菌株生長測試、配對實驗，以及配合分生之DNA比對技術，鑑定區分出四種，分別 為O. quercus、O. floccosum、O. pluriannulatum、O. abiocarpum，而仍有許多未完成 鑑定之種類待後續進一步探討。

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The fungi of Ophiostoma species commonly occur on woody plants, and usually adversely affect the commercial wood quality. Eighty eight Ophiostoma isolates have been isolated from imported wood by using MSC selective medium. Three groups of species can be morphologically distinguished according to the anamorphic state, among which Ophiostoma quercus in the group of Ophiostoma piceae complex is dominant. The identification is carried out morphologically, and then is further ascertained in combination of biology, mating type studies and molecular data. Consequently four species have been identified. They are O. quercus 、 O. floccosum 、 O. pluriannulatum, and O. abiocarpum. Lots of isolates are still pending to be identified. Further studies are required.

臺灣地區外擔子菌科之研究 施欣慧 1、謝煥儒 1、傅春旭 2 1

國立台灣大學植物病理與微生物學系，2 行政院農委會林業試驗所

Studies of Exobasidiaceae in Taiwan－Hsin-Hui Shih1, Huann-Ju Hsieh1 and Chuen-Hsu Fu2(1 Department of Plant Pathology and Microbiology, National Taiwan University, No. 1, Sec. 4, Roosevelt Road, Taipei, 10617, Taiwan，2 Division of Forest Protection, Taiwan Forest Research Institute, 6 Floor, Room 610, #67 San-Yuan Street, Zhongzheng District, Taipei, 10079, Taiwan) 於台灣地區共採集約一百多份外擔子菌科之標本，其中包含 14 種 Exobasidium spp.，1 種 Muribasidiospora sp.。Exobasidium 屬方面，2 種為新種，分別感染毛蕊木 和台灣馬醉木；E. cylindrosporum、E. pieridis 和 E. yoshinagai 則為新紀錄種。此外，

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於台灣地區首次發現由 Muribasidiospora 屬真菌感染大頭茶的病害，此為一新種。本研究 除形態分類鑑定外，亦分析 30 株 Exobasidium 分離株之 ITS1-5.8S rDNA-ITS2 序列， 其親緣樹狀圖結果顯示，外擔子菌大致可依其寄主分類群而分成三大類群，分別為：杜鵑 花屬、越橘亞科和茶科。 Recently, more than 100 exobasidiaceous specimens were collected, among which were composed of 14 Exobasidium species and 1 Muribasidiaspora. Two Exobasidium species occurred on Vaccinium japonicum var. lasiostemon and Pieridis taiwanensis were new species and three species, E. cylindrosporum on Rhododendron spp., E. pieridis on Lyonia ovalifolia var. ovalifolia and E. yoshinagai on R. mariesii, were newly recorded. Muribasidiospora occurred on Gordonia axillaries was also a new species in Taiwan. Analyzing the ITS1-5.8S rDNA-ITS2 regions of 30 Exobasidium isolates, the phylogenetic tree indicated that Exobasidium species were correlated to their respective hosts and could be grouped into three clades corresponded with host taxonomic groups, Rhododendron, Vaccinioideae and Theaceae.

分子診斷技術應用於偵測及定量檢測種媒和土壤傳播真菌 劉文禎 1、張裕民 1、潘時 1

2

、陳隆鐘 1

中興大學植物病理學系，2 中興大學生命科學系

Application of molecular diagnosis for detection and quantification of seedborne and soilborne fungi--Wen-Chen Liu1, Yu-Min Chang1, Shih-Mei Pan2 and Lung-Chung Chen1(1Department of Plant Pathology, National Chung Hsing University ， 2Department of Life Sciences, National Chung Hsing University) 一個國家或地區特定的疫病害蟲非疫區之認定，除了有利於農產品出口、減少進口之風

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險外，亦可提高農產品之市場競爭力。疫病害蟲非疫區的定義係指經科學方法證明無某種特 定有害生物發生且此狀態由官方加以維持的地區。而非疫區之建立及維持的三個主要階段為 建立非疫區的系統、維持非疫區之植物防疫檢疫措施和確認維持非疫區之查核。由於植物種 子種苗病蟲害檢查制度已是世界各國推動植物防疫檢疫工作極重要的一環，因此 為解決種 子種苗及使用資材之檢疫診斷問題，進而發展及建立更精確之檢疫技術，包括以 PCR 為主 軸之診斷技術(PCR-based methods)、PCR 酵素連結免疫抗體法(PCR--ELISA)之定量技 術及隨機引子核酸增幅多型性分析(RAPD)等技術。首先利用 RAPD 分析所得之 830 bp 特 異 性 核 酸 片 段 以 設 計 專 一 性 引 子 對 ， 及 應 用 PCR 技 術 來 檢 測 種 媒 真 菌 Alternaria brassicicola。結果引子對 Ab536r 及 Ab536f，可針對有病原性之 A. brassicicola 增幅 出 536 bp 的專一性條帶，此專一性引子對靈敏度可達 50 pg μl-1。進一步建立 PCR-ELISA 技術可直接進行增幅產物之定量分析，研究顯示當鏈黴卵白素濃度為 50 μg ml-1，以及探 針濃度為 1 pmol 時，以 DIG 標定於 primer 進行之核酸定量檢測系統有較佳的讀值表現 ， 比使用 DIG 標定於 dNTP 進行之核酸定量檢測系統較為經濟。在針對市售多種植物栽培介質， 如泥炭土、蛭石、珍珠石、水苔及樹皮等進行帶菌之多樣性調查中，目前發現有 Penicillium spp.、Aspergillus spp.、Peridiospora sp.及 Sporotrichum sp.等真菌存在。而在介質帶 菌分離及核酸萃取方面，將介質混合定量孢子後過濾，收集孢子並以加熱法獲得之核酸為 模板及以專一性引子對進行 PCR，可偵測出特異性片段。而在種子種苗方面，萃取其 DNA 及以專一性引子對進行 PCR，亦可偵測出專一性片段。另外參考國內外期刊文獻，收集有關 植物檢疫真菌之分子標記資料庫，包括有 Alternaria brassicae、A. brassiaicola、A. raphani 、 Fusarium oxysporum f. sp. lycopersici 、 Leptosphaeria maculans 、Plasmodiophora brassicae、Sclerotinia sclerotiorum、Rhizoctonia solani、Phytophthora capsici 和 Phytophthora infestans，以供檢疫種子種苗帶有檢 疫真菌分析之用。 關鍵詞：分子診斷、種子檢查、檢疫技術

The determination of a PFA (pest free area) in a country or an area will benefit export of plant products, reduce the risk of import and improve the competitiveness of plant products in markets. A pest free area is “an area in which a specific pest does not occur as demonstrated by scientific evidence and in which, where appropriate, this condition is being officially maintained”. Three main stages are considered in the establishment and subsequent maintenance of a PEA are systems to establish freedom, phytosanitary measures to maintain freedom and checks to verify freedom has been maintained. The requirement for phytosanitary is the quite important part of quarantine in the world ，thus we have developed and established the more accurate techniques, including PCR-based methods, PCR-ELISA for quantification and RAPD, to resolve the problems of phytosanitary. In this study, the 830 bp specific DNA fragment derived from RAPD analysis was used to design A. brassicicola specific primer pairs Ab536R (5 ´-ATATAAAGGCGGGTAACG-3´) and Ab536F (5´-AGCGCCTTATACTCCTTCT-3´). The

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primer pairs could amplify a specific 536-bp fragment from genomic DNA of A. brassicicola by PCR. The sensitivity of the primer pairs could detect 50 pg of genomic DNA of A. brassicicola. Furthermore, the establishment of PCR-ELISA could quantify PCR products directly. Our results showed that when utilizing DIG-labeled primer, 1 pmol probe and 50 μg ml -1 streptavidin, will near the results of utilizing DIG-labeled dNTP, 0.5 pmol probe and 10μg ml -1 streptavidin, but is more economic. For isolation of fungi from media and extraction of DNA, the spores inoculated to media were harvested. The template obtained by heat and specific primers were employed to PCR and a specific DNA fragment was amplified. For seed test, extraction of tissue DNA and specific primers were used to PCR and a specific DNA fragment was detected. Besides, according to the reference from the world, what the molecular marker of seaborne fungi which is including Alternaria brassicae 、 A. brassiaicola 、 A. raphani 、 Fusarium oxysporum f. sp. lycopersici 、 Leptosphaeria maculans 、 Plasmodiophora brassicae 、 Sclerotinia sclerotiorum 、Rhizoctonia solani、Phytophthora capsici and Phytophthora infestans. For the above studies, we can set up the accurate methods for seeds and seedlings quarantine systems. Keywords：Molecular diagnosis, Seed inspection, Phytosanitary technique

兩種台灣產集珠黴屬新種 何小曼 1、莊淑錚 1 國立台北教育大學自然科學教育學系

Two new species of Syncephalis from Taiwan:Syncephalis obliqua ＆ Syncephalis macrospora –Hsiao-Man Ho1, Shu-Cheng Chuang1, Dept. of Science education, National Taipei University of Education, 134. Sect. 2, Heping E. Rd. Taipei.

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本文報告兩種台灣產集珠黴屬新種：Syncephalis obliqua 及 Syncephalis macrospora. S. obliqua 與其他接近種類之不同為孢子囊孢子在囊泡上的生長分佈是斜的， 並具有較大的孢子囊孢子。而 S. macrospora 則與相近種類之最大不同為具有較長孢子囊 柄及較大孢子囊孢子。本文並提供與其他種類之比較與討輪。 Two new species of Syncephalis isolated from Taiwan are described and illustrated： Syncephalis obliqua and Syncephalis macrospora . S. obliqua is distinguished from other species by the asymmetrical placement of the merosporangia on the vesicle and larger cylindrical spores. S. macrospora is distinghished from the other species by longer sporophores and larger spores. A discussion comparing these new species with other taxa is included.

Sacchachitosan 對於兔子眼角膜 遭受鹼腐蝕傷口癒合之研究 劉曉娟 1.、蘇慶華 2 1 2

臺北醫學大學生物醫學材料研究所

臺北醫學大學生物醫學材料研究所及微生物及免疫學科

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Mode of action on SACCHACHITOSAN for Alkali-burned Cornea of Rabbits－Shiao-Chuan Liu1, and Ching-Hua Su2(1Graduate Institute of Biomaterials, Taipei Medical University,2Graduate Institute of Biomedical Materials and Department of Microbiology and Immunology,Taipei Medical University, 250 Wu-Hsing Street, 110 Taipei, Taiwan)

在角膜癒合的過程中，細胞激素跟生長因子都在癒合過程中扮演重要的角色。而傷口癒 合過程包括:發炎反應、細胞增生、細胞遷移、細胞分化、細胞外間質的合成，還有結締組織的 再修復。此篇研究為應用在兔子眼角膜遭受鹼灼傷的傷口癒合藥劑，而藥劑的來源為真菌類 細胞壁以及其他來源。文獻指出 SACCHACHITIN(松山靈芝細胞壁萃取物)可加速傷口癒合， 其功能為促進細胞增生、增加細胞激素以及生長因子的分泌、降低基質金屬蛋白酶的分泌,， 這些功能都對傷口癒合有幫助。SACCHACHITOSAN (deacetylated SACCHACHITIN)為 懸浮性物質，故 SACCHACHITOSAN 被選為來作為新藥劑的材料。在動物實驗之前，先將 SACCHACHITOSAN 加入角膜上皮細胞(SIRC)的培養液中與細胞反應，藉此觀察 SACCHACHITOSAN 與細胞增生以及其最佳劑量的關係。實驗證明，懸浮液濃度 200-300 μg/ml 的 SACCHACHITOSAN 具有促進細胞增生的功能。因此推論 SACCHACHITOSAN 或許可以當做眼角膜傷口癒合的新藥劑。 In response to corneal injury, cytokines and growth factors play a crucial role during the healing and reparative processes. These processes included inflammation, cell proliferation, migration and differentiation, synthesis of the extracellular matrix, and remodeling of connective tissue. The aim of the present study was to develop a new type wound healing drops derived from cell wall of fungi and other sources to meet the demands of alkali burn rabbits’ cornea. SACCHACHITIN extracted from cell wall of Ganoderma tsugae is reported as an accelerator of wound healing, and previous reports indicate that SACCHACHITIN is able to promote wound healing by inducing cell proliferation , increasing the secretion of cell cytokines and growth factors and decreasing matrix metalloproteinase by these cells provided an excellent environment for wound healing. SACCHACHITOSAN, a deacetylated SACCHACHITIN, is water-soluble, thus it is selected for the eye drops of this purpose. Before the animal experiments, SACCHACHITOSAN was estimated for its effects on the proliferation and dose dependent response on rabbit corneal epithelial cell line (Statens Seruminstitut Rabbit Cornea; SIRC). Preliminary results indicated that SACCHACHITOSAN performed as a good mediator for the growth of the cell line in the concentration of 200-300 μg/ml. It was suggested that the existence of SACCHACHITOSAN may help in development of better ocular drug.

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Further experiments on animals are no under investigation.

.

添加中草藥探討樟芝生物轉換之作用 謝 弘 1、張家祥 2、何偉真 1、謝昌衛 1* 1

大葉大學生物資源學系，2 大葉大學生物產業科技學系研究所

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Study on Chinese herbal medicine for bio-diversion in Antrodia cinnamomea－Ch'i -Hung Hsieh1, Chia-Hsiang Chang2, Wei-Chen Ho1, Ch'ang-Wei Hsieh1 (1Department of Bioresources, DA-YEH University，2Department and Graduate Program of Bioindustry Technology, DA-YEH University, 112 Shan-Chiao Street, 510 Chen Lin, Taiwan) 本研究之目的為將多種中草藥應用於樟芝固態培養中，於 30 天內觀察其色澤與生長型 態。菌絲體經過冷凍乾燥後針對其指標成分如三萜類化合物及多醣體於不同時間下進行含量 分析，並利用生物轉換之原理使本身不具藥效或生物活性的物質，經菌種轉換為具有生物 活性的發酵物。在粗三萜方面，靈芝及厚朴有較高之生物轉換率分別可達 3.41 與 2.72 倍； 在多醣含量分析上，紫蘇有較高之生物轉換率可達 2.2 倍。因此，如何使樟芝於人工培養中 能有效地產生高活性之產物將是我們的目標。 The objectives of this research were to study different kind of Chinese herbal medicine in solid culture of Antrodia cinnamomea that observed the tinctures and acervate conditions in 30 days. After freezing and desiccation, we aimed at index composition of mycelium such as triterpenoids and polysaccharides analyzed the contents in different times. Through mushroom, used the principle of bio diversion that the substance have none curative effect or bio-activity can transform leavening having bio-activuty. In triterpenoids, Ganoderma Lucidum and Magnolia officinalis have much more bio-diversion that achieved 3.41 and 2.72 time respectively. In polasaccharide, perilla have more bio-diversion that rechieved 2.2 time. Therefore, how to make Antrodia cinnamomea can account high-activity products in artificial culture efficiently are our destination.

樟芝生長因子之初步研究 曾朝豐、王朝儀、 繼光

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亞洲大學生物科技與生物資訊研究所-生科組

A preliminary study on the hyphal growth of Antrodia camphorata-- Chao-Feng Tseng, ChaoEi Wang and Chi-Guang Wu (Department of Biotechnology and Bioinformatics, Asia University, Taichung, Taiwan). 本研究利用不同固態培養基培養樟芝（Antrodia camphorata）。經由不同培養基配方 進行比較，發現利用樟木滅菌液配置之固態培養基，其樟芝菌絲生長較一般培養基良好， 利用不同氮原（酵母抽出物和麥芽抽出物）製備培養基，發現酵母抽出物製備之培養基菌 絲生長較慢，且長出之菌絲為白色菌絲。單用樟木滅菌液製備培養基（2% agar），其菌絲 生長最佳，但菌絲生長卻不密集。利用 HPLC 分析發現，樟芝菌絲萃取液與樟芝子實體萃取 液有相似的成分。 關鍵字：樟芝，生長因子，高效能液相層析 In this study, different composition of culture media was designed to cultivate Antrodia camphorata. The results indicated that wood extract of A. camphorata could enhance the hyphal growth, better than other nutrient amendments. Yeast extract amended medium did not stimulate hyphal growth, and the mycelial color was pale whiteand. In contrast, wood extract showed the best hyphal growth if compared with other treatments. Through the HPLC analysis, the composition of hyphal extract was closely similar to that from fruiting body. Keyword: Antrodia camphorata, growth factor, HPLC

自加馬射線照射之靈芝殘渣製備鹼多醣及其抗氧化特性

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婉莉、連佩盈、毛正倫 國立中興大學食品暨應用生物科技學系

Preparation of the alkali-polysaccharide extracted from γ-irradiated Ganoderma tsugae residue and its antioxidant properties--Wan-Lih Huang,Pei-Ying Lian and Jeng-Leun Mau (Department of Food Science and Biotechnology, National Chung-Hsing University,Taichung) 目前市面上經兩次熱水萃取後所剩之松杉靈芝（Ganoderma tsugae Murrill）殘渣 仍無任何經濟價值與開發。研究顯示加馬線照射可規律的降解靈芝渣，因此本研究目的為利 用照射加馬線 1 kGy 劑量於松杉靈芝殘渣，並萃取其照射後鹼多醣，與未照射靈芝渣之鹼 萃多醣進行抗氧化性質探討。結果顯示，在抗氧化力方面，經照射之靈芝鹼多醣，濃度在 5 mg/ml 時，抗氧化力即可達 61.54%。在還原力方面，經照射之靈芝鹼多醣，隨著濃度提 高而增加。而在清除 1,1-二苯基-2-苦味肼基團能力方面，經照射之靈芝鹼多醣於低濃度 0.5mg/ml（43.48%）較未照射靈芝鹼多醣（39.96%）清除效果佳。清除羥自由基能力 方面，在濃度 5-10mg/ml（16.41-39.75%）較未照射（0-2.26%）清除能力佳。而在螯 合亞鐵離子方面，經照射之靈芝鹼多醣在濃度 0.5-20mg/ml（68.50-100%）皆比未經照 射（55.62-98.94%）高，且隨著濃度提高而增加。結果顯示，加馬線照射對靈芝殘渣具有 破壞效果，可有效降解殘渣而提升其抗氧化效果。 The residue of Ganoderma tsugae Murrill, which has been extracted twice by hot water, has no economical value and exploitation. Besides, some research indicated gamma irradiation might degrade the residue and yield polysaccharide with reduced size. Therefore, the research was designed to apply 1 kGy gamma ray irradiation on the residues of G. tsugae in order to explore the utilization of this residue. The alkali- polysaccharide with 1 kGy γ-irradiation was compared with the control for its antioxidant properties. The alkali- polysaccharides with 1 kGy irradiation dose showed the best antioxidant activity (61.54%) at 5 mg/ml. The alkali-polysaccharides with irradiation increased its reducing power with concentration. The alkali-polysaccharide with 1 kGy γ-irradiation (43.48%) had higher scavenging ability of alkali-polysaccharides on DPPH radical at low concentration (0.5 mg/ml) than with the control (39.96%). The alkali-polysaccharide with 1 kGy γ-irradiation (16.41-39.75%) had higher scavenging ability of alkali-polysaccharides on OH radicals (5-10 mg/ml) than with the control (0-2.26%). The alkali-polysaccharide with 1 kGy γirradiation (68.50-100%) had higher chelating ability on ferrous ions (0.5-20 mg/ml) than with the control (55.62-98.94%) and increased with concentration.

固態發酵

肉絲菇小麥之非揮發性呈味成分 50

徐佳莉、簡韶妤、毛正倫 國立中興大學食品暨應用生物科技學系

Non-volatile taste components of solid state fermented termite fungal wheat--Jia-Li Syu, Shao-Yu Jia, Jeng-Leun Mau (Department of Food Science and Biotechnology, National Chung-Hsing University, 250 Kuokung Road, Taichung 40227, Taiwan, ROC) 本研究以固態發酵方式將雞肉絲菇菌種接種於小麥上培養 14 天，所得之產品（雞肉絲菇 小麥）經凍乾後，分析其非揮發性呈味成分，並與其菌絲體、濾液及小麥作比較。結果顯示， 在脂肪酸組成上，以亞麻油酸為主，其中以菌絲體之含量（73.67％）為最高。在可溶性糖 含量上，以濾液（320.73 mg/g）含量最高。此外，菌絲體所含的游離胺基酸以精胺酸及 麩胺酸為主，而游離胺基酸總量，以菌絲體（28.49 mg/g）含量最高。然而，在呈味特性 MSG-like 含量上以菌絲體、濾液、雞肉絲菇小麥與小麥的含量分別為 3.41、6.06、1.63 與 3.51 mg/g。在核苷酸方面，以菌絲體（9.71 mg/g）含量最高。除此之外，等價鮮味值上 以濾液（243.21 g/100g）與菌絲體（172.29 g/100g）皆高於雞肉絲菇小麥（ 0.93 g/100g）與小麥（3.30 g/100g）。綜合上述的結果，經固態發酵所得的雞肉絲菇小麥之 呈味成分含量並沒有較小麥所含者高，可能是由於雞肉絲菇菌絲於小麥基質中生長不足所 導致的，因此往後研究可再探討雞肉絲菇小麥的最適培養條件。 The research was to inoculate the mycelia of Termitomyces albuminosus into autoclaved polished wheat and incubate for 14 days to produce a product called termite fungal wheat. The product was freeze dried and its non-volatile components were analysed and compared to mycelia, filtrate and wheat. With regards to fatty acid component, linoleic acid was the major fatty acid, and the mycelia were the highest (73.67%). The content of soluble sugars of filtrate was the highest (320.73 mg/g). Furthermore, the major free amino acids of mycelia were arginine and glutamate. The content of total free amino acids for mycelia was the highest (28.49 mg/g). With regards to monosodium glutamate-like components, the contents of mycelia, filtrate, termite fungal wheat and wheat were 3.41, 6.06, 1.63 and 3.51 mg/g, respectively. The content of 5´-nucleotide was the highest in mycelia (9.71 mg/g). Besides, the equivalent umami concentration of filtrate (243.21 g/100 g) was higher than those of mycelia (172.29 g/100 g), wheat (3.30 g/100 g) and termite fungal wheat (0.93 g/100 g). Overall, the taste components of termite fungal wheat obtained from solid state fermentation was much less than those in wheat. This might be due to the inadequate growth of termite fungal in wheat. Therefore, the optimal condition for T. albuminosus to grow in wheat could be studied further.

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固態發酵樟芝米之抗 氧 化性質 林欣儀， 彩平，毛正倫 國立中興大學食品暨應用生物科技系

Antioxidant properties of solid-state fermentated Chang-chin rice--Shin-Yi Lin, Tsai-Ping Wu, Jeng-Leun Mau (Department of Food Science and Biotechnology, National Chung-Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan, ROC) 樟芝（Taiwanofungus camphorates Wu et al.）為一種珍稀的食藥用菇，含有許 多生理活性成份，本研究以胚芽米為固態發酵之基質，將樟芝接種於其上培養 24 天，所得 之樟芝米於凍乾後針對其抗氧化活性進行評估，並與其菌絲體及胚芽米互相比較。結果顯示 在熱水萃取方面，樟芝菌絲體萃取物之還原力則比樟芝米較好。在清除 1,1-二苯基-2-苦味 肼基團上樟芝米萃取物濃度 20 mg/ml 時其能力為 88.42％，比同濃度之抗壞血酸高。在螯 合亞鐵離子之能力上，樟芝米濃度達 20 mg/ml 時，其螯合能力為 100％。另外乙醇萃取 方面所得之萃取物在濃度為 20 mg/ml 時，三者的抗氧化能力達到最高。在還原力方面，樟 芝米都較其它兩者為高。樟芝米在清除 1,1-二苯基-2-苦味肼基團上，其濃度於 1 mg/ml 時， 即有高於 50％的清除能力。由此可知，胚芽米經過固態發酵為樟芝米後，可提高樟芝米之 抗氧化性質。 Taiwanofungus camphorates Wu et al. (Chang-chih or Niu-chang-ku) is a precious edible and medicinal mushroom due to the fact that it contains many physiologically active substances. The objectives of this study were to prepare Chang-chih rice using embryo rice as a substate, Changchih was initially grown on the rice for 24 days, and then freeze-dry and analyze its antioxidant properties and compare with mycelia, embryo rice and some commonly used antioxidants. With regard to reducing power of hot water extracts, the mycelia better than Chang-chih rice. At 20 mg/ml, scavenging ability of Chang-chih rice on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals were 88.42％, better than ascorbic acid that of at the same concentration. Chang-chih rice chelated ferrous ions by 100% at 20 mg/ml. All of ethanolic extracts had the highest antioxidant activity at 20 mg/ml. For reducing power, Chang-chih rice was higher than others. At 1 mg/ml, scavenging ability of Chang-chih rice on DPPH radicals were higher than 50%. Bsae on the results obtained, the Chang-chih rice prepared from the solid state fermentation of embryo rice possessed better antioxidant properties.

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液態發酵繡球菌之胞內與胞外多醣 活化巨噬細胞 RAW 264.7 產生抗腫瘤效應之研究 張春生、湯凱宇、陳

楨、陳健褀

南台科技大學生物科技研究所 The anti-tumor effect of polysaccharides and exopolysaccharides of Sparassis crispa under submerged fermentation from activated RAW 264.7 macrophages Chun-Sheng Chang, KaiYuer Tang, Chee-Jen Chen, Jian-Chyi-Chen (Southern Taiwan University of Technology. Department of Biotechnology. 1, Nan-Tai St, Yung-Kang City, Tainan ) 菇蕈類所含之多醣體具有許多功效，如降血糖、抗氧化及抗腫瘤等功效。由於繡球菌 (Sparassis crispa)子實體所含多醣，對於老鼠白血球症有強烈血管擴張與出血反應，患 有黏膜白斑病老鼠的造血反應於 cyclophosphamide 也有所提升。在 DBA/1 及 DBA/2 老 鼠也可明顯提昇 IgM 免疫球蛋白的數量，在子實體取得不易的情況下，本論文利用液態發 酵培養繡球菌之胞內與胞外多醣刺激巨噬細胞 RAW264.7 產生抑制腫瘤生長之激素，評估 其抗腫瘤效應。篩選六種不同的碳源，發現以玉米漿澱粉(Corn Starch)為碳源時，可得最 高菌體量為：11.98 g/L。在碳源為葡萄糖 (Glucose)時，可得最多胞內多醣為：117.71 mg/g。在碳源為蔗糖(Saccharose)時胞外多醣為最高：40.17 mg/g。在細胞功能性評估 方面，將菌絲體依序進行熱水萃取、冷鹼萃取及熱鹼萃取並進行透析後，分別取得 F-Ⅰ、F-Ⅱ 及 F-Ⅲ 三種多醣樣品，取相同多醣濃度刺激免疫細胞(Raw 264.7)後，取其上清液分別加 入癌細胞株，以碳源為果糖(Fructose)所獲得之 F-1，對於乳癌細胞(MCF-7)之細胞存活率 有最低值(50.16% ± 1.87%)； F-Ⅱ 及 FIII 皆以碳源為玉米漿澱粉時，有最低細胞存活率 分別為 39.99% ± 2.62 %，58.49 % ± 2.25%。對於肝癌細胞(HEP G2)無論胞外多醣、 F-Ⅰ、F-Ⅱ 或 F-Ⅲ 皆無明顯抑制率。從實驗結果顯示， 繡 球菌之胞內多醣能藉由刺激 RAW 264.7 來抑制 MCF-7 癌細胞的生長。 The experiments were to investigate the anti-tumor effect of polysaccharides and exopolysaccharides of Sparassis crispa under submerged fermentation from activated RAW 264.7 macrophages. Screening the six different carbon sources, the corn starch as the carbon source can harvest the highest concentration of biomass, 11.984 g/L. When glucose used as the carbon source,

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the highest production of polysaccharide per gram of mycelium was obtained at 117.71 mg/g. The highest production of exopolysaccharide per gram mycelium was 40.17 mg/g which was used saccharose as the carbon source. Polysaccharide fraction (F-I) was extracted the air-dried and powdered S. crispa with hot water. The resulting residue was then extracted with cold alkali to obtain the second polysaccharide fraction (F-II), and then extracted with hot alkali to obtain the third polysaccharide fraction (F-III). Those polysaccharide fractions were collected after extensive dialysis. To test the anti-tumor activity of mediator released from the macrophage Raw264.7 were stimulated by the polysaccharide fractions, respectively. The F-I cultured from the fructose as carbon source that showed the lowest survival ratio of MCF-7 cell line was 50.16% ± 1.87%. Both of the F-II and F-III were cultured from the corn starch as the carbon source. Their best anti-tumor activity for MCF-7 was showed at 39.99% ± 2.62 % and 58.49 % ± 2.25%, respectively. Those results showed the polysaccharide of S. crispa can apparently inhibit the cell growth of MCF-7 from activated Raw 264.7.

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利用紅 晶片分析 Monascus pilosus 與 Monascus purpureus 之基因表現差異 凃景瑜 、 英娥、廖麗玲 食品工業發展研究所，生物資源保存及研究中心

Comparison of Genes Expression in Monascus pilosus and Monascus purpureus using Microarray--Ching-Yu Tu, Ing-Er Hwang, Li-Ling Liaw (Food Industry Research and Development Institute, Bioresource Collection and Research Center, P.O. Box246, Hsinchu 300, Taiwan, R.O.C.) 紅麴菌為東方人常用的食品加工菌種，近年來許多研究結果指出紅麴菌具有產生多樣 二次代謝產物及水解酵素的生物特點，因此廣泛應用於保健食品、藥物開發與工業酵素等方 面之開發。本研究主要利用一含 596 個紅麴基因的寡核苷酸晶片探討紅麴菌屬 Monascus pilosus (BCRC38093)與 M. purpureus (BCRC38113) 於液態培養下所呈現的基因表現 圖譜差異。M. pilosus 為 monacolin K 的高產株；而 M. purpureus 為 GABA 的高產株， 其實驗結果可歸納如下：(1)屬於 monacolin K 高產株的 M. pilosus 於 monacolin K 的 基因叢組 mkA~mkI 皆較 M. purpureus 高出 2-20 倍的基因表現量。(2) M. purpureus 於醣解反應的多個重要酵素基因皆較 M. pilosus 表現高，顯示 M. purpureus 具有較活躍 的 醣 類 代 謝 能 力 。 (3) 於 水 解 酵 素 方 面 ， M. purpureus 於 alpha-amylase A 、 nonhemolytic phospholipase C、alpha-glucosidase、galactonate dehydratase、alphaxylosidase 、lipase 1、1,3-beta-glucan synthase component bgs2 與 oligo-1,6glucosidase 等 基 因 有 較 高 的 表 現 量 ； 而 於 lysophospholipase 、 3-phytase A 、uricase 、chitin synthase G、endochitinase 1、bifunctional endo-1,4-betaxylanase xylA、glucan 1,3-beta-glucosidase 與 acetylxylan esterase 等基因則表 現量較低。 In Eastern, Monasus is a fungus used extensively in the food industry. Many researches reported that Monascus could produce many secondary metabolites and hydrolysis enzymes. According to these results, Monascus was applied wildly in the development of healthy food, medicine and industrial enzymes. A Monascus microarray chip, included of 596 genes, was used to study different genes expression in M. pilosus (BCRC38093) and M. purpureus (BCRC31499), which were cultured at liquid medium. M. pilosus is a highly monacolin K producing strain and M. purpureus is a GABA-rich strain. The results from microarry analysis were summarized as follow：(1) Monacolin K genes cluster (mkA~mkI) were expressed 2-20 fold higher in M. pilosus than in M. purpureus. (2) In particular, transcription levels of most catabolic genes of the glycolytic pathway were higher in M. purpureus than in M. pilosus. It implies that M. purpureus expresses more glycolytic activity than M. pilosus. (3) We further studied the expression of genes encoding hydrolytic enzymes. M. purpureus gave the higher gene expression profile of alpha-

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amylase A、non-hemolytic phospholipase C、alpha-glucosidase、galactonate dehydratase、alphaxylosidase、lipase 1、 1,3-beta-glucan synthase component bgs2 and oligo-1,6-glucosidase genes and lower expression levels of lysophospholipase 、 3-phytase A 、 uricase 、 chitin synthase G 、endochitinase 1、bifunctional endo-1,4-beta-xylanase xylA、glucan 1,3-beta-glucosidase and acetylxylan esterasegenes.

紅 麴 大豆甲醇與乙醇萃取物之抗 氧 化性質及其抗 氧 化成分 李煜玲、毛正倫 國立中興大學食品暨應用生物科技學系

Antioxidant properties of methanolic and ethanolic extracts from Monascus fermented soybeans--Yu-Ling Lee and Jeng-Leun Mau（Department of Food Science and Biotechnology, National Chung-Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan） 本研究係以兩株紅 麴 菌 Monascus pilosus BCRC 31527 和 Monascus purpureus BCRC 31499 分別接種於蒸熟大豆上，經培養製成紅麴大豆產品，其經甲醇和乙醇萃取後， 所得到之萃取物進行抗氧化性質及抗氧化成分分析，並以未接種大豆作為控制組。結果顯示， 在抗氧化力（共軛雙烯法）方面，在 5 mg/ml 下，三種樣品的甲醇萃與乙醇萃效果相當， 抗氧化力範圍在 78.1-87.4%。在還原力效果上，三種樣品的甲醇萃比乙醇萃為佳，其中 31499 紅麴大豆效果最好。在 5 mg/ml 下，於捕捉 DPPH 自由基能力方面，甲醇萃的效果 比乙醇萃效果佳，而對三種樣品之間效果依序為 31499＞31527＞大豆，並最高可達到 95.2%。在螯合亞鐵離子能力上，其結果與捕捉 DPPH 自由基能力的結果相同。在抗氧化成 分方面，結果顯示，經由紅麴菌發酵過後的兩種紅麴大豆萃取物，其總酚化合物含量因此 增加。綜此，顯示接種之紅麴大豆，由於紅麴菌的生長，提高酚類化合物含量亦提高其抗氧 化性質。 The objective of this study was to evaluate the antioxidant properties of methanolic and ethanolic extracts from Monascus purpureus BCRC 31499 fermented soybeans (MFS-31499) and Monascis pilosus BCRC 31527 fermented soybeans (MFS-31527) as compared to uninoculated soybean product. Using the conjugated diene method, antioxidant activities of ethanolic and methanolic extracts were similar and in the range of 78.1-87.4% at 5 mg/ml. Concerning to three samples, methanolic extracts showed higher reducing powers than ethanolic extracts and MFS31499 exhibited better reducing power. Regarding to three samples, methanolic extracts showed better scavenging DPPH abilities than ethanolic extracts and in the descending order: MFS-31499 > MFS-31527 > soybeans. In chelating abilities in ferrous ions, the results showed similar pattern as those of scavenging DPPH abilities. The contents of total antioxidant components assayed for

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methanolic and ethanolic extracts were in the descending order: MFS-31499 > MFS-31527 > soybeans. Overall, the result of total phenols increased by incubation with Monascus, MFS-31499 and MFS-31527 showed higher phenolic components and better antioxidant activities.

機能性紅 黑豆之培養 李惠倫、李煜玲、毛正倫 國立中興大學食品暨應用生物科技學系

Development of functional Monascus Fermented Black Soybean Products.--Hu-Lun Lee, YuLing Lee and Jeng-Leun Mau (Departmemt of Food Science and Biotechnology, National ChungHsing University, 250 Kuokuang Road, Taichung, Taiwan) 本 篇 研 究 乃 利 用 黑 豆 作 為 基 質 ， 接 種 紅 麴 菌 株 Monascus purpureus Went 31499 ， 培 養 出 紅 麴 黑 豆 產 品 ， 並 利 用 分 析 其 麥 角 固 醇 、 monacolin K 、 異 黃 酮 (daidzin、daidzein、genistin、genistein) 及橘黴素含量，找出其最適培養條件。結果顯示， 不 同 水 分 及 接 菌 量 下 ， 以 50% 水 分 及 3% 接 菌 量 為 最 佳 ， 此 時 麥 角 固 醇 生 物 量 為 13mg/g；在不同培養溫度及天數下，以 25℃培養 10 天為最佳，可得 monacolin K 含量 （216 ppm），且無檢測出橘黴素含量；添加不同比例碳源的部分，以 10%葡萄糖為最 佳，可得 monacolin K 含量（140 ppm）與橘黴素含量（0.22ppm）；而在添加不同比 例氮源的部分，發現添加後並無提升 monacolin K 產量，反而具有抑制作用；此外，異黃 酮 方 面 ， 隨 著 培 養 天 數 上 升 daidzin 及 genistin 含 量 隨 之 下 降 ， 而 daidzein 及 genistein 則隨之上升。綜合上述條件，得最佳培養條件為以溫度 25℃、水分含量 50%、接 菌量 3%及 10%葡萄糖為碳源，經培養 10 天後，可得最高產量的 monacolin K。 The study used black soybean as the substrate and inoculated Monascus purpureus Went 31499 to cultivate the fermented black soybean product, and analyzed its content of ergosterol, monacolin K, isoflavones (daidzin, daidzein, genistin, genistein) and citrinin to find the suitable conditions. The results showed that under different moisture content and inoculation rates, the 50% moisture and 3% inoculation of Monascus purpureus Went 31499 yields better results, and the content of ergosterol is 13mg/g; The cultivation at 25 ℃ for 10 days under different temperature and days could get the highest amount of monacolin K (216 ppm), and did not detect out any citrinin. In carbon source, addition of 10% glucose could get the highest amount of monacolin K

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(140 ppm) and its citrinin content is 0.22 ppm; We found that addition of nitrogens could not increase the content of monacolin K, but reduce it. Besides, following the cultivation of days, the content of daidzin and genistin decreased, but daidzein and genistein increased. Above all, the optimal conditions were 50% moisture content, 3% inoculation rate, 10% glucose, then cultivated at 25℃ for 10 days, so that we could obtain the best yield of monacolin K.

台灣栽培香菇 SCAR 分子標記建立 蘇秋建、陳隆鐘 中興大學植物病理學系

Identification of Taiwan cultivated strains of Lentinula edodes by SCAR markers Chiu-Chien Su and Lung-Chung Chen (Department of Plant Pathology, National Chung Hsing University, Taichung, Taiwan 402, R. O. C.) 香菇是廣為世界栽培的主要食藥用菌之一，亦為台灣主要栽培的食藥用菌，本研究收 集目前台灣各地主要栽培香菇菌株 48 株及外國菌株 10 株。首先，篩選具栽培地區性代表 性菌株共 21 株與 10 株外國香菇菌株，運用逢機增幅多型性聚合酶鏈鎖反應，篩選出 10 個隨機引子都可以獲得較穩定的 RAPD 譜帶，增幅片段分子量在 0.18 kb-2.52 kb 之間， 單引子可增幅 DNA 片段數目在 7-19 個之間，運用 10 個引子隨機增幅結果應用電腦軟體 NTSYS-pc 依 UPGMA 分群法進行菌株遺傳差異比較，在 0.68 的 DNA 相似係數上將其歸 類為三群而台灣主要集中在中間一群，部分菌株與國外菌株相近，在 0.91 的 DNA 相似係 數上可分為七群，亦說明台灣栽培香菇遺傳生態廣泛，運用 RAPD 結果，篩選出 850 bp 特異性片段，經轉殖定序分析結果其長度為 884 bp，運用 primer 5.0 設計引子 LE7 Forward 與 LE7 Reverse。通過 SCAR-PCR 增幅結果，能夠將 RAPD 之特異片段成功的轉 變成特異性與穩定性更好的 SCAR 標記，此特異性片段長度為 623 bp，為了證明所獲得 DNA 模板的完整與特異性質，亦加入引子 ITS1 與 ITS4 來加以驗證，排除假陽性的之情況。 獲得的香菇菌株 SCAR 標記在收集的 58 個菌株獲得驗證，此片段在台灣目前 921 系列栽 培品種皆可增幅出來，並與國外菌株對照，國外菌株未有此片段增幅，驗證 SCAR 特異標 記在品種間的快速鑑定與檢測之可行性。 關鍵字 : 香菇、逢機增幅多型性聚合酶鏈鎖反應、核酸序列增幅區段 Lentinula edodes (Berk.)Pegler, the shiitake mushroom, is one of the most widely cultivated mushrooms worldwide. The purpose of the project is to investigate the characteristics of 48 isolates or cultivated strains and 10 foreign countries strains. 21 isolated from Taiwan and 10 foreign countries strains were selected. Single 10-base primers were used to generate randomly amplified polymorphic DNA (RAPD) markers in the shiitake mushroom, L. edodes. 10 primers

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produced polymorphisms in all 31 strains tested, producing 7-19 bands ranging from 0.18 to 2.52 kb.10 primers results from 27 primers RAPD profiles with amplification of 31 L. edodes isolates were select to analysis genetic similarity by UPGMA cluster analysis with computer software NTSYS-pc. We found that isolates of L. edodes had different genetic diversity. It indicated that 31 isolates of L.edodes could be divided into three groups under DNA similar coefficient of 0.68 mainly in Taiwan, some isolates were similar to foreigen countries. It indicated that 31 isolates of L. edodes could be divided into severn groups under DNA similar coefficient of 0.91. It indicated that Taiwan cultivate L. edodes have rich genetic diversity.We used results of RAPD to select special bands, cloned and sequenced, then used as bases for designing SCAR marks.We used primer 5.0 to design primer LE7 forward and primer LE7 Reverse.Then the RAPD marks were converted into SCAR marks,which were much more specific and stable and the band size was 623bp. In order to prove the DNA sequence completely and specifical characteristics we obtained.We also took primers ITS1 and ITS4 to prove the possible of false positive.When the SCAR mark was tested using 58 strains collected from Taiwan, its feasibility and reliability was validated. The special could amplify 921 strains and completed with foreign countries not the specifical band. The strain-specific SCAR marks were suitable for the rapid identification of L. edodes Key : Lentinula edodes, random amplified polymorphic DNA, sequence characterized amplified region.

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應用逢機增幅多型性聚合酶連鎖反應反轉錄 mRNA 探討香菇與側耳屬子實體發育過程基因表現之差異 蘇秋建、陳隆鐘 中興大學植物病理學系

Cloning and characterization of genes differentially expressed during fruit body development of Lentinula edodes and Pleurotus species by differential display of RAPD-Chiu-Chien Su and LungChung Chen (Department of Plant Pathology, National Chung Hsing University, Taichung, Taiwan 402, R. O. C. ) 菌類子實體發育過程中相關生理生化及基因表現之變化，一直是真菌學家所感興趣的 研究課題。為了解香菇與側耳屬子實體發育過程基因的表現情形，本研究以本實驗室篩選的 收集菌株香菇 LE16 及側耳屬菌株 P5 與 P6 為研究對象。為了分析香菇與側耳屬子實體發育 過程基因表現，篩選了不同的三個子實體發育時期，其分別為菌絲時期、原基時期與子實體 時期，快速萃取其總量 RNA 並且運用 RT-PCR 反轉錄為 cDNA，篩選出 28 個隨機引子增 幅 cDNA 片段，在香菇子實體發育過程不同時期發現不同的 cDNA 片段，引子 gal 篩選出 在子實體具有的 F1 (200 bp)之特異性片段，原基時期具有的 P2 (300 bp)特異性片段以 及子實體與原基皆具有的特異性片段 P1 與 F2 (250 bp)，經定序結果 F1 片段為 173 bp 與 NCBI 及 EST 比對結果其與 Neurospora crassa 表現疏水性蛋白基因相近，其他相關 片段則仍在定序分析中。在側耳屬中篩選 P6 菌株，引子 P13 能增幅出菌絲生長時期具有的 特異性片段 A (200 bp)，而 S23 能增幅出子實體時期具有的特異性片段 E (500 bp)與 F (690 bp)，P9 菌株則運用 S24 可以增幅在子實體特異性片段 G (300 bp)與 K (700 bp)，目前經轉殖定序結果，其分別為 A、E、F，將定序結果與 NCBI 及 EST 上登錄之序列 比對，其中 A 序列為未知，而 E 序列大小為 484 bp，則與 Ustilago maydis hyposis protein，而 F 序列大小為 684 bp，與 Nurospora crassa filment H protein 相近，未

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來將亦運用北方轉漬法加以佐證其是否為在不同時期的主要表現基因及純化與分析子實體 發育過程時期之不同表現蛋白。 關鍵字: 香菇、側耳屬、應用逢機增幅多型性聚合鏈所反應反轉錄 mRNA Knowledge of the seat of growth with drives morphogensis and appreciation of the balance between physical, biological phenomena and gene expressed intisisted by fungist. In this study, we studied on Lentinula edodes (LE16) and Pleurotus species (P6 and P9). To analysis genes involved in fruit body development of L. edoeds and Pleurotus species, mRNA from three different developmental stages: vegetative mycelium, and fruit body. Total mRNA were isolated and reverse-transcribed to cDNAs. Twenty eight random PCR amplifications were perform with cDNAs, which F1, F2, P1, P2 cDNA fragments from different stage in L. edoeds. By using primer gal, four cDNA clones specifically expressed in primordium (P1, P2)or mature fruit body (F1, F2) were detected. Sequence analysis and databases searchs revealed significant similarity with Neurospora crassa hypothetical protein partial mRNA (F1) and others are still studying. In Pleurotus species respect, Twenty eight random PCR amplifications were performed with the cDNAs, which A, E, F, cDNA fragments from different developmental stages. Sequence analysis and database searches reveraled significant similarity with Ustilago maydis hyposis protein (E), Nurospora crassa filment H protein ( F). In the future, we will use Northern analysis to confirme and protein extracted and analized by different fruit body development stages. Key world: Lentinula edodes, Pleurotus species, differential display of RAPD

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白木耳屬菌株液體發酵液應用於化妝保養品之研究 謝詠筌 1、陳 楨 2 1

南台科技大學生物科技研究所研究生，2 南台科技大學生物科技研究所

Study on the fermentation broth of Tremella strains for the potential use in cosmetic.--YungChuan Hsieh1、Chee-Jen Chen2 (Institute of Biotechnology, Southern Taiwan University of Technology) 實 驗 材 料 有 五 株 白 木 耳 屬 (Tremella) 菌 株 ， 菌 種 分 別 為 Tremella encephala T897、Tremella nivalis CCJ911、Tremella flava CCJ928、Tremella sp. CCJ933 及 Tremella fucifomis CCJ960。利用老鼠黑色素瘤細胞(B16F10)進行黑色素生合成酵素活性 試驗，CCJ897 菌株液體發酵液對於抑制黑色素生合成酵素活性之效果是五株菌株中最好 的，分別抑制 tyrosin hydroxylase 活性 11.44%及 dopa oxidase 活性 27.43%。UVA 照射會使黑色素細胞生成 NO(Nitric oxide)，而 NO 到達足夠量時又會誘導黑色素生合成， 藉由抗發炎試驗來評估白木耳菌株液體發酵液避免黑色素產生之作用；而抗發炎試驗是以 CCJ960 菌株液體發酵液降低 nitrite 產生效果最好，抑制 nitrite 產生 27.36%。利用過氧 化氫(H2O2)做為氧化壓力造成人類子宮頸癌細胞(Hela cell)之細胞損害，發現以 CCJ911 菌株液體發酵液最能降低 H2O2 所引起的細胞損害，具有保護作用可使受 H2O2 傷害之 Hela cell 存活率提高 17.15%。本研究中藉由不同方向及層面，探討白木耳菌株液體發酵 液是否可應用於化妝保養品之可行性評估。 The fermentation broth of five Tremella strains was compared and its character was explored in this study. The five Tremella strains: Tremella encephala CCJ897 、 Tremella nivalis CCJ911 、 Tremella flava CCJ928 、 Tremella sp. CCJ933 and Tremella fucifomis CCJ960. The

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inhibitory effect on the melanin synthetic enzyme via cell culture of mouse melanoma (B16F10) was studied. The culture T897 showed higher inhibitory activities, 11.44% to tyrosin hydroxylase and 27.43% to dopa oxidase, than the other strains respectively. UVA radiation may induce the generation of NO which will stimulate melanocytes to generate and accumulate melanin. NO may induce inflammatory, suggesting the anti-inflammatory response prevent pigmenting of the skin. The result indicated that the culture CCJ960 has inhibited LPS-induced nitrite production up to 27.36% better than the other strains. Meanwhile, the antioxidant activity showed that the culture CCJ911 increased 17.15% of viability of Hela cells prevented the damage from hydrogen peroxide. This study provides some clues towards the understanding the role and mechanisms of action of five Tremella strains by using liquid fermentation for the application in cosmetic.

低溫及機械刺激 對阿魏菇 (Pleurotus ferulae)原基誘導之影響 洪進雄 1、李欣樺 2 1

國立嘉義大學園藝系、2 國立嘉義大學園藝所

The effect of low temperature and mechanical damages on inducement of the primordia formation in Pleurotus ferulae—Chin-Hsiung Hung、Shin-Hua Lee (1Department of Horticultural Science, National Chia-Yi University、2Graduate Institute of Horticulture, National Chia-Yi University) 利用低溫刺激及機械刺激誘發阿魏菇 (Pleurotus ferulae)原基發生，以 10℃低溫處理 阿魏菇菌絲所得之原基出現所需天數為 33.4 日、培養皿出現原基之百分率為 73.3%。此外， 原基發生位置皆位於培養皿外圍的區域，故推測阿魏菇原基主要發生於幼嫩的菌絲。因此， 利用機械破壞處理菌絲，促使菌絲新生。但原基發生位置並不因機械處理而有所差異，推測 阿魏菇原基發生仍需菌絲發育至某種程度的成熟階段。但機械刺激可使原基發生平均日數由 43.3 日縮短為 32.2 日及培養皿出現原基之百分率由 36.7%提高為 42.9%，顯示機械刺 激處理阿魏菇菌絲有利於原基出現，因此 10℃低溫刺激配合機械刺激可有效促使阿魏菇原 基發生。 The stimulation of low temperature and mechanical damages were used to the mycelium and induce the primordia formation of Pleurotus ferulae, the average days of primordial formation by 10℃ treatments were 33.4 days 、the percentage of primordia appeared in the Petri dishes were 73.3%. and primordia were only located at peripheral regions of Petri dish, it would be regarded that primordial only appeared at the young mycelia. and mechanical treatment can induce

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regenerate of mycelia, but didn`t affect the location of primordial., it could be suggested that the formation of primodia need the certain mature stages of mycelia. The stimulation by mechanical damages to mycelia could shorten the average days of primordial formation from 43.3 days to 32.2 days. The percentage of primordia appeared in the Petri dishes were increased from 36.7 % to 42.9%, it showed that the mechanical stimulation could promote the formation of primordial, 10℃ and mechanical stimulation obtained the best induction results.

雞 腿菇多醣組成及其對癌細胞之毒性 蔡淑瑤 1、毛正倫 2 1

亞洲大學生活應用科學學系、2 國立中興大學食品暨應用生物科技學系

Polysaccharides composition of Coprinus comatus and its cytotoxic activities on cancer cells —Shu-Yao Tsai1 and Jeng-Leun Mau 2 (1 Department of Applied Life Science, Asia University; 2 Department of Food Science and Biotechnology, National Chung-Hsing University) 雞腿菇（Coprinus comatus (Mull.: Fr.) S. F. Gray）因其雞腿形狀而得名，亦稱毛 頭鬼傘，是一種食藥兼用的真菌類。利用鹼溶液及熱水萃取雞腿菇以製備多醣，以鹼溶液比 熱水能萃取出更多之雞腿菇多醣。由元素分析發現雞腿菇鹼萃多醣含 N、C 及 H 之量皆高於 雞腿菇熱水萃多醣所含。由中性單糖組成分析來看，發現雞腿菇熱水萃及鹼萃多醣主要是由 甘露糖和葡萄糖所組成。以膠體層析法得知雞腿菇熱水萃多醣的分子量較其鹼萃多醣大。另 以鹼萃及熱水萃所純化的雞腿菇多醣對癌細胞生長情況，結果發現，雞腿菇子實體和菌絲 體 的 鹼 萃 多 醣 對癌 細胞 之生 長抑 制率 IC50 在 A549 分別 為 0.24 和 2.58 mg/mL ； 在 HCT116 分別為 1.14 和 1.58 mg/mL；在 MCF-7 分別為 1.11 和 3.22 mg/mL；在 SKHep-1 分別為 2.56 和 8.46 mg/mL。綜上所述，雞腿菇多醣具有抑制癌細胞生長之能力， 但是其抑制機制為何，則有待進一步研究。 Coprinus comatus (Muller: Fries) S. F. Gray (Coprinaceae), the shaggy mane or chicken drumstick mushroom and also known as the lawyer’s wig mushroom, is a newly cultivated edible and medicinal mushroom. With regard to the hot water or alkali extraction from C. comatus, the yield was the higher for alkali extraction polysaccharides. In elemental analysis, nitrogen, carbon

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and hydrogen contents of alkali extracts polysaccharides from C. comatus were higher than that of their hot water extracts polysaccharides. Neutral sugars were mannose and glucose for all polysaccharides isolated. Using gel filtration, the molecular weights of hot water extracts from C. comatus were higher than that of their alkali extracts. Besides, the studies of the cancer cell viability of hot water and alkali extracts polysaccharides of C. comatus. IC50 values in A549 cell line were 0.24 and 2.58 mg/mL; in HCT116 cell line were 1.14 and 1.58 mg/mL; in MCF-7 cell line were 1.11 and 3.22 mg/mL; in SK-Hep-1 cell line were 2.56 and 8.46 mg/mL for fruit body in alkali extracts polysaccharide, and mycelia in alkali extracts polysaccharide from C. comatus, respectively. Overall, C. comatus contained abundant bioactive polysaccharides, tumor cytotoxicity. However, how to inhibition cancer cell grown of C. comatus polysaccharide need to further research.

探討營養對松茸菌絲體生長特性之影響 楊勝傑 1、李世豪 1、廖宇賡 2

國立嘉義大學 林產科學系所 1 森林暨自然資源系所 2

Study on the Effects of Nutrients on Growth Characteristics of Tricholoma matsutake— Sheng-Jye Yang1, Shih-Hao Lee1, Yue Ken Liao2 (1Department and Graduate Institute of Forest Products Science, 2Department and Graduate Institute of Forestry and Natural Resources, National Chiayi University, 300 University Road, 600 Chiayi, Taiwan) 松茸「Tricholoma matsutake（Ito et Imai）Sing.」學名為松口蘑，在分類學上隸屬蘑菇 目（Agaricales）、口蘑科（Tricholomataceae）、口蘑屬（Tricholoma）。松茸是聞名 於世的食用菌，但因自然生育地劇減，人工栽培困難等因子，是亟待世人開發的高價值真 菌。本研究針對減緩菌絲老化現象，而在培養基內進行不同營養物質添加（MNC、啤酒酵母 花生殼粒片萃取成份）之探討，以尋求最佳之松茸培養基。試驗結果顯示：以 MNC 加上乾 燥啤酒酵母，菌絲生長速度最快且菌絲體無老化現象；而添加花生殼萃取成份則次之；單 純 MNC 培養基內菌絲生長速度最為緩慢；於純啤酒酵母的培養基菌絲於第四週出現了典型 的老化現象，而在 MNC 培養基中並無觀察到此現象。 Tricholoma matsutake is attributed to Tricholomataceae. It is famous for edible fungi all over the

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world. However, due to tremendous decrease in nature culture land and difficulty to cultivate by human, it is highly desired to develop technology to cultivate this valuable fungi. Diminishing aging speed of mycelium is investigated by adding various nutrients （MNC, Ebios, extract from peanut hull）in the culture medium. It is expected to discover the optimal medium in this work. The results show: The growth speed of mycelium in MNC and Ebios culture medium is the best； The MNC and extract from peanut hull culture medium is the next; the MNC culture medium is the worst. The aging of mycelium in the Ebios culture medium is observed in the fourth week. There is no such phenomenon in the MNC culture medium.

不同真菌材料之幾丁聚醣 對 痤 瘡丙酸桿菌脂酶活性抑制之探討 方君仲 1.、蘇慶華 2 1 2

臺北醫學大學細胞及分子生物研究所，

臺北醫學大學生物醫學材料研究所及微生物及免疫學科

The Inhibitory Effect of Chitosans from Fungal Materials on the Activity of Lipase of Propionibacterium acnes --Chun-Chung Fang1 and Ching-Hua Su2 (1Graduate Institute of Cell and Molecular Biology, Taipei Medical University ， 2Graduate Institute of Biomedical Materials and Department of Microbiology and Immunology, Taipei Medical University, 250 Wu-Hsing Street, 110 Taipei, Taiwan) 由痤瘡丙酸桿菌(Propionibacterium acnes)所引發的粉刺 ( Acne vulgaris ) 、 丘疹、膿胞，好發於青春期之男女。痤瘡丙酸桿菌利用本身分泌的細胞外酵素 -脂酶 (Lipase)，分解皮脂獲得養分來源，並進一步誘發痤瘡的產生。幾丁聚醣(Chitosan)對細 菌及部分真菌具有抑菌作用之能力，且之前的研究發現幾丁聚醣(Chitosan)不但具有抑制 痤瘡丙酸桿菌生長的能力，更可能具有直接抑制其產生的細胞外酵素-脂酶(Lipase)活性

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的能力。目前利用 Gene cloning 技術將 lipase gene(GehA) contract 入載體 pDNR 中 並利用 ampicillin 篩選菌株；在經過 agarose gel 分析得到分子量大小為 1197bps 大小 之 DNA 與預期 GehA gene 之大小相同；目前正在運用另一具 T7 promoter 載體 p7GFP2-MCS-Rluc 並利用 IPTG 誘導方式於 E.coli 中大量表現 lipase protein。之後希望利 用來自根黴菌 Rhizopus stolonifer、雙紡錘孢子蟲草 Cordyceps bifusispora 及靈芝殘 渣 Ganoderma tsugae 萃取出之幾丁質，經去乙醯化所得之幾丁聚醣來比較其抑制 Lipase 活性的效果。 Propionibacterium acnes ( P. acnes ) played an important role in the pathogenesis of acne vulgaris . Extracellular lipases produced by P. acnes in vivo act as key enzyme in the hydrolyzation pathway of native sebum triacylglycerols to glycerols and free fatty acids ( FFA ). We contract lipase gene(GehA) into pDNR vector and then subclone this GehA gene into another p7-GFP2MCS-Rluc vector which has T7 promoter to overexpress lipase protein by IPTG induced in E.coli . Finally, we expect to investigate whether or not chitosan from different materials such as cell wall component from Rhizopus stolonifer、Cordyceps bifusispora and Ganoderma tsugaeand possessed the inhibitory potency on the growth of P. acnes and/or the activity of lipase. Putative lipase activity was detected by lipase zymography and the observation of the interactive precipitate between FFA and Victoria Blue B ( VBB ).

利用 IGS-RFLP 分析病原性及非病原性之唐菖蒲萎凋病菌 紀廷霖、陳瑞祥 國立嘉義大學生物科技研究所

IGS-RFLP analysis of pathogenic and nonpathogenic Fusarium oxysporum f. sp. gladioli. -- TingLin Chi and Ruey-Shyang Chen (Graduate Institute of Biotechnology, National Chiayi University, Chiayi 600, Taiwan) Fusarium oxysporum 經常表現出高度寄主專一性之特性，並且可依其寄主種類的 不同進一步區分成不同之分化型。本研究主要分析 68 株 F. oxysporum f. sp. gladioli 菌 株之遺傳歧異性。利用聚合酵素連鎖反應增幅核糖體 DNA 中之 intergenic spacer (IGS) 區域以進行限制酵素長度多型性分析。經由三種限制性酵素 BstEII、ScaI 和 XhoI 的作用， 病原性和非病原性的 F. oxysporum f. sp. gladioli 菌株因具有不同之圖譜形式而可區分 為不同群。 大部分病原性菌株皆歸屬於同一群，顯示其序列相似度相當高，可藉此推論病 原性菌株可能皆來自同一親源。

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Pathogenic isolates of Fusarium oxysporum often display a high degree of host specificity and may be subdivided into formae speciales based on the host species attacked. In the present study, sixty-eight isolates of F. oxysporum f. sp. gladioli were analyzed for genetic diversity. The intergenic spacer (IGS) regions of the rDNA of F. oxysporum f. sp. gladioli were amplified by PCR and an IGS-RFLP analysis was performed. Restriction digestion with BstEII, ScaI and XhoI allowed differentiation between pathogenic and nonpathogenic isolates of F. oxysporum f. sp. gladioli. Pathogenic and nonpathogenic isolates were separated into different groups based on RFLP patterns, although some nonpathogenic isolates grouped with pathogenic isolates. The population of pathogenic isolates was less diverse than that of the nonpathogenic isolates, suggesting that the pathogenic isolates were possibly of monophyletic origin.

本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案

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本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 本頁請找尋代替 案 真菌幾丁聚醣抗致突變性之評估 顏名聰 1、曾裕琇 2、毛正倫 3 1

嘉南藥理科技大學生活應用與保健系，2 高雄餐旅學院烘焙管理系 3 中興大學食品暨應用生物科技學系

Antimutagenic properties of fungal chitosans from shiitake stipes--Ming-Tsung Yen1 Yu-Hsiu Tseng2 and Jeng-Leun Mau3(1Department of Applied Life Science and Health, Chia Nan University of Pharmacy and Science, 60 Erchjen Road, Section 1, Jente, Tainan, Taiwan 71710, R.O.C. ， 2Department of Baking Technology and Management, National Kaohsiung Hospitality College, 1 Sung-Ho Road, Kaohsiung 812, Taiwan, R.O.C. ， 3Department of Food Science and Biotechnology, National Chung-Hsing University, 250 Kuokuang Road, Taichung, Taiwan 40227, R.O.C. 癌症發生的原因與飲食具有極大相關性，因此膳食所扮演的角色愈來愈被重視。同時亦 有研究發現我們的食物中存在有天然的致突變物質、致癌成分及抗致突變物質。幾丁質及其 衍生物因為具有抗致突變性，且具有抑制致突變物之代謝活性。故本研究藉由香菇柄製備真 菌幾丁聚醣對直接致變物之抗致突變性，以了解真菌幾丁聚醣抗致突變之功效。結果顯示， 真菌幾丁聚醣 B90、B120 與 C90、C120 之毒性試驗，在濃度 0.05〜5 mg/plate 不論有 無添加 S9 混合物，真菌幾丁聚醣 B90、B120 與 C90、C120 對 S. typhimurium TA98 及 TA100 菌 株 均 不 具 毒 性 ， 菌 數 存 活 率 分 別 在 100% 以 上 ； 致 突 變 試 驗 在 0.05 〜 5 mg/plate 濃度下，不論有無添加 S9 混合物，真菌幾丁聚醣 B90、B120 與 C90、C120 對 S. typhimurium TA98 及 TA100 菌株均不具致突變性，其致突變性比率範圍分別在 0.55 ～1.11 及 0.57～1.07。

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The cancer to come about the maximun to be bound up with of diet, this food to impersonate is being of attention. Also have been reported of discover that has natural to send turn of events, carcinogenic composition and antimutagenic material from food. Chitin and derivatives have antimutagenic and to restrain a sudden change of metabolism activity effect. Furthermore, the fungal chitosan of immediate in an opposite to send change matter of antimutagenic from shiitake stipes were studied. Fungal chitosan showed is no toxicity was found in Salmonella typhimurium TA98 and TA100 was observed at 0.05-5 mg/plate of fungal chitosan B90, B120, C90 and C120 with a survival of 100%, respectively. The result showed is no mutagenicity found in Salmonella typhimurium TA98 and TA100 was observed at 0.05-5 mg/plate of fungal chitosan B90, B120, C90 and C120 with the mutagenicity of rate range 0.55～1.11and 0.57～1.07, respectively.

以納豆菌生產γ-聚麩胺酸之研究 李順來*、鄭明福、許孟博 南台科技大學生物科技研究所

Study of production of poly(γ-glutamic acid) by Bacillius subtilis (natto) -- Shun-Lai Li*, Ming-Fwu Jeng and Meng-Bor Hsu ( Graduate Institute of Biotechnology, Southern Taiwan University of Technology, No.1, Nantai St, Yung-Kang City, Tainan, Taiwan) 聚麩胺酸[poly(γ-glutamic acid)；γ-PGA]為日本傳統發酵食品納豆中，水溶性微生 物高分子聚合物，可作為環境保護、保健食品、醫藥、化妝品等用途。一般而言，發酵液黏度 増加，代表 γ-PGA 產量提高。本研究以市售納豆菌 N1 為生產菌株，期望經由培養參數之篩 選，探討 γ-PGA 產率增高可行性。納豆菌以 37℃、150 rpm 培養實驗，得知下列結果: 一. 納豆菌 N1 培養於 LB 培養基中( Yeast extract 15g/L、Tryptone 5g/L、Sodium chloride 4g/L、Glucose 30g/L )，菌量成長較高。二.採用於 LB 培養基活化二次之菌種顯示:能提昇 菌 量 60 ％ 。 三 . 納 豆 菌 N1 發 酵 於 修 飾 之 M2 培 養 基 中 ( Monosodium glutamate monohydrate 171.7g/L、Glucose 80g/L、Peptone 15g/L、Yeast extract 5g/L、Urea 3g/L、K2HPO4 2g/L )，獲得較高 γ-PGA 產量。四. 發酵期間使用修飾過 M2 培養基饋料，

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能増加黏度 63.1％(提高 γ-PGA 產量)，同時縮減 13.5％發酵時間。 結論顯示: 採用於 LB 培養基活化二次之 N1 菌種，加入等差饋料之修飾過 M2 培養基培養，確能明顯提高 γPGA 產量。 Poly(γ-glutamic acid) (γ-PGA) is a water-soluble biopolymer that can be found in Japanese traditional fermented food, natto, which has potential applications in the environmental protection, health food, medicine and cosmetics. In general, the increase in broth viscosity implies the increase ofγ- PGA content. This study has focused on the screening of cultural parameters for high γ-PGA production by B. subtilis (natto) N1. In this study, B. subtilis (natto) was incubated at 37℃ and 150 rpm. The following results were obtained: 1, N1(natto) generated higher biomass as cultured in LB medium ( Yeast extract 15g/L, Tryptone 5g/L, Sodium chloride 4g/L and Glucose 30g/L ). 2, The biomass increased up to 60% by using the inoculum that was activated twice in LB medium. 3, Higher γ-PGA production was obtained when N1(natto) was fermented in modified M2 medium ( Monosodium glutamate monohydrate 171.7g/L, Glucose 80g/L, Peptone 15g/L, Yeast extract 5g/L, Urea 3g/L and K2HPO4 2g/L ). 4, Feeding modified M2 medium during fermentation could not only raise 63.1% of the broth viscosity, corresponding with higherγ- PGA production, but also reduce 13.5% of the fermentation time. In conclusion：The two-step activation procedure of N1 in the LB medium and then cultivated in the modified M2 medium with step-increase feeding strategy could significantly increaseγ- PGA product.

蜜環菌醱酵物增長戊巴比妥塩誘導小鼠睡眠效果 李順來、 雅芬、許孟博 南台科技大學生物科技所

The enhancement of pentobarbital-induced hypnosis in mice by fermentation products of Armillariella mellea.－Shun-Lai Li 、Ya-Fen Huang、Meng-Bor Hsu(Graduate Institute of Biotechnology ,Southern Taiwan University of Technology) 蜜環菌，分佈於全球各地區，為口磨科植物，也是偶見木棲腐生或木棲寄生的野菇之一 種，為食療皆可。本實驗利用蜜環菌與中藥材天麻及酸棗仁進行共同醱酵，所得醱酵物餵食 小鼠一週後進行睡眠實驗測試，以戊巴比妥塩誘導小鼠入睡，並觀察小鼠入睡時間與睡眠 時間是否改變。入睡時間是從注射戊巴比妥塩後到失去反射動作期間，而睡眠時間則是從失 去反射動作到恢復期間稱之。

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蜜環菌天麻醱酵物餵食組鼠較未餵食組小鼠睡眠時間延長一倍時間，餵食蜜環菌酸棗 仁發酵物也可延長 90%睡眠時間。而餵食不同劑量蜜環菌天麻及蜜環菌酸棗仁醱酵物小鼠 入睡時間較低於未餵食組別 62%到 90%。此結果証實蜜環菌複方醱酵產品對幫助睡眠具有 相當顯著的效果。 Armillariella mellea is one of the Tricholomataceae mushrooms that spread worldwide and can be used as food and medicine. This study is to explore the enhancement effect of the fermentation products of Armillariella mellea with Gastrodia dioscoreirhiza or Ziziphus spinosa on the pentobarbital-induced hypnosis in mice .A short period (7 days) of oral administration to mice twice daily with fermentation products was treated before hypnosis tests were conducted. The sleeping onset time and duration time were then measured on the pentobarbital-induced mice. The onset time was evaluated as the time period from pentobarbital injection to the loss of righting reflex and the sleeping duration time was measured by recording the duration from loss to recovery of righting reflex. The study showed that the sleeping duration time induced by pentobarbital (60mg/kg)was doubled by treating with the fermentation products of Armillariella mellea with Gastrodia dioscoreirhiza and prolonged 90% by that of Armillariella mellea with Ziziphus spinosa .The onset time by treating with the fermentation products of Armillariella mellea with Gastrodia dioscoreirhiza or Ziziphus spinosa was shorter 62% to 90% than that untreated .The results showed that the fermentation products of Armillariella mellea with Gastrodia dioscoreirhiza or Ziziphus spinosa did significantly improve sleep.

美白化妝品資材的開發與研究 許菱晏、 繼光 亞洲大學生物科技暨生物資訊研究所-生科組

A stu dy of the nat ural prod ucts for wh iteni ng Cosmetical use- Hsu,Ling-Yen, Wu, ChiGuang(Department of Biotechnology and Bioinformatics, Asia University, Taichung, Taiwan) 為了解決東方女性的膚色困擾, 近幾年來有許多的研究都試著從中草藥與微生物中去找尋 具有阻止色素沉澱,以達美白功效之酪胺酸酶抑制物，並可進一步應用到化妝品製造上。酪 胺酸酶 (Tyrosinase：EC 1.14.18.1) 為一含銅之多酚氧化酶，是皮膚黑色素形成與蔬果 褐變的重要因子。在研究中所使用的材料有枸杞油、柚子葉、樟芝、紅麴、黑木耳、酪梨葉，研

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究中除了枸杞油外,其他材料皆以純水加熱至90°C來進行萃取，檢測其體外抑制酪氨酸酶 活性的能力。本研究以維他命C作為比較之標準，經測試後, 各材料對於酪胺酸酶的抑制能 力分別為-枸杞油 60%, 柚子葉 56%, 樟芝 35%, 紅 13%, 黑木耳 7%, 酪梨葉 36%。 In order to whiten Oriental female's skin color; many researchers have dedicated to search for suitable agents from Chinese herbs and microbial resources to arrest pigmentation. Hopefully, these agents could be applied to the cosmetics manufacturing. Tyrosinase (EC1.14.18.1) is a copper-containing polyphenol oxidase and a key factor for the melanin synthesis. In this study, the materials include Lycium seed oil, Citrus grandis, Antrodia camphorata, Monascus sp., Auricularia polytricha. Except Lycium seed oil, other materials all were extracted with water at 90 ° C. To compare with Vitamin C as standard, the tyrosinase inhibitory capability for each test material was Lycium seed oil 60%, leaves of Citrus grandis 56%, Antordia camphorata 35%, Monascus 13%, Auricularia polytricha 7%, leaves of Avocado 36%.

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