Species identification of blood and bloodstains by high-performance liquid chromatography

SummaryA reverse-phase high-performance liquid chromatographic method for species identification of blood and bloodstains is described. The method employs a 300 Å 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.ZusammenfassungBeschrieben wird eine „reversedphase” HPLC-Methode zur Identifizierung und Spezifizierung von Blut und Blutspuren. Die Analysenmethode besteht aus einer SynChropak RP-4-Säule und einem ternären Lösungsmittelgradientensystem (Acetonitril/Trifluoressigsäure/Wasser). Mit dieser Methode wird durch die Detektion des Häm-Peaks die Probe als Blut identifiziert und die Herkunft (human/tierisch) gleichzeitig durch charakteristische Chromatogramme UV-spektrometrisch spezifiziert. Deformationen der chromatographischen Profile wurden bei älteren Blutspuren beobachtet, wobei sich aber die Retentionzeiten der Hauptkomponenten nur geringfügig änderten. Der Speziesnachweis gelang bei bis zu drei Monate alten Blutspuren. Die vorgestellte Methode hat den Vorteil der Einfachheit, der Schnelligkeit und der Empfindlichkeit für die forensische Praxis.

[1]  M. Brunori,et al.  Purification and functional properties of the hemoglobin components from the rat (Wistar). , 1981, European journal of biochemistry.

[2]  BRYAN J. CULLIFORD,et al.  Precipitin Reactions in Forensic Problems: A New Method for Precipitin Reactions on Forensic Blood, Semen and Saliva Stains , 1964, Nature.

[3]  A. Kutlar,et al.  Separation of normal and abnormal hemoglobin chains by reversed-phase high-performance liquid chromatography. , 1986, Journal of chromatography.

[4]  D. Teplow,et al.  High performance liquid chromatographic separation of the globin chains of non-human hemoglobins. , 1985, Hemoglobin.

[5]  H. Inoue,et al.  Identification of fetal hemoglobin in blood stains by high performance liquid chromatography , 2004, Zeitschrift für Rechtsmedizin.

[6]  H C Lee,et al.  Simultaneous identification and determination of species origin, ABH antigens and isoenzyme markers in the same bloodstain. , 1985, Forensic science international.

[7]  J. Gilman,et al.  Rat hemoglobin heterogeneity: genetic variation affecting hemoglobin proportions. , 1982, Hemoglobin.

[8]  D. Stevenson,et al.  A Review of Analytical Methods , 1986 .

[9]  P. W. Harris-Smith,et al.  Species identification of blood and saliva stains by enzyme-linked immunoassay (ELISA) using monoclonal antibody. , 1984, Journal of forensic sciences.

[10]  M. Oshima,et al.  Identification of species specific hemoglobin by isoelectric focusing. , 1982, Forensic science international.

[11]  Precipitin Reactions in Forensic Problems: Precipitin Sera in Forensic Problems , 1964, Nature.

[12]  O. Takenaka,et al.  Novel hemoglobin components and their amino acid sequences from the crab-eating macaque (Macaca fascicularis) , 2005, Journal of Molecular Evolution.

[13]  W. Schroeder,et al.  High Performance Liquid Cbromatographic Separation of Globin Chains on a Large-Pore C4 Column , 1984 .

[14]  L. Kobilinsky,et al.  Development of a radioimmunoassay technique for the detection of human hemoglobin in dried bloodstains. , 1988, Journal of forensic sciences.

[15]  G. Springer,et al.  BLOOD GROUP ACTIVITY OF GRAM-NEGATIVE BACTERIA , 1961, The Journal of experimental medicine.