Prognostic role of immunosuppressive acidic protein in patients with esophageal cancer.

Immunosuppressive acidic protein (IAP) suppresses several immune responses in vivo and in vitro , and high preoperative IAP levels could predict the impairment of the host's immunity. In this study prognostic significance of preoperative IAP levels was investigated in 68 esophageal cancer patients with curative resection and eight with non-curative resection. The curative group had significantly lower levels than the non-curative group (432 +/- 183 mg/mL vs. 739 +/- 235 mg/mL, P < 0.0001). The IAP levels were associated with T-status (P < 0.0001), lymphatic invasion (P < 0.05), and p-stages (P < 0.0001). When 5-year survival rate of patients with curative resection was compared by setting various cutoff values of IAP between high and low IAP groups, several cutoff points (400-580 mg/mL) were revealed to be significantly associated with survival. Setting cutoff value of IAP to 560 mg/mL resulted in a most significant difference of 5-year survival rate of patients between the high and low IAP groups (13.9% and 61.5%, P < 0.0001). These data indicate that pre-operative IAP level is a useful parameter to predict the prognosis of esophageal cancer patients after curative resection.

Identification of human urine stains by HPLC analysis of 17-ketosteroid conjugates.

A new method for identifying human urine stains utilizing high-performance liquid chromatographic (HPLC) analysis of five major 17-ketosteroid conjugates: dehydroepiandrosterone sulfate, etiocholanolone sulfate, etiocholanolone glucuronide, androsterone sulfate, and androsterone glucuronide was examined. Samples of urine stains were extracted with borate buffer solution (pH 9.3) and the extracts were applied onto a Sep-Pak tC18 cartridge. The analytes were eluted from the cartridge with methanol. The eluates were prelabeled with 2,4-dinitrophenylhydrazine in trichloroacetic acid-benzene solution and were separated by HPLC on a reversed-phase ODS column using a mobile phase of 80% methanol in a buffer consisting of 25 mM sodium acetate in 2% acetic acid. The eluates were monitored by a spectrophotometer at 380 nm. While all five 17-ketosteroid conjugates were clearly detected in the human urine stain samples, traces of only some of these conjugates were detected in the animal samples. Therefore, the presence of all five 17-ketosteroid conjugates indicated human specificity. In addition to the above finding, the properties of those five 17-ketosteroid conjugates were confirmed by electrospray ionization liquid chromatography-mass spectrometry (ESI-LC-MS).

Multiplex amplified product-length polymorphism analysis for rapid detection of human mitochondrial DNA variations.

A number of mutations in coding and noncoding regions of mitochondrial DNA (mtDNA) have previously been studied. In the present study, we simultaneously typed six mutation sites in the coding region by use of amplified product-length polymorphism (APLP) analysis. The mtDNA variations of 2471 individuals from 20 populations of Japanese, Korean, Chinese, and German were examined and classified into 18 haplotypes. Two of these haplotypes, B1 (estimated ancestral haplotype) and C1, were distributed among all populations tested. However, the haplotypes A1, A2, B2, B3, and C2 were mostly restricted to the Mongoloid populations, whereas haplotypes B5 and C5 appeared almost exclusively in the German population. Phylogenetic analysis by the neighbor-joining method revealed that the Japanese populations were more closely related to each other than to the other East Asian populations surveyed. The multiplex APLP method is suitable for large-scale screening studies of mtDNA variability because it is both rapid and economical.

Absolute configuration of (6S,9S)-roseoside from Polygonum hydropiper.

From Polygonum hydropiper L., a C13-norisoprenoid glucoside was isolated and its absolute configuration was established to be (6S,9S)-roseoside (1) by spectroscopic evidence and X-ray crystallographic analysis of its acetate derivative (2). In addition, the stereostructure of roseoside from Canthium subcordatum was revised to the (6S,9S) configuration.

New method of derivatization and headspace solid-phase microextraction for gas chromatographic-mass spectrometric analysis of amphetamines in hair.

A simple method for hair analysis of methamphetamine (MAMP) and amphetamine (AMP) by gas chromatography-mass spectrometry (GC-MS) was developed using simultaneous headspace solid-phase microextraction (HS-SPME) with derivatization. After alkaline-digestion of hair, the analytes derivatized with heptafluoro-n-butyryl chloride were adsorbed on a polydimethylsiloxane-coated fiber by HS-SPME and analyzed by GC-MS. Their mass spectra were, respectively, observable at 1 ng per sample. The standard curves in the range of 0.1-100 ng were linear. The intra-day coefficients of variation at each 0.5 ng were 12.5% for AMP and 3.8% for MAMP. The applicability of this method was demonstrated in some case studies.

Death following cupric sulfate emesis.

CASE HISTORY: A 25-year-old woman who had ingested about 20 tablets of diazepam 2.5 mg in a suicide attempt was given cupric sulfate 2.5 g in 1750 mL water as an emetic, but died 3 days later. On autopsy, death was attributed to acute hemolysis and acute renal failure due to copper poisoning. Copper concentrations were 5.31 microg/mL in whole blood, 19.0 microg/g in the liver, 8.9 microg/g in the kidney, 1.1 microg/L in the brain, 1.1 microg/g in the gastric wall, 1.5 microg/g in the jejunal wall, 0.3 microg/g in the colon wall, 4.6 microg/g in the gastric contents, and 12.6 microg/g in the intestinal contents (fresh weight). This case and 10 others from the Chinese medical literature provide additional evidence that cupric sulfate is a corrosive poison and contraindicated as an emetic.

[New development of hair examination! The searching of hair for the terminal residual substance of pleasant sensation].

Formerly, hair had been analyzed mainly in the field of individual identification. Otherwise, it is known that many substances are accumulated in hair such as metals like arsenic, mercury, alkaloids like caffeine, theophylline, drugs like opiate, stimulants, other drugs and poisons. So, by qualitative and quantitative analysis of such deposits in hair. It is possible to presume one's life manner during recent several months. This is the new development of hair analysis and it is expected that it may be wide of application. The main theme of this paper is hedonics. Nobody dislike hedonic sensation. So, it can be said everybody being living always coveting hedonic sensation. There are many hedonic sensers all over the body, and hedonic sensation transmitted by A10 nerve, so A10 nerve is called the hedonic nerve. Neurotransmitter of this A10 nerve is Dopamine. Dopamine is the endogenous hedonic material. Exogeneous hedonic one is stimulant like Methamphetamine. In this paper, relation between Dopamine, Methamphetamine and Tyramine, thought to be the terminal product from two formers is examined in the aspect of hair analysis. Catecholamines including Dopamine are made of Tyrosine. They have a Benzene-ring. Compounds with a Benzene-ring made of Tyrosine are relatively rare, but all of them have very strong action. Tyramine is made of Tyrosine, too. So the structure of Tyramine is essentially resemble to Dopamine, and the skeletal structure of Methamphetamine is identical to Dopamine, so the action is identical, too. Normally, excessive release of Dopamine in brain is controlled by the negative feedback of GABA system. But extrinsic stimulus as Methamphetamine put balance of the homeostasis between Dopamine system and GABA system in to disorder, and long term administration of Methamphetamine makes long remaining degeneration of synapse of Dopamine nerve. So, abnormal state of excess releasing of Dopamine happens easily. And the morbid state named stimulant psychosis occur. The volume of Tyramine in hair changes correlatively with the volume of released Dopamine in brain. Therefore it is suggested that quantitative analysis of Tyramine in hair enable to estimate the changes of Dopamine released in brain.

Headspace solid-phase microextraction and gas chromatographic-mass spectrometric screening for volatile hydrocarbons in blood.

Optimization for headspace solid-phase microextraction (SPME) was studied with a view to performing gas chromatographic-mass spectrometric (GC-MS) screening of volatile hydrocarbons (VHCs) in blood. Twenty hydrocarbons comprising aliphatic hydrocarbons ranging from n-hexane to n-tridecane, and aromatic hydrocarbons ranging from benzene to trimethylbenzenes were used in this study. This method can be used for examining a burned body to ascertain whether the victim had been alive or not when the burning incident took place. n-Hexane, n-heptane and benzene, the main indicators of gasoline components, were found as detectable peaks through the use of cryogenic oven trapping upon SPME injection into a GC-MS instrument. The optimal screening procedure was performed as follows. The analytes in the headspace of 0.2 g of blood mixed with 0.8 ml of water plus 0.2 microg of toluene-d8 at -5 degrees C were adsorbed to a 100-microm polydimethylsiloxane (PDMS) fiber for 30 min, and measured using the full-mass-scanning GC-MS method. The lower detection limits of all the compounds were 0.01 microg per 1 g of blood. Linearities (r2) within the range 0.01 to 4 microg per 1 g of blood were only obtained for the aromatic hydrocarbons at between 0.9638 (pseudocumene) and 0.9994 (toluene), but not for aliphatic hydrocarbons at between 0.9392 (n-tridecane) and 0.9935 (n-hexane). The coefficients of variation at 0.2 microg/g were less than 8.6% (n-undecane). In conclusion, this method is feasible for the screening of volatile hydrocarbons from blood in forensic medicine.

[A patient with multiple pulmonary metastases from colonic cancer who survived for 5 years by effectual administration of 5'-DFUR].

The patient was a 69-year-old woman who had been diagnosed with a bowel obstruction due to colonic cancer, with simultaneous multiple pulmonary metastases. The primary lesion was resected and 5'-DFUR was administered for 2 years at an out-patient clinic. During those 2 years, there was no change in CEA value and the pulmonary lesions were fading on the roentogenograms. It then became doubtful whether the pulmonary shadows were real metastases or not, and 5'-DFUR administration was discontinued. After stopping the medication, her CEA value rose and the tumor shadows increased in intensity. 5'-DFUR was therefore re-administrated and her CEA value declined. Afterwards, a re-elevation in CEA value was seen, and low-dose FP therapy was added on an out-patient basis. Anti-cancer chemotherapy of 5'-DFUR (oral) and low-dose FP (i.v.) was contributed to her 5-year survival.

A case of eosinophilic pleural effusion induced by pancreatothoracic fistula.

A 49-year-old man was admitted for evaluation of a left pleural effusion. Thoracenthesis yielded a hemorrhagic pleural effusion with a high percentage of eosinophils (15.9%). Although there were no significant abdominal signs, serological examinations demonstrated a marked increase of pancreatic enzyme activity. Moreover, abdominal CT demonstrated cystic changes between the tail of the pancreas and the spleen. Accordingly ERP was performed under pressure, and contrast medium draining from the pancreas was observed. Pancreatic pleural effusion in this patient consisted of pancreatic juice retained in the thoracic cavity, which resulted from intrapancreatic fistulation connecting to the thoracic cavity due to a pancreatic cyst caused by chronic pancreatitis. The present report indicates that we should investigate the retention of eosinophilic pleural effusion considering not only the possibility of thoracic disease, but also the possibility of a pleural effusion derived from abdominal diseases.

Gas chromatographic-mass spectrometric analysis of dichlorobenzene isomers in human blood with headspace solid-phase microextraction.

Headspace solid-phase microextraction (HS-SPME) was utilized for the determination of three dichlorobenzene isomers (DCBs) in human blood. In the headspace at 30 degrees C, DCBs were absorbed for 15 min by a 100-micron polydimethylsiloxane (PDMS) fiber. They were then analyzed by capillary column gas chromatography-mass spectrometry (GC-MS). By setting the initial column oven temperature at 20 degrees C, the three isomers were resolved at the baseline level. p-Xylene-d10 was used as the internal standard (I.S.). For quantitation, the molecular ion at m/z 146 for each isomer and the molecular ion at m/z 116 for I.S. were selected. For day-to-day precision, relative standard deviations in the range 3.2-10.7% were found at blood concentrations of 1.0 and 10 micrograms/ml. Each compound was detectable at a level of at least 0.02 microgram per 1 g of whole blood (by full mass scanning). HS-SPME-GC-MS, when performed at relatively low temperatures, was found to be feasible in toxicological laboratories. Using this method, the plasma levels of one patient who had drunk a pesticide-like material were measured.

Practical GC/MS analysis of oxidation dye components in hair fiber as a forensic investigative procedure.

The purpose of this study was to improve the reliability of discrimination (or identification) of dyed hair by analyzing chemical substances present in the hair, as an addition to the conventional macroscopical and microscopical examinations and ABO blood group examination. Oxidation hair-dye components such as p-phenylenediamine (PPDA), toluylene-2,5-diamine (T-2,5-DA), o-aminophenol (OAP), m-aminophenol (MAP), p-aminophenol (PAP) and p-amino-o-cresol (PAOC) were selected as the object of study. After alkaline-digestion, hair samples were adjusted to a pH of 12.6 to 12.8, and applied onto an Extrelut column. After 15 min, the components were simultaneously extracted and derivatized with n-hexane including 1% heptafluoro-n-butyryl (HFB) chloride. Their HFB derivatives within a condensed sample were diluted in ethyl acetate, and analyzed by gas chromatography-mass spectrometry (GC-MS) with full mass scanning or selected ion monitoring. For estimating their levels, 2,4,6-trimethylaniline was used as the internal standard. Standard curves obtained by preparing a 5 cm segment of control hair spiked with authentic substances were linear, ranging from 0.1 to 4.0 micrograms for PPDA and T-2,5-DA, and from 0.01 to 0.4 microgram for OAP, MAP, PAP and PAOC. The coefficient of variation of inter-day precisions (each n = 4) was below 16% for PPDA, below 20% for OAP and PAP and below 24% for T-2,5-DA, MAP and PAOC. These components were detectable even at 6 ng for PPDA, T-2,5-DA, MAP and PAP, 8 ng for OAP, and 4 ng for PAOC. Recovery percents using this procedure ranged from 54 to 86%. By using actual dyed hair samples this method was shown to provide high sensitivity for accurate detection.

Rupture of a giant middle cerebral aneurysm after blunt head injury.

A 20-year-old man was involved in a fight while under the influence of alcohol. He subsequently developed an intracerebral haemorrhage in the putamen with a minimal subarachnoid haemorrhage and died. An autopsy revealed a ruptured giant saccular aneurysm of the middle cerebral artery. We believe that the blunt head injury caused a rupture of the pre-existing giant saccular aneurysm either through the force transmitted from the blow, or by a possible transient elevation of the blood pressure secondary to the excitement of the fight, or by a combination of both factors.

Utility of caffeine analysis for forensic hair discrimination.

Utility of caffeine analysis for the purpose of forensic hair discrimination was evaluated. Methanol extracts of each 5-cm strand of scalp hair, after its incubation at 37 degrees C for 15 h, was analyzed by gas chromatography-mass spectrometry (GC-MS). For quantitative analysis of caffeine, caffeine-d3 was added as internal standard to the methanol extract. Hair specimens were obtained from three victims and from twenty-two volunteers. The results showed that caffeine levels in hair varied according to each individual; caffeine was detected up to a maximum level of 4.19 ng/5-cm hair. The concentrations of caffeine in hair obtained from four different regions of the scalp in twenty-two volunteers showed a coefficient of variation not greater than 24.7% for each individual. Based on our data, it was concluded that caffeine levels give relatively high reliability for discriminating a victim's hair from others at a criminal or accidental scene, although it should be used in combination with morphological examination and ABO blood group determination.

Simple extractive derivatization of methamphetamine and its metabolites in biological materials with Extrelut columns for their GC-MS determination.

A simple, extractive heptafluoro-n-butyrylation with Extrelut columns was devised to simultaneously measure methamphetamine (MAMP), amphetamine (AMP), 4-hydroxymethamphetamine (HMAMP), and 4-hydroxyamphetamine (HAMP) in biological materials by gas chromatography-mass spectrometry (GC-MS) using 4-methoxymethamphetamine-d5 as the internal standard. Human urine, human whole blood, and porcine skeletal muscle spiked with the stimulant standards were used for evaluating the method. After deproteinization and adjustment of the pH to 12.6, the sample was applied to an Extrelut column. Using the present method, AMP, MAMP, and HMAMP could be determined in an actual forensic case study.

The utility of volatile hydrocarbon analysis in cases of carbon monoxide poisoning.

A new approach to investigate the circumstances relating to carbon monoxide intoxication by analysing volatile hydrocarbons in the blood of cadavers is reported. Headspace gas chromatography/mass spectrometry was used to demonstrate the hydrocarbons. The results can be characterized into four categories depending on the compounds detected. In construction fire cases where no accelerants were found at the scene benzene, toluene and styrene were detected in the blood. In cases where gasoline was found in the fire debris surrounding the victim, high levels of benzene, toluene, ethylbenzene, xylene isomers, n-hexane and n-heptane were detected in the blood. In cases where kerosene was found in the fire debris around the victim, benzene, toluene, ethylbenzene, xylene isomers, C9-aromatics(n-propylbenzene, trimethyl-benzene isomers), n-octane, n-nonane and n-decane were detected in the blood. In cases where the victim was found inside a gasoline-fuelled automobile filled with exhaust gas, benzene, toluene, ethylbenzene, xylene isomers, C9-aromatics were found, but no aliphatic hydrocarbons such as components of petroleum. The analyses of the combustion gases of inflammable materials, exhaust gas, gasoline vapours and kerosene vapours were also performed to evaluate the results of the blood analyses. Consequently, some compounds are proposed as indicators to discriminate between inhaled gases i.e. styrene in common combustion gas, n-hexane and n-heptane as well as benzene, toluene and C9-aromatics in gasoline cases, n-nonane and n-octane as well as benzene, toluene and C9-aromatics in kerosene cases, and benzene, toluene, C9-aromatics but no aliphatic hydrocarbons in exhaust gas cases.

Identification of vegetable species in gastric contents using HPLC.

Identification of 16 vegetables, focusing on the influence of digestion in the stomach, was carried out on the basis of the types of flavonoids detected on chromatograms using HPLC. Among the 12 vegetables for which flavonoids were detected, the chromatographic patterns of the flavonoids in digested vegetables were the same as those of the corresponding raw vegetables, making it possible to identify the species of vegetables even after digestion. In our analysis, 5 mg of a freeze-dried sample was found be an adequate quantity to enable the detection of flavonoids. Brief practical cases are also described.

Brain death diagnosed by forensic analysis of drug distribution in human tissues.

Brain death was diagnosed by analyzing drug distribution in human tissues. Concentrations of diazepam and phenobarbital in human tissues, administered therapeutically to 3 patients in a critical condition, were determined by gas chromatography with a nitrogen phosphorus detector and gas chromatography-mass spectrometry. In 2 patients administered drugs while the brain was functioning, drug concentrations in the brain were high compared with those in the other tissues. In the other patient given the drugs after vital signs of the brain ceased, the level of drugs in the brain was much lower than seen in other tissues. We propose that a forensic diagnosis of patients who are brain dead can be made by comparing the concentrations of drugs in the brain given at the time of critical condition with concentration of these agents in other tissues. When cerebral function ceased can be roughly estimated by the variable distribution patterns of drugs and time of administration of these drugs.

Highly sensitive and rapid determination of theophylline, theobromine and caffeine in human plasma and urine by gradient capillary high-performance liquid chromatography-frit-fast atom bombardment mass spectrometry.

A sensitive and reliable analytical procedure has been established for the detection of theophylline (TH), theobromine (TB) and caffeine (CA) in human plasma and urine by gradient capillary high-performance liquid chromatography (HPLC)-frit-fast atom bombardment mass spectrometry (FAB-MS) (LC-frit-FAB-MS). Two capillary columns and a column-switching valve were used in this LC system to allow all of the sample injected to be introduced into the MS system. 7-Ethyltheophylline was used as the internal standard (I.S.). The xanthines in the specimen were extracted with an Extrelut column. The lowest detected amount was ca. 5 ng/ml using this method.

[Development of the method for analysis of theophylline and its related compounds using capillary high-performance liquid chromatography/fast atom bombardment-mass spectrometry].

A convenient, reliable and sensitive method for analysis of theophylline (TH), theobromine (TB) and caffeine (CA) in biological samples by capillary high performance liquid chromatography (HPLC)/fast atom bombardment (FAB)-mass spectrometry (MS) was developed using 7-ethyltheophylline (7-ETH) as an internal standard (IS). The capillary column (0.3 mm i.d.) enabled the introduction of entire effluent to the frit interface for FAB-MS; and a special column switching device was utilized for sample concentration and gradient formation. These conditions realized simultaneous determination of the compounds of as low as 50-500 ng/ml concentrations. Actually, the compounds in blood and urine could be identified and quantified for healthy subjects having TH and/or CA spontaneously. In addition, TH in human hair obtained from the patients with bronchial asthma, who had taken 400 mg TH daily for more than 2 years, could be detected by the capillary HPLC/FAB-MS. This method seems very useful for the analysis of clinical or forensic samples, because of its ability of final drug identification.