| The determination of time of death, or postmortem interval (PMI), is a matter of crucial importance in criminal investigations. Nowadays, its determination usually depends on theses indicators such as insect community succession on carrion, insect morphological changes and development span. Using those indicators, however, the growth and development stage could not be assessed accurately at some cases. For example, the larval body length and morphological characteristics of most necrophagous flies is difficult to be discerned as larvae are at the post-feeding stage, easily resulting in the determination of PMI. Indeed, changes in the composition and levels of some biochemical substances usually performed some regular patterns during insect growth and development. If those regular patterns are significantly associated with insect age, those biochemical substances, which can be rapidly measured by the suitable apparatus with standard methods, as the age indicators may supplement the shortage of the previous indicators so as to minimize the man-made error. Therefore, changes in compositions and levels of several biochemical substances, including cuticular hydrocarbon, pteridine, protein and uric acid during growth and development of some common species of necrophagous flies were measured and analyzed, and then the potential of using change patterns in those biochemical substances as age indicators in necrophagous flies was discussed. In addition, the application of cuticular hydrocarbon analysis to the taxonomic differentiation of the pupae and empty exuviae of six necrophagous flies, were investigated. The results are summarized as follows:1 Time-associated characteristics in cuticular hydrocarbons of the larvae from six necrophagous fliesGas chromatography (GC) and GC-mass spectrometry (GC-MS) was used to measure the time-associated, quantitative changes in cuticular hydrocarbons of the larvae from six necrophagous files, i.e. Aldrichina grahami, Chrysomya megacephala, Lucilia sericata, Achoetandrus rufifacies, Boettcherisca peregrina and Parasarcophaga crassipalpis. Some cuticular hydrocarbons suitable for age determination of the larvae were identified through stepwise multiple regression analysis of the relationship between relative abundance of different hydrocarbons and larval development time, which were gas chromatographic peak 14 (unknown), 33 (n-Pentacosane), 49 (Heptacosene b) and 79 (n-Hentriacontane) for Al. grahami, peak 2 (unknown) and 54 (n-Hentriacontane) for C. megacephala, peak 8 (Docosene ), 34 (3-Methyl-pentacosane) and 56 (2-Methyl-octacosane) for L. sericata, peak 15 (5-Methyl-tricosane), 41 (n-Octacosane) and 45 (n-Nonacosane) for Ac. rufifacies,peak 16(unknown) and 25 (11-, 13-, 15-Methyl-hentriacontane) for B. peregrina and peak 3 (unknown), 4 (Tricosene) and 23 (unknown) for P. crassipalpis. Additionally, the models were developed respectively to determine the larval age of different flies accurately.2 Temperature effects of time-associated characteristic in cuticular hydrocarbons of Aldrichina grahami larvaHighly significant correlation were observed between the relative abundance of some larval cuticular hydrocarbons associated with gas chromatographic peaks and the chronological age of Al. grahami larvae. Those gas chromatographic peaks included p30 (Pentacosene a^ p33 (n-Pentacosane), p48 (Heptacosene a) p49 (Heptacosene b) and p79 (n-Hentriacontane). Quantitative changes over time of these cuticular hydrocarbons were similar at different constant temperatures such as 16, 20, 24 and 28 . The temperature-depended model using cuticular hydrocarbons associated with gas chromatographic peaks 30, 33, 48, 49 and 79 as the variables, i.e. D=217.23/(T+1.54) x(l-2.264xjep3o-0.8632xxp33-2.454xxp48-6.619xxp49-1.948xjep79), was developed to promise for larval age grading of A. grahami accurately (R2=0.9118).3 Cuticular hydrocarbon composition in the pupae and its application in taxonomic differentiation for six necrophagous fliesGC-patterns and their time-assoc...
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