Development of Free Radical Products during the Greening Reaction of Caffeic Acid Esters (or Chlorogenic Acid) and a Primary Amino Compound
ESR spectra were measured directly on a marked greening reaction mixture of Et-caffeate and a primary amino compound in alkali solution under aeration. A clear hyperfine structure was commonly detected early in the greening reaction with different amino compounds. Its hyperfine spectrum split into seven peaks was analyzed and found to be due to the oxidized free radical product of the Et-caffeate using an authentic sample system. Another type of hyperfine ESR spectrum was observed later in the reaction, and was altered with different amino compounds. The hyperfine structure for n-butylamine split into 12 lines. The latter type of free radical products were assumed to be a semiquinone type radical compounds of the trihydroxy benzacridine derivative, which was identified as the principal structure of the green and yellow pigments formed by this greening reaction system. A formation mechanism of the green pigment and related products involving these free radical products is proposed.
Greening pigments produced reaction of ethyl caffeate with methylamine.
Little is known about the greening reaction of chlorogenic acid with amino acid in alkaline conditions. The isolation and characterization of the pigment were attempted by using ethyl caffeate and methylamine instead of chlorogenic acid and amino acid. The green compounds were purified by using Sephadex G-10 and thin layer chromatography, and were analyzed by ESR and C-13 MNR. The ESR spectrum of F3 (corresponding to purified pigment) in a liquid state showed that the structure of the C-OH moiety seemed to give either the quinone type or its polymerized form. The ESR spectrum of F3 in a solid state showed that the g-value of this fraction was 2.003. On the basis of the C-13 NMR spectrum, the signals assigned to two carbons (C1" and C-2") of the alcohol moiety of ethyl caffeate were observed clearly in F3. Considering the chemical shift of C-1 in ethyl caffeate, the C-1 in F3 could exist as -C=N-CH3. F3 seemed to be a precursor of the greening compound.
Spectrophotometric Studies on the Behavior of Chlorogenic Acid in Alkaline Aqueous Solutions
クロロゲン酸の緑変機構を明らかにする研究の一環として, まず, 室温で, 数種のアルカリ性水溶液におけるクロロゲン酸の経時的な変化, およびクロロゲン酸と含窒素化合物との反応で起こる緑変について, 紫外・可視領域の分光測定的検討を行なった。pH 9付近のMenzel緩衝液, 炭酸ナトリウムアルカリ性水溶液におけるクロロゲン酸の吸収の最も強いλmaxは, 水溶液中での場合より, 42nm長波長領域ヘシフトすることが観察された。またクロロゲン酸の吸収スペクトルの経時的変動について, pH 9付近のMenzel緩衝液, 炭酸ナトリウム, 炭酸水素ナトリウムなどのアルカリ性水溶液中での検討から, クロロゲン酸は366nmにλmaxを示す黄色を呈する物質 (366物質) に変化し, このものを経て自動酸化されることが明らかになった。そして, 366物質があらわれているときにアラニン, フェニルアラニン, グルタミン酸およびメチルアミンなどを添加した場合だけ, 新たに可視領域に特徴的なλmax 675～695nmを示す緑色液が得られた。また, 上記緑色液のλmax 675～695nmの吸光度は, いずれの含窒素化合物を用いたときも, λmax 366nmの吸光度の減少に対応して増加していくことが認められた。したがって, pH 9付近のアルカリ性水溶液中で生じる366物質が, アラニン, フェニルアラニン, メチルアミンなどのアミノ基をもつものと反応し, 緑変することが明らかになった。