Shoichi Yada

Simultaneous Determination of Carboxyhemoglobin and Methemoglobin in Victims of Carbon Monoxide Poisoning

Concentrations of methemoglobin (Met-Hb) and carboxyhemoglobin (HbCO) were simultaneously determined in blood samples from the victims of city gas poisoning, fires, and exhaust fumes poisoning. All the samples contained concentrations of HbCO, although the concentrations for victims of city gas poisoning were significantly higher than those for victims of fires or exhaust fumes poisoning. Only negligible amounts of Met-Hb were detected in the samples from cases of city gas poisoning, while 4.9 to 31.6% of Met-Hb were found in all but one (1.8%) of the samples of the other two cases. Heat denaturation was considered to be the main cause of Met-Hb production in cases of fires, and inhalation of oxides of nitrogen in cases of exhaust fumes poisoning.

A practical method for the accurate determination of methemoglobin in blood containing carboxyhemoglobin

Kinetics of the oxidation of carboxyhemoglobin (HbCO) by potassium ferricyanide was studied photometrically in a weakly acid solution. An increase in the absorbance at 630 nm reached a maximum within 10 min when over a 100-fold excess of ferricyanide to hemoglobin iron was used. A slight decrease in the absorbance was observed after completion of the reaction when over a 500-fold excess of the reagent was used. In the presence of 0.4% Sterox SE, the absorbance began to decrease without complete oxidation. From these findings, a simple, rapid and accurate method for the determination of methemoglobin (Met-Hb) in blood was devised. The method was compared with two other methods, using 11 blood samples containing various amounts of HbCO, and proved to be suitable for blood containing elevated HbCO as well as for ordinary blood.

Forensic application of rapid analysis of carboxyhemoglobin in blood using an oxygen electrode.

Blood carboxyhemoglobin (HbCO) levels in forensic victims were measured by a new and rapid oxygen-electrode method for successive determinations of HHb, HbO2 and HbCO. It was found that the method is applicable to old denatured blood when combined with the cyanmethemoglobin method. Methemoglobin (Met-Hb) in blood was also estimated by subtracting the sum of HHb, HbO2 and HbCO from total hemoglobin (total Hb.) Blood Met-Hb levels in six victims of city gas poisoning were practically negligible, while those in five victims of fire were 13.5-26.3%.

A simple spectrophotometric method for the determination of carboxyhemoglobin in blood

The spectrophotometric method for the determination of carboxyhemoglobin (HbCO) in blood reported by Fretwurst and Meinecke was modified so as to give the same values of percentage HbCO (HbCO%) as those determined by the oxygen electrode method. Values of HbCO% of nine practical samples determined by both the oxygen electrode method and the present method were nearly identical regardless of the presence of methemoglobin (Met-Hb) in blood. The present method is suitable for forensic practice.

Detection of Thyroglobulin in Bloodstains as an Aid in the Diagnosis of Mechanical Asphyxia

Mechanical forces applied to the neck region are known to release certain amounts of thyroglobulin into circulation. In this experiment, an attempt was made to detect thyroglobulin in bloodstains as an aid in the diagnosis of mechanical asphyxia. Experimental bloodstains containing thyroglobulin at concentrations of 1, 2, 5 and 10 mu g/mL were prepared on a sheet of filter paper. Small pieces of bloodstains, measuring approximately 2.4 cm2 in area, were extracted with 0.1 mL of distilled water and the extracts were tested against an antihuman thyroglobulin serum by precipitation-electrophoresis. Bloodstains containing more than 1 mu g/mL of thyroglobulin formed distinct precipitin lines for up to one month of storage, while bloodstains containing more than 5 mu g/mL of thyroglobulin formed distinct precipitin lines for up to three months of storage. The present results suggest that the bloodstains can be utilized in the diagnosis of mechanical asphyxia.

Plasma Thyroglobulin as an Indicator of Mechanical Asphyxia—Comparison of Plasma Thyroglobulin Levels by Radioimmunoassay and the Results of Precipitation-Electrophoresis:

Thyroglobulin levels in the plasma of victims of mechanical asphyxia or other causes of death were determined by radioimmunoassay, and the results were compared with those obtained by precipitation-electrophoresis. The plasmas containing over 190 ng/0.1 ml of thyroglobulin showed positive reaction by the precipitation-electrophoresis and were restricted to the victims of mechanical asphyxia. Four out of 15 samples of mechanical asphyxia showed plasma levels of 29–170 ng/0.1 ml and 5 out of 15 levels of 0.10 ng/0.1 ml. Ten samples obtained from victims of other causes of death showed the levels of 0–10 ng/0.1 ml except for the two victims suspected of suffering a bruise on their neck region (144 and 116 ng/0.1 ml). The simple precipitation-electrophoresis technique was recommended as the supplementary test for the diagnosis of mechanical asphyxia.

Phosphoglucomutase1 (PGM1) and 6-phosphogluconate dehydrogenase (PGD) types in the human hairbulb

Phosphoglucomutase1 and 6-phosphogluconate dehydrogenase types present in human red cells were also found in human hairbulbs. These phenotypes were detectable up to 2 weeks following extraction of the hair. The results are useful in medicolegal individualization of the human hair.

Kinetic analysis of anaerobic metabolism in rats during acute cyanide poisoning

In order to investigate the changes in energy metabolism during acute anoxia, blood levels of various metabolites were analysed in cyanide-poisoned rats. After intraperitoneal injection of a sublethal dose of potassium cyanide (5 mg/kg), blood samples were obtained by cervical dislocation at intervals of 5 min until 30 min. Lactate and lactate/pyruvate ratio (L/P) in plasma concomitantly changed with cyanide; increased rapidly at 5 min, remained fairly constant until 20 min and then began to decrease at 25 min. In contrast, the products of ATP degradation, oxypurines and inorganic phosphate (Pi), increased gradually until 25 min and then began to decrease at 30 min. Allantoin in plasma scarcely increased throughout the experiments. The results indicate that the rapid activation of anaerobic ATP formation by glycolysis was followed by the increase in ATP degradation in cyanide-poisoned rats, Thus, increase in plasma oxypurines could be regarded as an indicator for severe anoxic states in tissues with massive ATP degradation.

A new reagent for the rapid determination of total hemoglobin as hemiglobincyanide in blood containing carboxyhemoglobin

Kinetics of conversion of HbCO to HiCN with various K3Fe(CN)6-KCN reagents was examined photometrically. Complete conversion of HbCO to HiCN required about 8 hr, 150 min, and 50 min with Drabkins reagent, that of van Kampen and Zijlstra, and that of Taylor and Miller, respectively. The absorbance at 540 nm was stabilized practically in 5 min even in CO-saturated blood when a reagent containing a 1600-fold excess of K3Fe(CN)6 to Hb iron was used. A slight increase in the absorbance was observed around 504 nm in the HiCN solution produced with the reagent. The absorbance at 540 nm in the solution, however, was stable until at least 120 min after mixing. From these findings, a reagent containing a 1600-fold excess of K3Fe(CN)6 to Hb iron and an incubation time of 5 min were used for the complete conversion of HbCO to HiCN. The stabilization of the absorbance at 540 nm was often prolonged to 10 min when the reagent was stored over 2 weeks. The reagent was compared with two other reagents, using 20 blood samples containing various amounts of HbCO, and proved to be suitable for the determination of total Hb in blood containing elevated HbCO as well as in ordinary blood.

Hyperuricemia in rats during acute carbon monoxide poisoning.

The effect of carbon monoxide (CO) inhalation on plasma levels of uric acid and hypoxanthine in rats was investigated. Exposure to 3% CO caused respiratory arrest within about 2 minutes. The plasma uric acid level of CO-treated rats increased to 157% above that of ether-treated rats. When rats were exposed to 1% or 0.8% CO, the exposure periods until the onset of respiratory arrest were prolonged, and plasma uric acid levels at respiratory arrest were further elevated. Plasma uric acid levels at respiratory arrest increased with prolongation of the exposure periods. Under our experimental conditions, hypoxanthine or xanthine was not detected in plasma of CO-treated rats. These results are discussed in relation to the hyperuricemia in hemorrhagic shock or hypoxemia: CO-induced hyperuricemia can be attributed to the stimulated degradation of adenine nucleotides under tissue anoxia, and thus could be an excellent parameter of tissue anoxia.