| Photodynamic therapy (PDT) is a developing cancer treatment modality. It has many advantages over current cancer treatment modalities, such as surgery, chemotherapy, and radiation therapy. One of the current focuses in this field is the development of anticancer photosensitizer for PDT. Pentalysine 2-carbonylphthalocyanine zinc [ZnPc-(Lys)5] is a novel anionic photosensitzer, which shows a higher level tumor cell uptake and 20-fold higher phototoxicity on tumor cells compared to anionic zinc phthalocyanine photosensitizers. ZnPc-(Lys)5 is expected to be an ideal anticancer photosensitizer. Currently, there were few reports on the phthalocyanine derivatives photosensitizer analysis in vivo. The determination of the free concentration phthalocyanine derivatives photosensitizer has not been reported.In the first part, it were studied pharmacokinetic parameters and tissue distribution of ZnPc-(Lys)5 in tumor-bearing mice. The experiments using the SDS micellar system for the determination of ZnPc-(Lys)5 in tumor-bearing mice(H22).In plasma the linear range of ZnPc-(Lys)5 were 10 ~500 nmol/L, r =0.9993; the limit of detection was 3 nmol/L, the limit of quantification was10 nmol/L; the relative recoveries were 96.20% ~ 102.74%; the relative standard deviations (RSD) were all under 10%. In tissue the linear range of ZnPc-(Lys)5 were 25 ~1250 ng/mL; the relative recoveries were 94.93% ~ 103.74%; the relative standard deviations (RSD) were all under 10%.After i.v. single dosing at 1 mg/kg to tumor-bearing mice, the ZnPc-(Lys)5 in the plasma and tissue sample were determinated using micellar enhanced spectrofluorometric method. The plasma concentration-time curve was simulated and pharmacokinetic parameters were calculated by 3P97 software. The pharmacokinetics fitted as an open two-compartment model. The main pharmacokinetic parameters were estimated to be follows: 2.07 h for t1/2α, 48.14 h for t1/2β. The pharmacokinetic parameters suggest that ZnPc-(Lys)5 was distributed fast and eliminated slowly. ZnPc-(Lys)5 in the tissue distribution order of tumor-bearing mice: liver> tumor > kidney> lung> heart> intestine> spleen> muscle> skin> brain. It was hardly detected in brain.In the second part, comparison of ultrafiltration and solvent precipitation (adding cold acetone, cold ethanol, trichloroacetic acid, trifluoroacetic acid, respectively) for the measurement of non-protein-bound pentalysine 2-carbonylphthalocyanine Zinc. Experimental results show that the cold acetone protein precipitation method was more suitable for measure the free concentration of pentalysine 2-carbonylphthalocyanine zinc in plasma. The linear range for pentalysine 2-carbonylphthalocyanine zinc were10~500nmol/L, r=0.9993; the extraction recoveries were 81.65% ~ 86.16%; plasma protein binding rate were 88.51%~92.80%. The cold acetone precipitation method is an ideal method to measure the free concentration of pentalysine 2-carbonylphthalocyanine zinc in plasma.In the third part, we established a method for determining the concentration of pentalysine 2-carbonylphthalocyanine zinc in mice plasma by HPLC. After a simple pretreatment, pentalysine 2-carbonylphthalocyanine zinc was analyzed on CNW C18 column (4.6 mm×250 mm, 5μm) and detected by fluorescence detector. The detection wavelengths were set at 610nm as excitation wave, 690nm as emission wave. The mobile phase consisted of methanol-water(containing 0.05% TFA, respectively) at a flow rate of 1.0 mL/min. Column temperature:25℃. The linear calibration curves were obtained in the concentration range of 2-500 nmol/L, (r=0.9990). The lowest limit of quantification was 0.5 nmol/L.The recoveries were 86.90 %-88.80 % in mice plasma,and the intra- and inter-day precision were less than 10% in all case. The HPLC method was specific, sensitive and accurate. And the method was satisfied for determining the concentration of pentalysine 2-carbonylphthalocyanine zinc in mice plasma. |
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