Antemortem Blood Research Articles

A hydrophilic interaction liquid chromatography electrospray tandem mass spectrometry method for the simultaneous determination of γ-hydroxybutyrate and its precursors in forensic whole blood

A liquid-chromatography–tandem-mass-spectrometry method using pneumatically assisted electrospray ionisation (LC–ESI-MS/MS) was developed for the simultaneous determination of γ-hydroxybutyric acid (GHB), γ-butyrolactone (GBL) and 1,4-butanediol (1,4-BD) in human ante-mortem and post-mortem whole blood. The blood proteins were precipitated using a mixture of methanol and acetonitrile, and the extract was cleaned-up by passage through a polymeric strong cation exchange sorbent. Separation of the analytes and their structural isomers was obtained using a column with a zwitterionic stationary phase. Matrix-matched calibrants, combined with isotope dilution, were used for quantitative analysis. GHB was determined in both positive and negative ion modes. The relative intra-laboratory reproducibility standard deviations were better than 10% and 6% for blood samples at concentrations of 2mg/L and 20–150mg/L, respectively. The mean true extraction recoveries were 80% for GHB and greater than 90% for GBL and 1,4-BD at concentration levels of 20–50mg/L. The limits of detection were approximately 0.5mg/L for GHB and GBL, and 0.02mg/L for 1,4-BD in ante-mortem blood. The corresponding lower limits of quantification were less than 1mg/L for GHB and GBL, and less than 0.1mg/L for 1,4-BD. GBL was unstable in whole blood freshly preserved with a sodium fluoride oxalate mixture, but the stability could be improved significantly by preservation with a sodium fluoride citrate EDTA mixture.

Postmortem Redistribution of Fentanyl in the Rabbit Blood

Postmortem redistribution of fentanyl in the rabbit was investigated after application of the 50-μg/h Durogesic pain patch. Patches were applied for 48 hours. Two cycles of patch administration were used before characterization of the postmortem redistribution. Fentanyl showed marked redistribution into the femoral and pulmonary veins of the rabbit through 48 hours after the animals were humanely killed and the pain patches removed. The plasma concentration of 2.34 ng/mL in the femoral blood before killing the animals increased 5.6-fold by 48 hours after patch removal to 13.2 ng/mL. This postmortem concentration is approximately 3-fold the C(max) determined during antemortem pharmacokinetic analysis, 4 ng/mL, which was achieved 24 hours after the application of the second 50-μg/h Durogesic pain patch. After blood sampling for 48 hours after animal termination with patch removal compared with sampling for 48 hours from animals not terminated and with patch removal, the exposure ratios in the terminated animals were approximately 30-fold, indicating that between the postmortem redistribution of fentanyl and the cessation of hepatic clearance of fentanyl in the rabbit, the postmortem redistribution of fentanyl leads to an elevated measures of postmortem blood concentrations relative to antemortem blood concentrations.

A Liquid Chromatography-Electrospray Tandem Mass Spectrometry Method for the Determination of Antiarrhythmic Drugs and Their Metabolites in Forensic Whole Blood Samples

A liquid chromatography-tandem mass spectrometry method, using pneumatically assisted electrospray ionization was developed for the determination of amiodarone, desethylamiodarone, propafenone, N-depropylpropafenone, 5-OH-propafenone, flecainide, and sotalol in human antemortem and postmortem whole blood. A mixture of methanol and acetonitrile was used to extract the samples, and the clear extracts obtained from the protein precipitation were injected directly onto an ethyl-linked phenyl LC column. The isotope dilution technique was applied for quantitative analysis. The relative intralaboratory reproducibility standard deviations were 5-9% at a concentration range of 1-5 mg/L and 9-12% at a concentration of 0.1 mg/L. The mean true recoveries were greater than 91% in the concentration range of 0.05-5 mg/L. The detection limits were in the range of 8-18 µg/L.

Is the formula of Traub still up to date in antemortem blood glucose level estimation?

According to the hypothesis of Traub, also known as the 'formula of Traub', postmortem values of glucose and lactate found in the cerebrospinal fluid or vitreous humor are considered indicators of antemortem blood glucose levels. However, because the lactate concentration increases in the vitreous and cerebrospinal fluid after death, some authors postulated that using the sum value to estimate antemortem blood glucose levels could lead to an overestimation of the cases of glucose metabolic disorders with fatal outcomes, such as diabetic ketoacidosis. The aim of our study, performed on 470 consecutive forensic cases, was to ascertain the advantages of the sum value to estimate antemortem blood glucose concentrations and, consequently, to rule out fatal diabetic ketoacidosis as the cause of death. Other biochemical parameters, such as blood 3-beta-hydroxybutyrate, acetoacetate, acetone, glycated haemoglobin and urine glucose levels, were also determined. In addition, postmortem native CT scan, autopsy, histology, neuropathology and toxicology were performed to confirm diabetic ketoacidosis as the cause of death. According to our results, the sum value does not add any further information for the estimation of antemortem blood glucose concentration. The vitreous glucose concentration appears to be the most reliable marker to estimate antemortem hyperglycaemia and, along with the determination of other biochemical markers (such as blood acetone and 3-beta-hydroxybutyrate, urine glucose and glycated haemoglobin), to confirm diabetic ketoacidosis as the cause of death.

Postmortem Drug Redistribution

Postmortem drug concentrations in blood may not always reflect antemortem drug concentrations in blood due to the movement of the drugs after death. The phenomenon is referred to as Postmortem Redistribution (PMR). Mechanisms involved in postmortem redistribution are both complicated and poorly understood. However, postmortem drug concentrations in blood do follow some generally accepted trends that aid with interpretation; the characteristics of the drug itself can be used to indicate if a drug is subject to PMR. Large changes in blood drug concentrations are predicted for basic, lipophilic drugs with a high volume of distribution (>3L/kg). When PMR occurs, blood specimens drawn from the central body cavity and heart generally will have higher drug concentrations postmortem than specimens drawn from peripheral areas, most commonly the femoral vein. The diffusion of drugs from organ tissue into the blood may explain the observed phenomenon [1]. To compensate for PMR, postmortem blood specimens are recommended to be collected from at least two areas of the body at autopsy; a peripheral area and a central area (often the heart), so that a comparison can be made. While antemortem blood specimens are especially helpful in interpreting postmortem blood drug concentrations, relevant specimens are only very rarely available. In a set of case studies of six drugs, drug concentrations in postmortem femoral blood specimens always exceeded the antemortem concentrations in five cases, suggesting that even peripheral blood may still exhibit some redistribution [2]. The study did not, however, report the postmortem interval been death and autopsy. It appeared that a partial answer to the difficulties associated with interpretation of postmortem drug concentrations was provided by two papers published in the 1990s. The first provided detailed information about blood drug concentrations attained from different sites for over fifty drugs [3]. The second provided a tabular list of the drug concentrations from both cardiac and peripheral blood samples expressed as a ratio of cardiac to peripheral blood (C/P) for over one hundred drugs [4]. The C/P ratio became the accepted benchmark with the accepted guideline that “high ratios” were associated with “potential for redistribution” [4]. This guideline was repeated in a review published a few years later that republished the C/P ratios for many of the drugs included in the Dalpe-Scott and coworker’s paper

A 2‐YEAR‐OLD BOY WITH HEMOLYTIC UREMIC SYNDROME AND PNEUMOCEPHALUS

Clostridium septicum infection following hemolytic uremic syndrome is rare and carries a poor prognosis, especially when the brain is involved. We report a case of a previously healthy 2-year-old boy who presented with two days of anuria and bloody diarrhea. He was admitted to the local children's hospital with a diagnosis of hemolytic uremic syndrome, presumably secondary to E. coli O157. He soon required intubation and was noted to have fixed and dilated pupils. Head CT revealed left frontal subcortical white matter vasogenic edema and scattered pockets of pneumocephalus. The patient expired 14 hours after admission. Antemortem blood cultures grew C. septicum. Gross pathologic examination of the brain revealed a large intraparenchymal cerebral hemorrhage in the left frontal and parietal lobes. There was extensive cystic changes as well. Microscopic examination revealed vacuolization and diffuse colonization with rod-shaped bacteria, but without the expected tissue response. There have been only six previously reported cases of C. septicum infection following hemolytic uremic syndrome, four of which had brain involvement. Mortality rate is high, with the only known survivor among those with brain involvement having a brain abscess rather than diffuse pneumocephalus.

Determination of benzodiazepines in ante-mortem and post-mortem whole blood by solid-supported liquid–liquid extraction and UPLC–MS/MS

A solid-supported liquid–liquid extraction ultra performance liquid chromatography tandem mass spectrometry (UPLC–MS/MS) method was developed and validated for the determination of benzodiazepines commonly found in Norway, for use in cases with suspected driving impairment and autopsy cases by analysis of human whole blood samples. The following compounds were included: alprazolam, bromazepam, clonazepam, diazepam, flunitrazepam, lorazepam, midazolam, nitrazepam, nordiazepam (metabolite of diazepam), oxazepam and phenazepam. Aliquots of 500μL whole blood were added 500μL of borate buffer pH 11 and extracted by solid-supported liquid–liquid extraction on ChemElut® columns using three times 2.5mL of methyl tert-butyl ether. Deuterated analogues were used as internal standards (IS) for all analytes, except for midazolam, phenazepam and bromazepam which had no commercially available deuterated analogues at the time the method was developed, and therefore used diazepam-d5, flunitrazepam-d7 and nitrazepam-d5, respectively. The analytes were separated using UPLC with a 2.1×100mm BEH C18-column, 1.7μm particle size, and quantified by MS/MS using multiple reaction monitoring (MRM) in positive mode. Two transitions were used for the analytes and one transition for the IS. The run time of the method was 8min including equilibration time. The concentrations of the benzodiazepines in the method span a broad range varying from the lowest concentration of 0.005μM for flunitrazepam to the highest of 20μM for oxazepam. The calibration curves of extracted whole blood standards were fitted by second-order calibration curves weighted 1/x, with R2 values ranging from 0.9981 to 0.9998. The intermediate precision had a CV (%) ranging between 2 and 19%. Recoveries of the analytes were from 71 to 96%. The LLOQs for the analytes varied from 0.0006 to 0.075μM and the LODs from 0.005 to 3.0nM. Matrix effects were studied by post extraction addition and found to be between 95 and 104% when calculated against an internal standard. A comparison with two other LC–MS methods was performed during method validation. Good correlation was seen for all analytes. The method has been running on a routine basis for several years, and has proven to be very robust and reliable with good results for external quality samples. The method also meets the requirements of the legislative limits for driving under the influence of non-alcohol drugs to be introduced in the Norwegian legislative system from 2012.

Hypostasis in the heart and great vessels of non-traumatic in-hospital death cases on postmortem computed tomography: Relationship to antemortem blood tests

Objectives The purpose of this study was to investigate hypostasis in the heart and great vessels (HHGV) by postmortem computed tomography (PMCT) in cases of non-traumatic in-hospital deaths and to analyze relationships between clarity of HHGV and antemortem blood markers. Material and methods The study included 31 cadavers of patients who died while receiving treatment in our academic tertiary-care hospital between April and December 2009. Antemortem blood tests including both hematological and blood coagulation markers were performed within 48 h before death, and PMCT and conventional autopsies were performed. The findings of HHGV were assessed in the following six locations: the superior vena cava, right atrium, right ventricle, thoracic aorta, left atrium, and left ventricle. The clarity of HHGV on PMCT was divided into two groups using attenuation of the left ventricular wall as a reference: no/obscure HHGV group and clear HHGV group. The relationships between clarity of HHGV and six blood markers [red blood cell (RBC) count, hemoglobin (Hb), hematocrit (Ht), prothrombin percentage activity (PT%), activated partial thromboplastin time (aPTT), and fibrinogen] were evaluated statistically. Results We observed no/obscure HHGV in 14 patients (45%) and clear HHGV in 17 patients (55%). The level of fibrinogen in clear HHGV group was significantly higher than in no/obscure HHGV group ( P = 0.029). No relationships were found between clarity of HHGV and other five blood markers. Conclusion HHGV is a common PMCT finding. Clear HHGV on PMCT was related to higher level of fibrinogen in non-traumatic in-hospital death cases.

Current awareness of piperazines: pharmacology and toxicology

Although many piperazine derivatives exist, only a limited number have been studied, whereby they have been found to be generally stimulant in nature resulting from dopaminergic, noradrenergic, and predominantly serotoninergic effects in the brain. Reported toxic effects include agitation, anxiety, cardiac symptoms (e.g. tachycardia) and sometimes seizures. As for many drugs, they are primarily metabolized by cytochrome P450 with subsequent possible glucuronidation and/or sulfation. Their abuse has been relatively recently observed in the last decade with only a few identified in biological fluid (primarily 1-benzylpiperazine (BZP) and 1-(3-trifluoromethylphenyl)piperazine (3-TFMPP)) despite publications of a number of analytical methods. Even when detected, however, the toxicological significance of their presence is often difficult to ascertain as many cases involve other drugs as well as a wide and overlapping range of concentrations found in blood (both in life and after death). This paper reviews the current pharmacological and toxicological information for piperazine derivatives and also includes new ante-mortem and post-mortem blood data.

A study of blood alcohol stability in forensic antemortem blood samples

The effect of long-term storage on alcohol stability in preserved forensic antemortem blood samples was investigated. Thirty-two whole blood case samples (each with two tubes of blood) were used for this study. One tube from each case was analyzed for blood alcohol concentration (BAC) for court proceedings of driving under the influence (DUI), and all blood samples were then stored under refrigeration. After the storage time (ranging from 13 to 39 months) both tubes of blood for each case were reanalyzed for BAC and the results were compared to the original analysis. Seven samples originally negative for alcohol analysis remained negative. The comparative data for 25 samples demonstrated various losses in BAC in both tubes. A significant loss with a mean of 0.015g/dL, was observed in previously opened tubes compared to a mean loss of 0.010g/dL in unopened tubes. In order to determine the effect of other storage conditions, the same blood samples were then stored at room temperature for 6 months followed by 38°C for 7 and 28 days and analyzed for BAC at the end of each storage time period. The seven alcohol negative cases remained negative when stored at room temperature or at 38°C. Six months of storage at room temperature decreased BAC further for both tubes of the alcohol positive cases with a mean loss of 0.014g/dL. Further storage at 38°C for 7 days did not cause any significant change in BAC. Storage at 38°C for 28 days caused some loss in BAC which was determined to be significant by statistical analysis.

Fatality from Drinking Denatured Alcohol and Hypothermia

A 19-year-old non-diabetic female suffering from irritable bowel syndrome was found unconscious outdoors in the month of October. She was severely hypothermic and rushed to hospital for life-saving treatment. Evidence emerged that the victim had attempted suicide by drinking denatured alcohol (T-Red). According to the manufacturer of this product, it contains > 85% (v/v) ethanol, ~5% (v/v) acetone, 1-2% (v/v) ethyl acetate, and ∼3% (v/v) methyl ethyl ketone (MEK), but no isopropanol. A venous blood sample taken on admission to hospital contained ethanol (660 mg/100 mL), acetone (25 mg/100 mL), isopropanol (78 mg/100 mL), and MEK, although the latter was not quantified. Despite intensive care, the patient died 21 h after admission and postmortem femoral blood contained ethanol (390 mg/100 mL), acetone (14 mg/100 mL), isopropanol (53 mg/100 mL), and MEK. During oxidative metabolism of ethanol, there is a shift in the redox state of the liver to a more reduced potential as reflected in a raised NADH/NAD(+) ratio, which impacts on other NAD-dependent biochemical reactions, including reduction of acetone to isopropanol. The lower concentrations of ethanol, acetone, and isopropanol in postmortem blood compared with antemortem blood indicate the metabolism of these substances during the 21-h survival period when the patient received emergency hospital treatment.

Fatal septic shock caused by candida myocarditis associated with malignant lymphoma and Epstein-Barr virus mediated disorder

A 60-year-old man with known epilepsy was admitted to our hospital due to hypotension, fever and arrhythmia. He was treated medically and myocardial infarction was ruled out. Treatment of septic shock was established according to the guidelines of the early goal-guided therapy including vasopressors, inotropic agents, mechanical ventilation, and hemofiltration in combination with empirical treatment with antimicrobial drugs and hydrocortisone. The patient’s condition deteriorated rapidly into multiorgan failure resulting in a fatal outcome. Antemortem blood cultures were sterile. Autopsy findings were compatible with Epstein-Barr virus mediated lymphoproliferative disorder, malignant lymphoma, disseminated candidiasis and candida myocarditis. Post-mortem blood and tissue cultures were positive for growth of candida glabrata and candida albicans, respectively. The post-mortem examination documented Epstein-Barr virus mediated lymphoproliferative disorder and malignant small cell lymphoma associated with candida myocarditis. Prior to death, the patient did not receive antiviral or antifungal treatment. Fatal candida myocarditis associated with septic shock secondary to disseminated candidiasis in a 60-years old male patient with prior unknown immune compromised state caused by Epstein-Barr virus mediated lymphoproliferative disorder and malignant lymphoma is described.

Two Fatal Intoxications with the New Designer Drug Methedrone (4-Methoxymethcathinone)

We report two fatalities involving the new designer drug methedrone, 4-methoxymethylcathinone. Blood was extracted with ethyl acetate after the addition of sodium hydroxide followed by evaporation and derivatization with TFAA before gas chromatography-mass spectrometry analysis. Hair was decontaminated and cut into segments, and after overnight extraction with acetonitrile/methanol/20 mM ammonium formate buffer (pH 3) (10:10:80), samples were analyzed by liquid chromatography-tandem mass spectrometry. The first case was treated in hospital, and blood was collected for drug screening. The concentration of methedrone in antemortem blood was 13.2 μg/g and in postmortem femoral blood 8.4 μg/g. The second case presented with 9.6 μg methedrone/g femoral blood, and in a hair sample, methedrone was detected in five short segments suggesting exposure to the drug during the months prior to death. In living abusers, the blood concentration range was 0.2-4.8 μg/g (n = 11). We conclude that use of methedrone may result in accidental death owing to its toxic properties and that the blood concentrations found in the two cases are close to those seen in the living. This suggests a rather narrow "therapeutic" window and emphasizes the danger in taking this kind of drug for recreational purposes.

Fentanyl in postmortem forensic toxicology

Context. Fentanyl is a powerful opioid used for the induction of anesthesia as well as for the management of severe pain. In recent years, transdermal fentanyl “patches” have become popular for outpatient management of chronic pain. The high potency and outpatient availability of fentanyl has also made it a highly sought-after drug of abuse. Deaths in which fentanyl is detected challenge medical examiners who must decide whether a given case represents therapeutic administration or overdose. Objectives. The current review seeks to present data about fentanyl that are relevant to the interpretation of postmortem blood and tissue fentanyl concentrations as well as to highlight areas which can be helpful or misleading in the evaluation of deaths potentially related to fentanyl exposure. Methods. Standard searching of the PubMed database for studies, case series, and case reports involving fentanyl kinetics, chemistry, and postmortem behavior was performed. Search terms typically included “postmortem” and “fentanyl,” as well as “kinetics” and “analysis,” where appropriate. Additional references were located as citations from reviewed papers or as conference abstracts. Results and Conclusions. The postmortem behavior of fentanyl is influenced heavily by pH changes and the antemortem kinetic behavior of the drug, especially, by its distribution. Postmortem blood fentanyl concentrations do not correlate directly with antemortem blood concentrations. Without adequate evaluation of kinetic data, investigative information and consideration of postmortem changes, misinterpretation of postmortem fentanyl results is likely.

Fatal Dog Bite in the Absence of Significant Trauma

Sudden and unexpected nontraumatic death in individuals with asplenia or hyposplenia is usually due to fulminant bacterial sepsis, most often involving Streptococcus pneumoniae, Neisseria meningitidis, and Hemophilus influenzae. We report a case of a previously well 40-year-old man who died 5 hours after hospital admission. At autopsy Waterhouse-Friderichsen syndrome was identified and Capnocytophaga canimorsus was subsequently isolated on antemortem blood cultures. Infection of humans with this organism is most often due to dog bite or contact. Upon specific inquiry it was ascertained that 2 days before admission the deceased had suffered a superficial bite to his hand by his pet Staffordshire Bullterrier dog. His relevant history included a previous splenectomy following blunt abdominal trauma. Asplenia and hyposplenia at autopsy should prompt microbiological testing with consideration of unusual organisms such as C. canimorsus. Although histories of animal contact or injury are often not available at the time of autopsy, this should also be considered in cases of apparent fulminant sepsis. In individuals with asplenia or hyposplenia, dog bites do not have to involve excessive tissue trauma, vascular compromise, or blood loss to be lethal.

Unique histological characteristics of Scedosporium that could aid in its identification

Scedosporium prolificans has been increasingly recognized as an etiological agent of disseminated mycelial infections in profoundly immunocompromised patients. Reported herein is a case of disseminated S. prolificans infection in a patient undergoing anti-neoplastic chemotherapy for acute myeloid leukemia. Antemortem blood culture yielded S. prolificans, which was confirmed on conventional morphological examination and polymerase chain reaction-based DNA sequencing targeting internally transcribed spacer regions. Histopathology of autopsy specimens indicated fungal infection in the heart, lungs, liver, kidneys, spleen, pancreas and gastrointestinal tract, with the development of hemorrhagic and ischemic necrosis. The infecting fungus had developing septate hyphae and was identified as belonging to the genus Scedosporium, on in situ hybridization of tissue. The combination of haphazardly branching hyphae and lemon-shaped conidia appeared to be the most useful distinguishing features to allow differentiation of this fungus from other filamentous fungi in tissue. Three other unique histopathological characteristics of the fungus were noted: (i) parallel hyphae bridged at right angles to produce letter-H patterns; (ii) intravascular conidiation; and (iii) purple conidia in tissue, though these are usually described as brown in most text books. Precise histopathology, in addition to other techniques such as in situ hybridization, can aid in the identification of etiological fungi.

Postmortem Acidification of Blood/Organs Induces an Increase in Flecainide Concentration in Cardiac Blood and the Contribution of the Lungs to This Increase

Postmortem acidification of blood and the contribution of this phenomenon to increased flecainide concentrations in cardiac blood were evaluated in rabbits. Flecainide was administered intravenously, antemortem peripheral blood was collected 15 min after administration, then rabbits were sacrificed. Blood and organs were collected immediately or 24 h after death, or immediately or 24 h after performance of cardiac massage. Postmortem left/right cardiac blood and organs showed lower pH than antemortem blood, and flecainide concentrations in all postmortem blood samples were higher than those in antemortem blood. Increased flecainide concentrations in cardiac blood were enhanced by postmortem cardiac massage and postmortem interval. In perfusion experiments using rabbit lung and heart, even if the flecainide concentration in inflow was kept constant, outflow concentrations were 2- to 3-fold higher than in inflow when inflow pH changed from 7.4 to 5.5. In contrast, flecainide concentration in outflow decreased immediately and then remained low when pH of perfusate changed from 5.5 to 7.4. These results demonstrate that flecainide accumulates in the lungs before death, and this accumulated flecainide releases into blood following postmortem acidification of blood/organs.

Postmortem candidaemia: marker of disseminated disease

AimThe significance of finding Candida species in heart blood cultures obtained at postmortem examination has never been studied. This article describes the findings of autopsy patients with postmortem candidaemia and...

Postmortem identification of hyperglycemia

The detection of diabetic coma postmortem requires accurate biochemical analysis. Due to continuous consumption of glucose by surviving cells postmortem, blood glucose levels decrease rapidly. Therefore, vitreous fluid has been used as a substitute in forensic practice, since it has a very low cell count. It has been repeatedly reported that the sum value of vitreous glucose and lactate should be used to estimate the original antemortem blood glucose level, based on the assumption that pre-existing glucose is gradually converted to lactate under anaerobic conditions during agonal phase and the early postmortem period. In this study, we applied a strategy including consistent sampling of vitreous fluid from the centre of both eyes of deceased subjects as soon as possible after arrival at the morgue, and immediate bedside analysis using a blood gas instrument. In total, 3076 cases were included during 2004–2006. We found that, after an initial drop of vitreous glucose during the very early postmortem period, the levels stayed stable for appreciable time postmortem. Analysis of a second sample collected at autopsy 1–3 days later gave similar results ( R 2 = 0.90). In contrast, the vitreous lactate levels showed a steady increase. This implies that the sum value of glucose and lactate increases with postmortem time, as reflected by vitreous potassium level. In fact, a statistically significant difference in the sum value was seen between subjects with potassium below 10 mmol/L ( n = 1086) and above 20 mmol/L ( n = 531), p < .001. In addition, in this large material, we did not identify a single case with circumstantial indication of hyperglycemia that only showed high vitreous lactate. We therefore suggest that vitreous glucose alone should be used to diagnose hyperglycemia postmortem and that the limit of 10 mmol/L should have a good specificity for diabetic coma, which theoretically would equal an original blood glucose value of about 26 mmol/L. As to the methodology, we found that sonication, centrifugation and addition of fluoride to the samples are unnecessary procedures when using a blood gas instrument. The strategy resulted in a doubling of the number of diabetic coma identified at our department compared to preceding period when analysis only was performed on selected cases.

Comparison of extraction efficiencies and LC–MS–MS matrix effects using LLE and SPE methods for 19 antipsychotics in human blood

Antipsychotic drugs are frequently associated with sudden death investigations. Detection of these drugs is necessary to establish their use and possible contribution to the death. LC-MS(MS) methods are common; however accurate and precise quantification is assured by using validated methods. This study compared extraction efficiency and matrix effects using common liquid-liquid and solid-phase extraction procedures in both ante-mortem and post-mortem specimen using LC-MS-MS. Extraction efficiencies and matrix effects were determined in five different blank blood specimens of each blood type. The samples were extracted using a number of different liquid-liquid extraction methods and compared with a standard mixed-mode solid-phase extraction method. Matrix effects were determined using a post-extraction addition approach-the blank blood specimens were extracted as described above and the extracts were reconstituted in mobile phase containing a known amount of analytes. The extraction comparison of ante-mortem and post-mortem blood showed considerable differences, in particular the extraction efficiency was quite different between ante-mortem and post-mortem blood. Quantitative methods used for determination of antipsychotic drugs in post-mortem blood should establish that there are no differences in extraction efficiency and matrix effects, particularly if using ante-mortem blood as calibrator.