井上 博之

The anaphylatoxin C3a or its desArg form (C3a/desArg) generated during complement activation could be detected in the vicinity of incised skin wounds of guinea pigs using immunoblotting methods. The C3a/desArg peptides were detectable immediately after injury in local sites up to 3 mm from the wound edge. In subsequent determinations of up to at least 3-day-old antemortem wounds, the maximum concentration of these peptides was largely localized up to 6 mm from the wound edge at 2 h after injury. In postmortem wounds, however, these peptides were undetectable. When they were released in antemortem wounded tissues they could be detected up to 1 day at 22-degrees-C after death. These results suggest that the detection of C3a/desArg in wounds using immunoblotting methods can be useful for distinguishing ante- from postmortem wounds.

In order to examine the vital reaction in wounds, catecholamines, serotonin (5-HT) and their metabolites in the incised skin wounds of guinea pigs were analyzed simultaneously by high-performance liquid chromatography (HPLC) using electrochemical detection (ECD). The principal changes in the levels of these compounds in vital wounds were as follows: a considerable decrease in norepinephrine (NE) content was observed 12 - 24 h after injury which persisted to at least 7 days. 3,4-Dihydroxyphenylacetic acid (DOPAC) decreased slightly for up to 30 min and then showed no significant difference compared with postmortem levels. Epinephrine and dopamine were barely detected by the HPLC-ECD method employed. 5-HT concentrations which showed an increase up to 24 h showed maximum levels 800-mu-m from the wound edge at 10 and 30 min after injury. 5-Hydroxyindoleacetic acid (5-HIAA) was significantly higher than the postmortem level over almost the entire period of these experiments. A 5-HIAA content of at least twice the postmortem level was observed 800-mu-m from the wound edge of a 10-min-old vital wound. Therefore, 5-HIAA is a likely candidate as a new marker for evaluating the vital reaction in wounds. The vital characteristics of NE, DOPAC, 5-HIAA and 5-HT in 10-min-old wounds persisted for up to 12 h at room temperature after death. These results suggest that the HPLC-ECD method used here is very useful for simultaneous examination of the vital reaction in wounds from the earliest to the later stages of the wound-healing period.

A method using reverse-phase high-performance liquid chromatography (HPLC) for the identification of fetal hemoglobin (Hb F) and the simulataneous estimation of bloodstain age is described. Umbilical cord and neonatal bloodstains can be differentiated from adult stains by the presence of gamma-globin chains which are characteristic of Hb F. With this method, cord and neonatal blood could be distinguished from adult blood in stains up to 32 weeks old. The age of the stain was estimated from the ratio of the peak area of the alpha-globin chain to that of heme on the same chromatogram. The ratio decreased gradually with an increase in the age of the stain up to 20 weeks old. Studies performed at each time period revealed no significant difference in the ratios of cord and neonatal bloodstains or in the ratios of cord and adult stains. The regression equation calculated from the ratios (y) and the ages of stains in weeks (x) expressed logarithmically is y = 2.5758 - 0.2497 ln (x) and the coefficient of correlation is -0.7491 (n = 252, P < 0.001). The present method, having the advantages of simplicity, speed and sensitivity, should be of great value to forensic science.

A reverse-phase high-performance liquid chromatographic method for species identification of blood and bloodstains is described. The method employs a 300 angstrom pore SynChropak RP-4 column and ternary solvents (acetonitrile-trifluoroacetic acid-water) and can not only identify a species by its characteristic chromatogram, but also simultaneously demonstrates that it is of blood origin by the existence of the heme peak. Deformations in chromatographic profiles obtained with older bloodstains were observed, but the retention times of heme and the major peaks showed only minor changes. The species could be identified from bloodstains at least 3 months old and the present method has the advantage of simplicity, speed and sensitivity in the practice of forensic science.