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Misleading results of ethanol analysis in urine specimens from rape victims suffering from diabetes
Journal of Clinical Forensic Medicine (2000) 7, 144 –146 © APS/Harcourt Publishers Ltd 2000
SHORT REPORT
Misleading results of ethanol analysis in urine specimens from rape victims suffering from diabetes A. W. Jones,1 A. Eklund,1 A. Helander2 1
Department of Forensic Chemistry, University Hospital, Linköping, Sweden Department of Clinical Neuroscience, Karolinska Institutet & Hospital, Stockholm, Sweden
2
SUMMARY. We report appreciably high concentrations of ethanol (82 and 102 mg/dL) in specimens of urine collected from two victims of date rape. Both girls (aged 15 and 18 years) suffered from diabetes mellitus, but adamantly denied drinking any alcohol before or after the incident. The presence of glycosuria and high risk of fungal infections in female diabetics suggests that ethanol was produced in vitro by fermentation after voiding. Making a routine test for sugar in the urine and ensuring that the sampling tubes contain sufficient sodium or potassium fluoride to inhibit glycolysis are recommended practices. A specific marker for post-sampling synthesis of ethanol might also be used such as the 5HTOL/5HIAA ratio. © APS/Harcourt Publishers Ltd 2000
Journal of Clinical Forensic Medicine (2000) 7, 144 –146
enzymes involved in glycolysis and can, therefore, prevent fermentation of glucose if the urine specimens are infected with microorganisms.8 When urine samples are taken in connection with clinical examination of rape victims or in other medical circumstances, fluoride preservatives are seemingly not always included in the sampling tubes. This creates a problem when toxicological results of ethanol analysis are interpreted. We report two instances of finding appreciably high concentrations of ethanol (82 and 102 mg/dL) in urine specimens from victims of date rape. Two girls, aged 15 and 18 years, denied drinking alcoholic beverages before the incidents and, interestingly, both suffered from type-1 diabetes mellitus. The forensic reports of high concentration of ethanol in urine led the police to suspect that the girls had been under the influence of alcohol at the time of the alleged rape and their credibility was, therefore, questioned.
BACKGROUND Glycosuria is one of the classic consequences of diabetes mellitus and the excess glucose excreted in urine can sometimes reach very high concentrations.1 Moreover, glucose is an excellent substrate for production of ethanol by fermentation and a glycosuria of 1% w/v (56 mmol/L) could theoretically produce a urine-ethanol concentration of approximately 500 mg/dL (109 mmol/L).2 Urine specimens are occasionally infected with bacteria or yeast such as Candida albicans which readily catalyses the fermentation of glucose.3 Yeast infections in the urinary tract and vagina are not uncommon,4 and those suffering from diabetes mellitus are especially prone to fungal infections.5,6 Diabetics, therefore, represent a high-risk group for post-sampling production of ethanol in urine.7 When specimens of urine are collected from drunk drivers or during medico-legal necropsy, the sampling tubes should always contain enough sodium or potassium fluoride to give a final concentration of at least 1% w/v.2 The fluoride ions inhibit several of the
CASE REPORTS During police investigations into the alleged rape of a girl aged 15 years, specimens of blood and urine were obtained from the victim approximately 15 h after the incident. The biological specimen was sent for toxicological analysis of ethanol as well as illicit and prescription drugs. The concentration of ethanol in blood was negative (<10 mg/dL), although traces of acetone was detected. The presence of illicit and prescription drugs were negative in the blood sample. The
A.W. Jones, A. Eklund, Department of Forensic Chemistry, University Hospital, SE-581 85 Linköping, Sweden. A. Helander, Department of Clinical Neuroscience, Karolinska Institute & Hospital, Stockholm, Sweden. Correspondence to: A.W. Jones 144
Ethanol in urine specimen from diabetic rape victims Table 1
145
Results of ethanol analysis in urine specimens from two victims of date rape, both of whom suffered from diabetes
Urine Specimen
Glucose1
First analysis of ethanol2
Second analysis of ethanol3
5HTOL/5HIAA ratio4
Girl aged 15 years Girl aged 18 years
++++ ++++
82 mg/dL5 102 mg/dL
550 mg/dL 800 mg/dL
14 nmol/µmol 6 nmol/µmol.
1 :Paper-strip test gave maximum response (~50 mmol/L);2:On arrival of specimens at the laboratrory; 3:After storage for approximately 3 months at ~4°C; 4:The reference value for 5HTOL/5HIAA in non-drinkers is <15 nmol/µmol; 5: Specimen also contained 11 mg/dL acetone
urine specimen contained 82 mg/dL ethanol and 11 mg/dL acetone, but illicit and prescription drugs were negative (Table 1). Because of the long interval between the rape and the time when biological fluids were obtained (~15 h), the police interpreted the ethanol concentration of 82 mg/dL in urine to mean that the girl was probably drunk at the time of the sexual assault. However, she adamantly denied drinking any alcoholic beverages before or after the offence and her credibility was, therefore, called into question in court. Because the girl suffered from type-1 diabetes and might have had glycosuria, expert testimony was sought to interpret the result of the ethanol analysis when the case was heard in the appeals court. Moreover, the lack of any fluoride preservative in the urine collection tubes made it highly likely that ethanol was produced after sampling. Finding a urine-alcohol concentration of 82 mg/dL and a zero blood-alcohol concentration in samples taken nearly simultaneously is hard to explain unless the person had held their urine for many hours after a period of heavy drinking. Blood-ethanol concentration steadily decreases to zero through metabolism as urine continues to be produced and is accumulated in the bladder. The abnormal urine/blood-alcohol relationship should have given a clue that something was suspect with the analytical results. A renewed analysis of the same urine specimen after it was stored for several months at 4°C showed the presence of a large amount of sugar and the concentration of ethanol had now increased dramatically (550 mg/dL), confirming an on-going fermentation process (Table 1). Evidence for post-sampling formation of ethanol was obtained by measuring the ratio of serotonin metabolites, 5-hydroxytryptophol (5HTOL) to 5hydroxyindoleacetic acid (5HIAA) in the urine sample.9 The urinary ratio of 5HTOL/5HIAA is normally very low (<15 nmol/µmol), but this increases appreciably after drinking alcohol owing to competitive inhibition of 5HIAA formation via the enzyme aldehyde dehydrogenase by acetaldehyde derived from oxidation of ethanol. Furthermore, the increased ratio of NADH/NAD+ during hepatic metabolism of
ethanol helps to promote formation of 5HTOL at the expense of 5HIAA.10,11 The 5HTOL/5HIAA ratio was 14 nmol/µmol (mean of two determinations), which is below the threshold value of 15 nmol/µmol used in clinical practice, and also considerably less than expected for a person with a urine-ethanol concentration of 82 mg/dL. At this alcohol concentration, the 5HTOL/5HIAA ratio often exceeds 200 nmol/µmol.11 In diabetics with ketoacidosis, β-hydroxybutyrate is converted into acetoacetate and the NADH/NAD+ ratio increases, although much less so than after drinking alcohol. The acetoacetate is further decarboxylated into acetone in a non-enzymatic reaction.1,8 It seems likely that the altered redox state (NADH/NAD+) in this subject, as indicated by the finding of acetone in blood and urine might have raised the 5HTOL/5HIAA ratio slightly to a value close to the threshold limit of 15 nmol/µmol.10 In a similar though unrelated case of an alleged rape of an 18-year-old girl, who also suffered from type-1 diabetes, specimens of blood and urine were taken ~19 h after the incident and sent for toxicological analysis. The blood sample was negative for ethanol as well as illicit and prescription drugs. However, the urine contained 102 mg/dL ethanol and was also positive for 7-amino-flunitrazepam and even contained 0.03 µg/mL carboxy-tetrahydrocannabinol (carboxy-THC). The 7-amino-flunitrazepam is a metabolite of flunitrazepam, which is the active substance in Rohypnol®, a potent benzodiazepine hypnotic and a drug which has been associated with so called ‘date rape’.12 Carboxy-THC is a metabolite of THC, the active ingredient in cannabis. Indeed, the girl admitted to the police that she had shared a handmade cigarette with her attacker and that she was also given an orange drink. She immediately felt drowsy and rested on the bed where the sexual assault took place. The girl vaguely remembers being aroused from her sleep and that sexual intercourse had definitely occurred. The results of toxicological analysis of urine indicates that the girl had taken the drug flunitrazepam some time earlier and that she had also smoked cannabis. The degree of impairment produced from
146
Journal of Clinical Forensic Medicine
these drugs cannot be deduced from the toxicological analysis of urine. However, the girl was adamant that she had not consumed any alcohol before or after the alleged rape so the police wanted an explanation for the negative blood alcohol concentration and the 102 mg/dL reported in urine. The police report also mentioned that the girl suffered from diabetes and that she had experienced poor glycaemic control at the time of the incident. The urine specimen was re-analyzed. It had been stored in a refrigerator at 4°C for several months, except for when the specimen was processed for analysis. The concentration of ethanol had now risen to 800 mg/dL confirming an on-going fermentation process (Table 1). A paper-strip test (Redia-Test, Boehringer Mannheim) for glucose gave the maximum response (~50 mmol/L). In this case, the urinary 5HTOL/5HIAA ratio was 6 nmol/µmol, being well within normal limits for those who have not been metabolizing alcohol, and thereby verifying the girl’s story that she had not consumed any alcohol.
CONCLUSION There are several lessons to be learnt from these two case reports. First, care is needed when urinary ethanol is measured in diabetic patients with genital candidiasis. Second, the tubes used to collect specimens of urine for alcohol analysis should always contain at least 1% sodium or potassium fluoride to prevent fermentation of any glucose that might be present. Third, a rapid dip-stick test for glycosuria should be applied routinely whenever urine specimens are found to contain ethanol. Fourth, if postsampling synthesis of ethanol is suspected, an aliquot of the urine specimen should be kept at room temperature in an airtight tube for a few days and then reanalyzed. An appreciable increase in the concentration of ethanol from the first analysis suggests ongoing fermentation of sugar in the urine specimen. Alternatively, a specific marker for post-sampling synthesis of ethanol could be used such as the 5HTOL/5HIAA ratio.9 Finding an abnormally high ratio speaks in favour of drinking ethanol some time earlier, whereas a low ratio leads to the conclusion that ethanol was synthesised in vitro after sampling. Also it might be possible to search for a specific urinary metabolite of ethanol such as ethyl glucuronide.
The presence of this metabolite would support recent consumption of ethanol because a very small amount (<1%) of the amount consumed is conjugated with glucuronic acid in the liver and excreted in urine.13 Further investigation showed that the tubes used for sampling blood and urine were included in a ‘rapekit’ purchased by the police from a small private company. The kit was prepared after consultation with various government forensic organizations.14 Although the Vacutainer tube used for sampling blood contained heparin and fluoride as preservatives, the need for including fluoride or some other enzyme inhibitor in the tubes used for collecting urine was not insisted upon leading to the consequences outlined in this article. REFERENCES 1. Sanson TH, Levine SN. Management of diabetic ketoacidosis. Drugs 1989; 38: 289–300. 2. Jones AW, Hylén L, Svensson E, Helander A. Storage of specimens at +4°C or addition of sodium fluoride (1%) prevents formation of ethanol in urine inoculated with Candida albicans. J Anal Toxicol 1999; 23: 333–336. 3. Saady JJ, Poklis A, Dalton HP. Production of urinary ethanol after sample collection. J Forensic Sci 1993; 38: 1467–1471. 4. Kremery S, Dubrava M, Kremery Jr V. Fungal urinary tract infections in patients at risk. Int J Antimicrob Agents 1999; 11: 289–291. 5. Gubbins PO, McConnell SA, Penzak SR. Current management of funguria. Am J Health Syst Pharm 1999; 56: 1929–1935. 6. Alexander WD, Wills PD, Eldred N. Urinary ethanol and diabetes mellitus. Diab Med 1988; 5: 463–464. 7. Alexander WD. Postmortem urinary alcohol is unreliable in diabetics. Brit Med J 1998; 317: 206. 8. Burtis CA, Ashwood ER (editors). Tietz Textbook of Clinical Chemistry, 2nd edn. London: WB Saunders Company, 1986, 66–67. 9. Helander A, Beck O, Jones AW. Distinguishing ingested ethanol from microbial formation by analysis of urinary 5hydroxytryptophol and 5-hydroxyindoleacetic acid. J Forensic Sci 1995; 40: 95–98. 10. Helander A, Beck O, Jones AW. Laboratory testing for recent alcohol consumption: Comparison of ethanol, methanol, and 5-hydroxytryptophol. Clin Chem 1996; 42: 618–624. 11. Jones, AW, Helander A. Disclosing recent drinking after alcohol has been cleared from the body. J Anal Toxicol 1996; 20: 141–142. 12. Simmons MM, Cupp MJ. Use and abuse of flunitrazepam. Ann Pharmacother 1998; 32: 117–119. 13. Schmitt G, Drönner P, Aderjan R, Skopp G. Blood sample containing 1.44 promille ethanol doesn’t contain ethylglucuronide – case report [in German]. Blutalkohol 1997; 34: 371–378. 14. Ansell R. Securing evidence after sexual offences is an important task for physicians [in Swedish]. Läkartidningen 1998; 95: 4626–4631.
SHORT REPORT
Misleading results of ethanol analysis in urine specimens from rape victims suffering from diabetes A. W. Jones,1 A. Eklund,1 A. Helander2 1
Department of Forensic Chemistry, University Hospital, Linköping, Sweden Department of Clinical Neuroscience, Karolinska Institutet & Hospital, Stockholm, Sweden
2
SUMMARY. We report appreciably high concentrations of ethanol (82 and 102 mg/dL) in specimens of urine collected from two victims of date rape. Both girls (aged 15 and 18 years) suffered from diabetes mellitus, but adamantly denied drinking any alcohol before or after the incident. The presence of glycosuria and high risk of fungal infections in female diabetics suggests that ethanol was produced in vitro by fermentation after voiding. Making a routine test for sugar in the urine and ensuring that the sampling tubes contain sufficient sodium or potassium fluoride to inhibit glycolysis are recommended practices. A specific marker for post-sampling synthesis of ethanol might also be used such as the 5HTOL/5HIAA ratio. © APS/Harcourt Publishers Ltd 2000
Journal of Clinical Forensic Medicine (2000) 7, 144 –146
enzymes involved in glycolysis and can, therefore, prevent fermentation of glucose if the urine specimens are infected with microorganisms.8 When urine samples are taken in connection with clinical examination of rape victims or in other medical circumstances, fluoride preservatives are seemingly not always included in the sampling tubes. This creates a problem when toxicological results of ethanol analysis are interpreted. We report two instances of finding appreciably high concentrations of ethanol (82 and 102 mg/dL) in urine specimens from victims of date rape. Two girls, aged 15 and 18 years, denied drinking alcoholic beverages before the incidents and, interestingly, both suffered from type-1 diabetes mellitus. The forensic reports of high concentration of ethanol in urine led the police to suspect that the girls had been under the influence of alcohol at the time of the alleged rape and their credibility was, therefore, questioned.
BACKGROUND Glycosuria is one of the classic consequences of diabetes mellitus and the excess glucose excreted in urine can sometimes reach very high concentrations.1 Moreover, glucose is an excellent substrate for production of ethanol by fermentation and a glycosuria of 1% w/v (56 mmol/L) could theoretically produce a urine-ethanol concentration of approximately 500 mg/dL (109 mmol/L).2 Urine specimens are occasionally infected with bacteria or yeast such as Candida albicans which readily catalyses the fermentation of glucose.3 Yeast infections in the urinary tract and vagina are not uncommon,4 and those suffering from diabetes mellitus are especially prone to fungal infections.5,6 Diabetics, therefore, represent a high-risk group for post-sampling production of ethanol in urine.7 When specimens of urine are collected from drunk drivers or during medico-legal necropsy, the sampling tubes should always contain enough sodium or potassium fluoride to give a final concentration of at least 1% w/v.2 The fluoride ions inhibit several of the
CASE REPORTS During police investigations into the alleged rape of a girl aged 15 years, specimens of blood and urine were obtained from the victim approximately 15 h after the incident. The biological specimen was sent for toxicological analysis of ethanol as well as illicit and prescription drugs. The concentration of ethanol in blood was negative (<10 mg/dL), although traces of acetone was detected. The presence of illicit and prescription drugs were negative in the blood sample. The
A.W. Jones, A. Eklund, Department of Forensic Chemistry, University Hospital, SE-581 85 Linköping, Sweden. A. Helander, Department of Clinical Neuroscience, Karolinska Institute & Hospital, Stockholm, Sweden. Correspondence to: A.W. Jones 144
Ethanol in urine specimen from diabetic rape victims Table 1
145
Results of ethanol analysis in urine specimens from two victims of date rape, both of whom suffered from diabetes
Urine Specimen
Glucose1
First analysis of ethanol2
Second analysis of ethanol3
5HTOL/5HIAA ratio4
Girl aged 15 years Girl aged 18 years
++++ ++++
82 mg/dL5 102 mg/dL
550 mg/dL 800 mg/dL
14 nmol/µmol 6 nmol/µmol.
1 :Paper-strip test gave maximum response (~50 mmol/L);2:On arrival of specimens at the laboratrory; 3:After storage for approximately 3 months at ~4°C; 4:The reference value for 5HTOL/5HIAA in non-drinkers is <15 nmol/µmol; 5: Specimen also contained 11 mg/dL acetone
urine specimen contained 82 mg/dL ethanol and 11 mg/dL acetone, but illicit and prescription drugs were negative (Table 1). Because of the long interval between the rape and the time when biological fluids were obtained (~15 h), the police interpreted the ethanol concentration of 82 mg/dL in urine to mean that the girl was probably drunk at the time of the sexual assault. However, she adamantly denied drinking any alcoholic beverages before or after the offence and her credibility was, therefore, called into question in court. Because the girl suffered from type-1 diabetes and might have had glycosuria, expert testimony was sought to interpret the result of the ethanol analysis when the case was heard in the appeals court. Moreover, the lack of any fluoride preservative in the urine collection tubes made it highly likely that ethanol was produced after sampling. Finding a urine-alcohol concentration of 82 mg/dL and a zero blood-alcohol concentration in samples taken nearly simultaneously is hard to explain unless the person had held their urine for many hours after a period of heavy drinking. Blood-ethanol concentration steadily decreases to zero through metabolism as urine continues to be produced and is accumulated in the bladder. The abnormal urine/blood-alcohol relationship should have given a clue that something was suspect with the analytical results. A renewed analysis of the same urine specimen after it was stored for several months at 4°C showed the presence of a large amount of sugar and the concentration of ethanol had now increased dramatically (550 mg/dL), confirming an on-going fermentation process (Table 1). Evidence for post-sampling formation of ethanol was obtained by measuring the ratio of serotonin metabolites, 5-hydroxytryptophol (5HTOL) to 5hydroxyindoleacetic acid (5HIAA) in the urine sample.9 The urinary ratio of 5HTOL/5HIAA is normally very low (<15 nmol/µmol), but this increases appreciably after drinking alcohol owing to competitive inhibition of 5HIAA formation via the enzyme aldehyde dehydrogenase by acetaldehyde derived from oxidation of ethanol. Furthermore, the increased ratio of NADH/NAD+ during hepatic metabolism of
ethanol helps to promote formation of 5HTOL at the expense of 5HIAA.10,11 The 5HTOL/5HIAA ratio was 14 nmol/µmol (mean of two determinations), which is below the threshold value of 15 nmol/µmol used in clinical practice, and also considerably less than expected for a person with a urine-ethanol concentration of 82 mg/dL. At this alcohol concentration, the 5HTOL/5HIAA ratio often exceeds 200 nmol/µmol.11 In diabetics with ketoacidosis, β-hydroxybutyrate is converted into acetoacetate and the NADH/NAD+ ratio increases, although much less so than after drinking alcohol. The acetoacetate is further decarboxylated into acetone in a non-enzymatic reaction.1,8 It seems likely that the altered redox state (NADH/NAD+) in this subject, as indicated by the finding of acetone in blood and urine might have raised the 5HTOL/5HIAA ratio slightly to a value close to the threshold limit of 15 nmol/µmol.10 In a similar though unrelated case of an alleged rape of an 18-year-old girl, who also suffered from type-1 diabetes, specimens of blood and urine were taken ~19 h after the incident and sent for toxicological analysis. The blood sample was negative for ethanol as well as illicit and prescription drugs. However, the urine contained 102 mg/dL ethanol and was also positive for 7-amino-flunitrazepam and even contained 0.03 µg/mL carboxy-tetrahydrocannabinol (carboxy-THC). The 7-amino-flunitrazepam is a metabolite of flunitrazepam, which is the active substance in Rohypnol®, a potent benzodiazepine hypnotic and a drug which has been associated with so called ‘date rape’.12 Carboxy-THC is a metabolite of THC, the active ingredient in cannabis. Indeed, the girl admitted to the police that she had shared a handmade cigarette with her attacker and that she was also given an orange drink. She immediately felt drowsy and rested on the bed where the sexual assault took place. The girl vaguely remembers being aroused from her sleep and that sexual intercourse had definitely occurred. The results of toxicological analysis of urine indicates that the girl had taken the drug flunitrazepam some time earlier and that she had also smoked cannabis. The degree of impairment produced from
146
Journal of Clinical Forensic Medicine
these drugs cannot be deduced from the toxicological analysis of urine. However, the girl was adamant that she had not consumed any alcohol before or after the alleged rape so the police wanted an explanation for the negative blood alcohol concentration and the 102 mg/dL reported in urine. The police report also mentioned that the girl suffered from diabetes and that she had experienced poor glycaemic control at the time of the incident. The urine specimen was re-analyzed. It had been stored in a refrigerator at 4°C for several months, except for when the specimen was processed for analysis. The concentration of ethanol had now risen to 800 mg/dL confirming an on-going fermentation process (Table 1). A paper-strip test (Redia-Test, Boehringer Mannheim) for glucose gave the maximum response (~50 mmol/L). In this case, the urinary 5HTOL/5HIAA ratio was 6 nmol/µmol, being well within normal limits for those who have not been metabolizing alcohol, and thereby verifying the girl’s story that she had not consumed any alcohol.
CONCLUSION There are several lessons to be learnt from these two case reports. First, care is needed when urinary ethanol is measured in diabetic patients with genital candidiasis. Second, the tubes used to collect specimens of urine for alcohol analysis should always contain at least 1% sodium or potassium fluoride to prevent fermentation of any glucose that might be present. Third, a rapid dip-stick test for glycosuria should be applied routinely whenever urine specimens are found to contain ethanol. Fourth, if postsampling synthesis of ethanol is suspected, an aliquot of the urine specimen should be kept at room temperature in an airtight tube for a few days and then reanalyzed. An appreciable increase in the concentration of ethanol from the first analysis suggests ongoing fermentation of sugar in the urine specimen. Alternatively, a specific marker for post-sampling synthesis of ethanol could be used such as the 5HTOL/5HIAA ratio.9 Finding an abnormally high ratio speaks in favour of drinking ethanol some time earlier, whereas a low ratio leads to the conclusion that ethanol was synthesised in vitro after sampling. Also it might be possible to search for a specific urinary metabolite of ethanol such as ethyl glucuronide.
The presence of this metabolite would support recent consumption of ethanol because a very small amount (<1%) of the amount consumed is conjugated with glucuronic acid in the liver and excreted in urine.13 Further investigation showed that the tubes used for sampling blood and urine were included in a ‘rapekit’ purchased by the police from a small private company. The kit was prepared after consultation with various government forensic organizations.14 Although the Vacutainer tube used for sampling blood contained heparin and fluoride as preservatives, the need for including fluoride or some other enzyme inhibitor in the tubes used for collecting urine was not insisted upon leading to the consequences outlined in this article. REFERENCES 1. Sanson TH, Levine SN. Management of diabetic ketoacidosis. Drugs 1989; 38: 289–300. 2. Jones AW, Hylén L, Svensson E, Helander A. Storage of specimens at +4°C or addition of sodium fluoride (1%) prevents formation of ethanol in urine inoculated with Candida albicans. J Anal Toxicol 1999; 23: 333–336. 3. Saady JJ, Poklis A, Dalton HP. Production of urinary ethanol after sample collection. J Forensic Sci 1993; 38: 1467–1471. 4. Kremery S, Dubrava M, Kremery Jr V. Fungal urinary tract infections in patients at risk. Int J Antimicrob Agents 1999; 11: 289–291. 5. Gubbins PO, McConnell SA, Penzak SR. Current management of funguria. Am J Health Syst Pharm 1999; 56: 1929–1935. 6. Alexander WD, Wills PD, Eldred N. Urinary ethanol and diabetes mellitus. Diab Med 1988; 5: 463–464. 7. Alexander WD. Postmortem urinary alcohol is unreliable in diabetics. Brit Med J 1998; 317: 206. 8. Burtis CA, Ashwood ER (editors). Tietz Textbook of Clinical Chemistry, 2nd edn. London: WB Saunders Company, 1986, 66–67. 9. Helander A, Beck O, Jones AW. Distinguishing ingested ethanol from microbial formation by analysis of urinary 5hydroxytryptophol and 5-hydroxyindoleacetic acid. J Forensic Sci 1995; 40: 95–98. 10. Helander A, Beck O, Jones AW. Laboratory testing for recent alcohol consumption: Comparison of ethanol, methanol, and 5-hydroxytryptophol. Clin Chem 1996; 42: 618–624. 11. Jones, AW, Helander A. Disclosing recent drinking after alcohol has been cleared from the body. J Anal Toxicol 1996; 20: 141–142. 12. Simmons MM, Cupp MJ. Use and abuse of flunitrazepam. Ann Pharmacother 1998; 32: 117–119. 13. Schmitt G, Drönner P, Aderjan R, Skopp G. Blood sample containing 1.44 promille ethanol doesn’t contain ethylglucuronide – case report [in German]. Blutalkohol 1997; 34: 371–378. 14. Ansell R. Securing evidence after sexual offences is an important task for physicians [in Swedish]. Läkartidningen 1998; 95: 4626–4631.