Chemical Modification of Chlorophylls for Photodynamic Therapy

发现在光照射下，氯苯基分解DNA。为了用于光动力疗法，叶绿素必须进行化学修饰以积累到癌细胞中。然而，叶绿素对热、酸碱和光高度敏感，会发生脱金属或异构化，因此，很难通过普通的化学反应改变其结构。我们尝试通过酶如蛋白酶或脂肪酶的催化来水解和酯交换叶绿素。但由于叶绿素微溶于水，因此必须使用有机溶剂，有机溶剂对酶活性有强烈的抑制作用。通过圆二色光谱的测定，发现有机溶剂对酶的抑制主要是由于二级结构的改变。然后，研究了添加剂对酯交换反应中蛋白酶活性和稳定性的影响，发现环糊精对酶的冷冻干燥显著提高了酶的活性和稳定性。光敏剂如Photofrin和准分子激光目前用于光动力疗法。为了开发更有效的光动力治疗方法，采用脱镁叶绿酸a钠盐作为光敏剂进行了体外实验。使用光吸收法测量脱镁叶绿酸在癌细胞中的积累。发现脱镁叶绿酸被有效地掺入癌细胞中。4 h后达到最大浓度。结果表明，脱镁叶绿酸类化合物是一种比较稳定的光敏剂。

It was found that chlorophrylls decompose DNA under light irradiation. To use for photodynamic therapy, chlorophylls must be chemically modified to accumulate into cancer cells. However, chlorophylls are highly sensitive to heat, acid-base, and light, suffering from demetallation or isomerization, and therefore, it is difficult to modify the structure by ordinary chemical reactions. We attempted at hydrolysis and transesterification of chlorophylls by the catalysis of enzymes such as proteases or lipases. However, chlorophylls were only slightly soluble in water, and therefore use of organic solvents was necessary.Organic solvents strongly inhibited the activity of enzymes. The changes in structures of proteases were detected by measurements of CD spectrum, and it was found that inhibition of enzymes by organic solvents was mostly due to modification of secondary structures. Then, the effects of additives on activity and stability of proteases were investigated for transesterification, and it was found that colyophilization of enzymes with cyclodextrins dramatically increased the activity and stability of the enzymes.Photo-sensitizers, such as Photofrin, and eximer laser are currently used for photodynamic therapy. To develop more effective methods of photodynamic therapy, in vitro experiments were made using soditum salt of pheophorbide a as a sensitizer. Accumulation of the pheophorbide into cancer cells was measured using light absorption method. It was found that the pheophorbide was effectively incorporated into cancer cells. Maximum concentration was attained after 4 h. The results suggest that pheophorbides are potential candidate of light-sensitizer that are relatively stable.