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酵素法による魚類筋肉NADおよびATPの定量条件に関する吟味
https://fra.repo.nii.ac.jp/records/2005385
https://fra.repo.nii.ac.jp/records/20053857ba496b1-9e0d-4677-84ee-458e95c2a08f
| Item type | 紀要論文 / Departmental Bulletin Paper(1) | |||||
|---|---|---|---|---|---|---|
| 公開日 | 2024-05-22 | |||||
| タイトル | ||||||
| タイトル | 酵素法による魚類筋肉NADおよびATPの定量条件に関する吟味 | |||||
| 言語 | ja | |||||
| タイトル | ||||||
| タイトル | A Study on Enzymatic Method for Determining NAD and ATP in Fish Muscle | |||||
| 言語 | en | |||||
| 言語 | ||||||
| 言語 | jpn | |||||
| 資源タイプ | ||||||
| 資源タイプ識別子 | http://purl.org/coar/resource_type/c_6501 | |||||
| 資源タイプ | departmental bulletin paper | |||||
| アクセス権 | ||||||
| アクセス権 | metadata only access | |||||
| アクセス権URI | http://purl.org/coar/access_right/c_14cb | |||||
| 著者 |
江平, 重男
× 江平, 重男 |
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| 抄録 | ||||||
| 内容記述タイプ | Abstract | |||||
| 内容記述 | Coenzymes, Nicotinamide Adenine Dinucleotides (oxidized and reduced forms of NAD and NADP) and ATP in animals tissues have been enzymatically determined by many workers mostly in the field of biochemistry or clinical biochemistry. However, studies on the accuracy of the methods applied to fish muscle are obscure. In this report, accuracy of enzymatic methods for the determination of NAD and ATP in fish muscle was examined. Muscle was extracted by using perchloric acid and neutralized as in the case of measuring index for estimating freshness of fish, K value, because K value has come to be widely measured in a study on keeping freshness or post mortem changes in fish muscle. NAD was assayed by employing ethanol-alcohol dehydrogenase (ADH) system, and ATP was determined by a coupled reaction of glucose-hexokinase (HK)-glucose-6-phosphate dehydrogenase (G-6-PDH) system. The methods were essentially based on those described by MATSUHASHI and LAMPRECHT et al., respectively, except that the amount of sample extract was increased upto 1.0 ml in order to minimize the error in determination. The results obtained were summarized as follows; 1) It was confirmed that perchloric acid extract of fish muscle prepared according to the method described for measuring freshness estimation index, K value, could be used as it is as sample solution for the assay of NAD as well as ATP. 2) Recoveries of NAD added to deionized water, muscle extract, and muscle slices were 97-104% in the range of 10-400 nmoles per assay solution (Fig. 1), 97-103% in the same range (95% confidence limits of the mean: 100 ± 0.8% for 100 nmoles per assay solution (Table 3)), and 97-99% for 4 μmoles added to per g muscle (Table 4). 3) NADP-specific activities contained as impurity in the ADH preparation used was less than 1% of NAD-specific activities in the range upto 200 nmoles of NADP per assay solution (Table 6). Taking into account the NADP content reported in fish muscle, interference by this contaminating activity was thought to have no effect on the accuracy. 4) The range of NAD concentration in fish muscle to be effectively determined by this method was calculated to be 0.1-4 μmoles per g muscle. 5) When G-6-PDH preparation suspended in ammonium sulphate solution was used, all procedures for the assay of ATP were carried out within 30 min (Fig. 4). With regard to crystalline lyophilized preparation, however, 80-120 min were needed. This discrepancy was considered to be caused by its low solubility in water. 6) Recoveries of ATP from the extract of fish muscle decreased with its concentration, showing more than 95% in the range of 80-220 nmoles per assay solution (Fig. 5). | |||||
| 言語 | en | |||||
| 抄録 | ||||||
| 内容記述タイプ | Abstract | |||||
| 内容記述 | 1. 魚類生鮮度判定恒数,K値測定用に調製した過塩素酸抽出液は,そのままNADおよびATP 測定試料として使用しうることを認めた。2. 上記抽出液中の NADは,アルコール脱水素酵素を用いて精度よく測定しうることを認めた。脱イオン水,筋肉抽出液,および筋肉片に添加した NADの回収率は,それぞれ1反応液当り10~400 nmolesの濃度範囲で97~104%(Fig.1),反応液当り10~400 nmolesの範囲で97~103%(添加量 100 nmolesの回収率平均値の95%信頼区間:100±0.8%)(Table 3),筋肉1g当り4μmolesに対し,97~99%であった(Table 4)。3. 使用したアルコール脱水素酵素標品中に含まれる不純物酵素の NADP-特異活性は,1反応液当り 200 nmloes 以下の添加NADP に対し,NAD-特異活性の1%以下にすぎなかった(Table 6)。報告された魚類筋肉 NADP 含量から,不純物の NADP-特異活性に基づくNAD 測定誤差は無視しうると考えられた。4. 本酵素法による NAD 有効測定範囲は,0.1~4.0μmoles/g であった。5. ヘキソキナーゼ,NADPおよびグルコース-6-リン酸脱水素酵素を用いるATPの定量において,凍結乾燥したグルコースー6-リン酸脱水素酵素は水に溶解しにくい傾向が認められ,測定に80~120分を要したが,硫酸アンモニア懸濁標品では30分以内に測定を終了しえた(Fig. 4)。6. 筋肉抽出液に加えた ATPの回収率は,1反応液当り25 nmoles以下ではばらつきが大きく,80 nmoles以上では濃度とともに減少する傾向がみられた。最適測定濃度は反応液当り80~220 nmolesと判断され,95~103%の回収率が得られた(Fig. 5)。 | |||||
| 言語 | ja | |||||
| 書誌情報 |
ja : 東海区水産研究所研究報告 en : Bulletin of Tokai Regional Fisheries Research Laboratory 巻 109, p. 57-76, ページ数 20, 発行日 1983-02-15 |
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| 出版者 | ||||||
| 出版者 | 東海区水産研究所 | |||||
| 言語 | ja | |||||
| 出版者 | ||||||
| 出版者 | Tokai Regional Fisheries Research Laboratory | |||||
| 言語 | en | |||||
| ISSN | ||||||
| 収録物識別子タイプ | PISSN | |||||
| 収録物識別子 | 0040-8859 | |||||
| 書誌レコードID | ||||||
| 収録物識別子タイプ | NCID | |||||
| 収録物識別子 | AN00156834 | |||||
| 情報源 | ||||||
| 識別子タイプ | Local | |||||
| 関連識別子 | tokai_k_109_57 | |||||
| 関連サイト | ||||||
| 識別子タイプ | URI | |||||
| 関連識別子 | https://agriknowledge.affrc.go.jp/RN/2010280531 | |||||
| 言語 | ja | |||||
| 関連名称 | 日本農学文献記事索引(agriknowledge) | |||||
