Photodynamic therapy (PDT) is currently being used as an alternative therapeutic modality for a variety of malignant tumors. This study was performed to show an efficient preparation of second generation of photosensitizer chlorin e6 (Ce6) with high yield and purity, and to test antitumor activity of Ce6-induced PDT (Ce6-PDT) both in vitro and in vivo using a rat tumor model. Three-week-old male Sprague-Dawley (SD) rats were inoculated s.c. on the right flank with 5x106 RK3E-ras cells. The animals were admin-istered i.v. with Ce6 (10 mg/kg) and 24 h later, PDT was performed using a laser diode at a light dose of 100 J/cm2. Ce6-PDT generated reactive oxygen species and led to significant growth inhibition in RK3E-ras cell. In addition, Ce6-PDT induced apoptosis through the activation of caspase-3 and its downstream target, PARP cleavage. The protein level of anti-apoptotic bcl-2 was also reduced by Ce6-PDT in RK3E-ras cells. In in vivo experiments, application of Ce6-PDT led to a significant reduction of tumor size. PCNA immunostaining and TUNEL assay revealed that Ce6-PDT inhibited tumor cell proliferation and increased apoptosis. These findings suggest that the newly purified Ce6-PDT can effectively arrest tumor growth by inhibiting cell proliferation and inducing apoptosis.  ( source: Oncology Reports, 2009, 22(5). 1085-1091. Doi: 10.3892/or_00000540. Efficient preparation of highly pure chlorin e6 and its photodynamic anti-cancer activity in a rat tumor model. Authors: Yeon-Hee Moon, Seong-Min Kwon, Hyo-Jun Kim, Kwan-Young Jung, Jong-Hwan Park, Soo-A Kim, Yong-Chul Kim, Sang-Gun Ahn, Jung-Hoon Yoon. Affiliations: Department of Pathology, School of Dentistry, Chosun University, Gwangju 500-759, Korea, http://www.spandidos-publications.com/or/22/5/1085 ). Photodynamic therapy (PDT) is currently being used as an alternative therapeutic modality for a variety of malignant tumors. This study was performed to show an efficient preparation of second generation of photosensitizer chlorin e6 (Ce6) with high yield and purity, and to test antitumor activity of Ce6-induced PDT (Ce6-PDT) both in vitro and in vivo using a rat tumor model. Three-week-old male Sprague-Dawley (SD) rats were inoculated s.c. on the right flank with 5x106 RK3E-ras cells. The animals were admin-istered i.v. with Ce6 (10 mg/kg) and 24 h later, PDT was performed using a laser diode at a light dose of 100 J/cm2. Ce6-PDT generated reactive oxygen species and led to significant growth inhibition in RK3E-ras cell. In addition, Ce6-PDT induced apoptosis through the activation of caspase-3 and its downstream target, PARP cleavage. The protein level of anti-apoptotic bcl-2 was also reduced by Ce6-PDT in RK3E-ras cells. In in vivo experiments, application of Ce6-PDT led to a significant reduction of tumor size. PCNA immunostaining and TUNEL assay revealed that Ce6-PDT inhibited tumor cell proliferation and increased apoptosis. These findings suggest that the newly purified Ce6-PDT can effectively arrest tumor growth by inhibiting cell proliferation and inducing apoptosis.  ( source: Oncology Reports, 2009, 22(5). 1085-1091. Doi: 10.3892/or_00000540. Efficient preparation of highly pure chlorin e6 and its photodynamic anti-cancer activity in a rat tumor model. Authors: Yeon-Hee Moon, Seong-Min Kwon, Hyo-Jun Kim, Kwan-Young Jung, Jong-Hwan Park, Soo-A Kim, Yong-Chul Kim, Sang-Gun Ahn, Jung-Hoon Yoon. Affiliations: Department of Pathology, School of Dentistry, Chosun University, Gwangju 500-759, Korea, http://www.spandidos-publications.com/or/22/5/1085 ).           Methods to make Chlorin E6 make Chlorin E6 Chlorin E6 is a natural product, and usually can be made from Live chlorella (Chlorella ellipsoidea, see the following pictures) Chlorin E6 is a natural product, and usually can be made from Live chlorella (Chlorella ellipsoidea, see the following pictures)                     The following procedure was reported in Oncology Reports, 2009, 22(5). 1085-1091 by Korean scientists Yeon-Hee Moon et al.   Live chlorella (Chlorella ellipsoidea) 100 g (dried weight) was sequentially washed with 500 ml of water and 300 ml of 50% ethanol in water to remove polar materials and the residue was extracted twice with 500 ml of 100% ethanol to obtain chlorophyll a rich fraction (extraction yield 4.3%). Stirring the combined ethanol solution of chlorophyll a in 1 N HCl (pH 2.5) for 3 h at room temperature afforded pheophytin in the form of precipitates. The precipitate was dissolved in dichloromethane washed with distilled water, dried with anhydrous sodium sulfate, and rotary-evaporated to dryness. The residue was purified by a chromatography using neutral alumina (Aldrich, Brockmann, ~150 mesh) with a gradient elution from 30% dichloromethane in n-hexane to 100% dichloromethane. The main green band was collected and evaporated to dryness. The crystalline powder was dissolved in acetone, adjusted pH 12.0 with 1 N NaOH, and stirred for 12 h. The precipitated Ce6 was filtered, washed with acetone and dissolved in 100 ml of water, and filtered to remove insoluble impurity. After lyophilization of the filtered water solution, a fine black powder of Ce6 was obtained. The purity of Ce6 is 93-98% (yield of Ce6: 1% from dried weight of chlorella). The following procedure was reported in   Live chlorella (Chlorella ellipsoidea) 100 g (dried weight) was sequentially washed with 500 ml of water and 300 ml of 50% ethanol in water to remove polar materials and the residue was extracted twice with 500 ml of 100% ethanol to obtain chlorophyll a rich fraction (extraction yield 4.3%). Stirring the combined ethanol solution of chlorophyll a in 1 N HCl (pH 2.5) for 3 h at room temperature afforded pheophytin in the form of precipitates. The precipitate was dissolved in dichloromethane washed with distilled water, dried with anhydrous sodium sulfate, and rotary-evaporated to dryness. The residue was purified by a chromatography using neutral alumina (Aldrich, Brockmann, ~150 mesh) with a gradient elution from 30% dichloromethane in n-hexane to 100% dichloromethane. The main green band was collected and evaporated to dryness. The crystalline powder was dissolved in acetone, adjusted pH 12.0 with 1 N NaOH, and stirred for 12 h. The precipitated Ce6 was filtered, washed with acetone and dissolved in 100 ml of water, and filtered to remove insoluble impurity. After lyophilization of the filtered water solution, a fine black powder of Ce6 was obtained. The purity of Ce6 is 93-98% (yield of Ce6: 1% from dried weight of chlorella).