Identification mark and identification method for accurately distinguishing drinking death and post-death wine filling

Abstract

The invention belongs to the technical field of judicial expertise, aims to solve the problem that false positive is easy to occur in ethanol source judgment in the current judicial expertise, and provides an identification mark and an identification method for accurately distinguishing drinking death and post-death wine filling. The identification marks are the concentration ratio of ethanol in heart blood to vitreous humor, ethanol in lower limb muscles and testis, and EtG and EtS in the lower limb muscles and testis. Experimental basis is provided for explaining an ethanol source problem in a forensic examination process through dispersion of ethanol injected into a dead rabbit in a dead body and generation conditions of EtG and EtS. And if the concentration ratio of the ethanol in the central blood to the vitreous humor is far greater than 1 in the actual detection process, the ethanol is possibly injected after death. Therefore, in the process of a forensic examination, the concentration ratio of ethanol in muscles and testis of lower limbs and in heart blood and vitreous humor should be detected in addition to detection of EtG and EtS in the heart blood by judging whether to drink or not.

Description

Technical Field

[0001] The present invention belongs to the technical field of judicial identification, and specifically relates to an identification mark and an identification method for accurately distinguishing death from drinking and posthumous ingestion of alcohol. Background Art

[0002] In the process of forensic identification, determining the source of ethanol has always been a key and difficult problem for forensic scientists. During the process of decomposition, nutrients in body fluids and tissues will ferment to produce ethanol under the action of microorganisms[1]. Or, in the case of alcohol being poured into the body, it is inaccurate to judge whether alcohol was consumed before death by simply detecting ethanol. Therefore, identifying biomarkers of ethanol entering the body before death, being produced after death, and being poured with alcohol after death is the key to explaining the source of ethanol.

[0003] Schmitt and Helander [2-3] et al. began to conduct preliminary research on ethyl glucuronide (EtG) and ethyl sulfate (EtS) as biomarkers of ethanol. About 2% of the ethanol that enters the body is metabolized through non-oxidative pathways [4], of which 0.5% of the ethanol uses UDP-glucuronic acid as a cofactor and is catalyzed by UDP-glucuronosyltransferase (UGT) superfamily enzymes to form EtG [5], and 0.1% of the ethanol reacts with 3'-phosphoadenosine-5'-phosphosulfate to form EtS under the catalysis of cytoplasmic sulfotransferases (SULTs) [6]. EtG and EtS have a long detection time (EtG in urine can reach 80 hours)[7-8]. Recently, there is also evidence that EtS in ex vivo blood has high stability at 37℃, 25℃, 4℃ and -20℃, and EtG has high stability under low temperature conditions (4℃ and -20℃). EtG is produced after adding ethanol to ex vivo blood and storing it at 37℃ for 7 days[9]. Most of the previous studies on the production of EtG and EtS used blood after drinking alcohol or blank blood with ethanol standard added. In actual cases, people may be fed alcohol after death to fake drinking before death. Summary of the invention

[0004] In order to solve the problem of false positives in the current judicial identification of the source of ethanol, the present invention provides an identification mark and an identification method for accurately distinguishing death from drinking and posthumous alcohol infusion.

[0005] The present invention is achieved by the following technical scheme: an identification mark for accurately distinguishing death from drinking and posthumous alcohol infusion, the identification mark being the ethanol concentration ratio in cardiac blood and vitreous humor, ethanol in lower limb muscles and testicles, and EtG and EtS in lower limb muscles and testicles.

[0006] The identification markers are the ethanol concentration ratio in cadaver heart blood and vitreous humor samples, ethanol in lower limb muscle and testis, and EtG and EtS in lower limb muscle and testis.

[0007] The identification method for accurately distinguishing death due to drinking and posthumous alcohol infusion using the identification mark, if the ratio of the ethanol concentration in the heart blood and vitreous humor of the corpse sample is less than 1, the sample was drunk before death; If the ratio of ethanol concentration in the heart blood and vitreous humor samples of the corpse sample is ≥ 1, and ethanol in the lower limb muscles and testicles is positive, the sample was drunk before death; If the ethanol in the lower limb muscles and testicles of the corpse sample is negative, and the EtG and EtS in the lower limb muscles and testicles are positive, the sample was consumed alcohol before death; If the ratio of ethanol concentration in the heart blood and vitreous humor sample of the corpse sample is ≧1, ethanol in the lower limb muscles and testicles is negative, EtG and EtS in the lower limb muscles and testicles are negative, and the sample was administered with alcohol after death.

[0008] The present invention provides an experimental basis for explaining the source of ethanol in the forensic examination process by analyzing the diffusion of ethanol in the body of a rabbit after death and the production of EtG and EtS. The results show that under high temperature conditions, the infusion of ethanol in a short period of time after the death of a rabbit will cause the production of EtG and EtS in body fluids and tissues, which brings new challenges to the determination of the source of ethanol. Therefore, in the forensic examination process, the ethanol concentration in the heart blood and vitreous humor samples, the ethanol in the lower limb muscles and testicles, and the EtG and EtS in the lower limb muscles and testicles should be detected to determine whether alcohol is consumed.

[0009] The experimental results of the present invention show that in the actual case investigation process, if the concentration ratio of ethanol in the heart blood and vitreous humor is much greater than 1, it indicates that ethanol may have been infused after death. In addition, in addition to detecting EtG and EtS in the heart blood, the lower limb muscles and testicles should also be tested to assist in determining whether alcohol was consumed. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] Figure 1 is the chromatogram of EtG and EtS in rabbit blood; Figure 2 is the chromatogram of EtG and EtS in rabbit liver; Figure 3 The concentrations of ethanol in body fluids and tissues at different gavage times after death; Figure 4 The concentrations of EtG in body fluids and tissues at different gavage times after death; Figure 5 The concentrations of EtS in body fluids and tissues at different gavage times after death. DETAILED DESCRIPTION

[0011] In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all the embodiments; based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0012] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs, and the disclosure and materials cited therein are hereby incorporated by reference.

[0013] Technical equivalents to the specific embodiments described that are apparent to those skilled in the art using no more than routine experimentation are intended to be encompassed by this application.

[0014] The experimental methods in the following examples are all conventional methods unless otherwise specified. The instruments and equipment used in the following examples are all conventional laboratory instruments and equipment unless otherwise specified; the experimental materials used in the following examples are all purchased from conventional biochemical reagent stores unless otherwise specified.

[0015] In order to clarify the source of ethanol in corpses, ethanol was injected into rabbit corpses and the concentrations of endosomes and metabolites EtG and EtS in body fluids and tissues were measured.

[0016] I. Animal experiment: 15 New Zealand white rabbits (Experimental Animal Center of Shanxi Medical University), male, weighing 2.0kg±0.2kg, fasted overnight before the experiment. The rabbits were fixed on the operating table, the trachea was clamped after the gastric tube was inserted, and after death (exhalation and heartbeat stopped), they were divided into 5 groups and placed at 37℃. Immediately after death, 1, 2, 4 and 8 hours after death, the dose of LD 50 Ethanol (LD 50 =6.3 g / kg) (n=3). 24 hours after intragastric administration, the heart blood, inferior vena cava blood, vitreous humor, liver, lung, kidney, brain, spleen, left lower limb muscle and testis were collected for detection of EtG and EtS.

[0017] 2. Main reagents and instruments: chromatographic grade methanol, chromatographic grade acetonitrile (Sigma-Aldrich, USA); chromatographic grade formic acid (Aladdin, Shanghai, China); tert-butyl alcohol, ethanol standards (Forensic Science Institute, Shanghai, China); 25% ammonia water (Damao, Tianjin, China); nitrogen (Anxu Hongyun Technology, Shanxi, China); Bond-Elut NH2 solid phase extraction cartridge (Agilent, USA); ethyl glucuronide (EtG) (Sigma-Aldrich, USA), deuterated ethyl glucuronide (D5-EtG, IS) (Sigma-Aldrich, USA), deuterated ethyl sulfate (D5-EtS, IS) (Sigma-Aldrich, USA); ethyl sulfate (EtS) (Sigma-Aldrich, USA). Ultrapure water used each time was freshly prepared by a water purifier.

[0018] Agilent 1260 liquid chromatograph-Agilent 6460 triple quadrupole tandem mass spectrometer (Agilent, USA); headspace sampler-gas chromatography system (Agilent, USA); Pressure+ 48 solid phase extractor (Biotage, Switzerland); Votex-genie2 vortex mixer (Scientific Industries, USA); Euro fully automatic nitrogen blowdown instrument (Eurotech Co., Ltd., Shanghai, China); Milli-Q water purifier (Millipore, USA); pipettes (1-10μL, 10-100μL, 100-1000μL,) (Eppendorf, Germany); hydrogen-air all-in-one machine HA-500 (China HP Analytical Technology Research Institute, Beijing, China); constant temperature climate chamber (Ningbo Yanghui Instrument Co., Ltd., Ningbo, China).

[0019] 3. Sample processing 1. Ethanol extraction: The method described in reference

[10] , for calibration samples, quality control samples and other samples used for methodological verification, take 100 μL of ethanol working solution of a certain concentration and 500 μL of tert-butyl alcohol internal standard working solution (40 μg / mL) and add them to the sample bottle, seal the aluminum cap with a silicone rubber pad with sealing forceps, mix well, and prepare for injection. For the body fluid sample to be tested, take 100 μL of body fluid into the injection bottle. For tissue samples, take 1 g of the homogenized tissue to be tested, add 2 mL of deionized water, vortex mix for 5 minutes, centrifuge at 3500 rpm for 5 minutes, take 100 μL of the supernatant into the injection bottle, and analyze the rest with the calibration sample using a headspace gas chromatography system (HS-GC).

[0020] 2. EtG and EtS extraction: The extraction method established in references [10,11]: Take 100 μL of whole blood in a 1.5 mL centrifuge tube, add 10 μL of mixed internal standard working solution (D5-EtG, D5-EtS, 2.5 μg / mL), and 400 μL of deionized water. Vortex for 30 seconds to mix evenly, and wait for sample loading.

[0021] 1.2 mL of methanol (0.3% formic acid) solution and 1.2 mL of deionized water were used to activate the solid phase extraction column in sequence. The nitrogen pressure was 2-3 Psi. After the sample was fully dispersed in the filler, 1.2 mL of methanol (0.3% formic acid) solution was added for elution. The above process was performed at a flow rate of 1 drop every 2 seconds. Dry for 10 minutes under a 5 Psi nitrogen flow, and add 1.2 mL of 5% NH4OH methanol-water solution for elution at the same flow rate. The eluate was dried in a 35°C water bath under nitrogen flow. Redissolved with 200 μL of 10% acetonitrile. After filtering through a filter membrane, it was analyzed using high-performance liquid chromatography-tandem mass spectrometry (LC-MS / MS).

[0022] Take 100 mg of the homogenized tissue to be tested, add 20 μL of mixed internal standard working solution (D5-EtG, D5-EtS, 25 μg / mL), 100 μL of deionized water and 800 μL of acetonitrile, shake and mix for 5 minutes, ultrasonicate in an ice bath for 30 minutes, centrifuge at 13000 rpm at 4°C for 10 minutes, take the supernatant and dry it in a 35°C water bath under nitrogen flow. Redissolve with 200 μL of 10% acetonitrile. After filtering through a filter membrane, analyze using high-performance liquid chromatography-tandem mass spectrometry (LC-MS / MS). 3. Instrument conditions Headspace gas chromatography (HS-GC): According to the national standard (GAT1073-2013), the headspace autosampler parameters were set as follows: heating box temperature 65°C; quantitative loop temperature 105°C; transfer line temperature 110°C; gas phase circulation time 3.5 min; sample bottle heating equilibrium time 10.0 min; sample bottle pressurization time 0.10 min; quantitative loop filling time 0.10 min; quantitative loop equilibrium time 0.05 min; injection time 1.0 min.

[0023] The gas chromatographic column was Agilent DB-ALC1 (30m*0.32mm*1.8μm) column, the column temperature was constant at 40°C, flow rate: 8mL / min. Inlet temperature: 150°C, split mode, split ratio 10:1. FID detector temperature: 250°C, H2 flow: 30mL / min, air flow: 360mL / min, tail blow flow N2: 25mL / min.

[0024] LC-MS / MS conditions: Chromatographic column: Agilent Zorbax SB-C18 (2.1*100mm; 3.5-Micron); mobile phase: water-acetonitrile (90:10), flow rate 0.4mL / min; injection volume: 5μL, column temperature 25℃; analysis time: 4.0min.

[0025] Ionization method: electrospray ionization (ESI); working mode: MRM (multiple reaction monitoring) mode. The mass spectrometry parameters of each target compound are shown in Table 1.

[0026] Table 1 MRM parameters of target compounds Note: 1) indicates quantitative ion.

[0027] 4. Methodological Validation: This methodological validation was completed according to industry guidelines

[12] and included specificity, limit of detection (LOD), lower limit of quantification (LLOQ), linearity, precision and accuracy, extraction recovery, matrix effect (ME), and stability.

[0028] 1. Specificity: Take 10 blank rabbit blood and rabbit livers from different sources, add 20% LLOQ dose of EtG and EtS, and analyze them by LC-MS / MS according to the above sample processing method to investigate specificity.

[0029] 2. LOD and LLOQ: The concentration of the analyte in the sample when the signal-to-noise ratio (SNR) is ≥3 is the limit of detection (LOD), and the lowest concentration of the analyte in the sample when SNR is ≥10 is the lower limit of quantitation (LLOQ).

[0030] 3. Linearity: Take 100 μL of blank rabbit blood and 1 g of blank rabbit liver and add ethanol standard to prepare working curve blood and tissue samples with a concentration range of 5 mg / 100 mL to 800 mg / 100 mL. After processing according to the above sample processing method, analyze by HS-GC.

[0031] EtG and EtS standard solutions were added to 100 μL blank rabbit blood to prepare working curve blood samples with concentration ranges of 25 ng / mL~2500 ng / mL and 40 ng / mL~2500 ng / mL, respectively. EtG and EtS standard solutions were added to 100 mg blank rabbit liver to prepare working curve tissue samples with concentration range of 0.05 ng / mg~20 ng / mg. The samples were processed according to the sample processing method in 1.3 and analyzed by LC-MS / MS.

[0032] 4. Precision and accuracy: Add ethanol standards to blank rabbit blood and rabbit liver to prepare four concentrations of LLOQ (5 mg / 100 mL), L (10 mg / 100 mL), M (200 mg / 100 mL), and H (800 mg / 100 mL), and analyze them after the above treatment. There are 6 parallel samples for each concentration, 24 samples in each batch, and a total of 5 batches of tests. Batches 1, 2, and 3 were tested in the morning, noon, and evening of the same day for intra-day accuracy and precision calculations, and batches 4 and 5 were tested at noon on the second and third days for inter-day accuracy and precision calculations. Accuracy and precision were calculated based on the working curve.

[0033] EtG and EtS standards were added to blank rabbit blood to prepare four concentrations of LLOQ (25 ng / mL, 40 ng / mL), L (100 ng / mL), M (1000 ng / mL), and H (2500 ng / mL). EtG and EtS standards were added to blank rabbit liver to prepare four concentrations of LLOQ (0.05 ng / mg), L (0.1 ng / mg), M (1 ng / mg), and H (20 ng / mg). The samples were processed according to 1.3 and analyzed. The precision and accuracy were calculated as above.

[0034] 5. Recovery and matrix effects: The recovery of EtG and EtS in blood and liver during the extraction process was determined by comparing the analyte concentrations of samples spiked before extraction and those of standards spiked at three concentrations (L, M, H) after extraction (n=6).

[0035] Blank blood and liver samples were extracted and spiked with EtG and EtS standards to evaluate matrix effects (n=6). The peak area ratios of EtG and EtS in matrix-spiked samples (A) at L, M, and H concentrations were compared to the peak area ratios of water-substituted samples (B) at the same concentrations. Matrix effects were defined as the percentage ratio of A to B. 100% indicates no matrix effect, >100% indicates ionization enhancement, and <100% indicates ionization suppression.

[0036] 6. Stability: Sample preparation and accuracy and precision are the same, with three batches of samples. The first batch was analyzed after being processed according to the above sample processing method. The second batch was analyzed after being processed according to the above sample processing method after three freeze-thaw cycles (-20℃~room temperature); the third batch was analyzed after being processed according to the above sample processing method in the injector for 24 hours. The repeated freeze-thaw stability and injection chamber stability are expressed as deviations. Deviation = (true concentration - measured concentration) / true concentration.

[0037] V. Experimental Results 1. Methodological validation results A. Specificity: The EtG and EtS detection methods in this experiment have strong specificity, good target separation effect, and no interference from the matrix in blood and tissues. Figure 1 , Figure 2 .

[0038] B. LOD, LLOQ and linearity: Ethanol has a good linear relationship in the range of 5 mg / 100 mL to 800 mg / 100 mL, and the correlation coefficient R 2 All are greater than 0.9999. For samples with too high ethanol concentration, dilute them first, and the final result is the measured concentration * dilution multiple. EtG and EtS in blood samples have good linear relationships in the range of 25ng / mL~2500ng / mL and 40ng / mL~2500ng / mL, respectively, and the correlation coefficient is R 2 The linear relationship between EtG and EtS in the liver was good in the range of 0.05g / mg~20ng / mg, and the correlation coefficient R 2 All were greater than 0.99. The LOD, LLOQ and linear equation are shown in Table 2.

[0039] Table 2: Detection limit, quantification limit and linearity of ethanol, EtG and EtS Note: 1) Ethanol unit is mg / 100mL; 2) Blood results; 3) Liver results.

[0040] C. Precision, accuracy, recovery, matrix effect and stability: The results of recovery, matrix effect and stability are shown in Table 3. The inter-day precision and intra-day precision were both within 1.50~6.37%, and the inter-day accuracy and intra-day accuracy were both between 86.72~104.33%, which met the industry requirements.

[0041] Table 3 Recovery, matrix effect and stability of EtG and EtS Note: 1) Blood results; 2) Liver results.

[0042] 2. Animal experiment results: LD of rabbits at 0, 1, 2, 4, and 8 hours after death 50After ethanol gavage at 37°C, the concentrations of ethanol, EtG, and EtS in the fluids and organs were detected after 24 hours. The results showed that ethanol diffused in the rabbit corpses. The degree of ethanol diffusion in body fluids (cardiac blood, inferior vena cava blood, and vitreous humor) decreased with the increase of postmortem gavage time, and the degree of diffusion in organs was consistent. Ethanol was not detected in the left lower limb muscles and testicles in the five postmortem gavage time groups, while ethanol was detected in the spleen only in the 0-hour postmortem gavage group and in the brain only in the 1-hour postmortem gavage group. Figure 3 When the postmortem interval was long (4h and 8h), EtG and EtS were not detected in all the body fluids and tissues tested, while when the postmortem interval was short (0h, 1h, 2h), EtG and EtS were detected in the heart blood, liver, spleen, and brain, only EtG was detected in the inferior vena cava blood, vitreous humor, and kidney, and only EtS was detected in the lung. The specific concentrations are shown in Tables 4 and 5. EtG and EtS were not detected in the left lower limb muscle and testis at the five postmortem gavage times. Figure 4 , Figure 5 In addition, in the blank group, ethanol, EtG, and EtS were not detected in all body fluids and tissues.

[0043] Table 4 Ethanol concentrations in body fluids and tissues of rabbits at different gavage times after death (mg / 100mL) Note: - indicates the concentration is below the detection limit.

[0044] Table 5 Concentrations of EtG and EtS in body fluids and tissues at different gavage times after death Note: - indicates the concentration is below the detection limit; 1) Concentration unit is ng / mL; 2) Concentration unit is ng / mg.

[0045] EtG and EtS are non-oxidative metabolites of ethanol and are widely used to distinguish between alcohol intake before death and ethanol production after death. However, some studies have shown that EtG will be produced when ethanol is added to blood at 37°C[9]. In actual criminal investigations, there are cases where alcohol is poured into corpses to falsify the scene. There are currently no clear identification indicators and methods for distinguishing between ethanol entering the body before death and its production after death, and whether EtG and EtS will be produced after alcohol is poured into the body after death.

[0046] The present invention simulated the phenomenon of alcohol infusion at different times of death under high temperature conditions using rabbits, and simultaneously detected ethanol, EtG and EtS in their body fluids and organs. The results showed that the ratio of ethanol concentration in heart blood and vitreous humor was 13.00±10.40, while the existing literature reported that in the study of ethanol redistribution in dogs after death, the ratio of ethanol concentration in heart blood and vitreous humor was 0.94±0.03

[13] (less than 1) 24 hours after death. This result suggests that the ratio of ethanol concentration in heart blood and vitreous humor in the two cases of postmortem ethanol infusion and premortem drinking is greater than the latter. This is because the heart is closer to the stomach than the vitreous body, and heart blood is more susceptible to postmortem diffusion than vitreous humor. Therefore, when judging premortem drinking and postmortem alcohol infusion, the ratio of ethanol concentration in heart blood and vitreous humor can be used as a reference. Ethanol, EtG and EtS were not detected in the blank group of this experiment, thus excluding the influence of postmortem ethanol on the research results.

[0047] From Table 5, we can see that when ethanol was infused within a short period of time (0-2 hours) after death, EtG or EtS was detected in all body fluids and organs except the left lower limb muscles and testicles. As a sample replacing blood, vitreous humor is less affected by microorganisms and postmortem redistribution [14-15] and is widely used in toxicological identification. In this experiment, ethanol was gavaged into the rabbits immediately after death, and ethanol, EtG and EtS were detected in the vitreous humor, with concentrations of 458.90 mg / 100 mL, 95.19 ng / mL and 232.50 ng / mL, respectively. Ethanol and EtG were detected in the vitreous humor 1 hour after death, with concentrations of 63.42 mg / 100 mL and 40.17 ng / mL, respectively. Therefore, there is a risk of false positives when simply relying on the detection of EtG and EtS in the vitreous humor to determine whether alcohol was consumed during life. A comprehensive judgment can be made based on the concentrations of EtG and EtS in tissues far from the heart and stomach, such as the lower limb muscles and testicles.

[0048] In the body, ethanol combines with glucuronides and sulfates to form EtG and EtS, respectively, through the catalysis of UGTs and SULTs. When the body dies, the enzymes in the body are not immediately inactivated, and some ethanol can still be metabolized under the catalytic action of the enzymes [1]. Since this experiment was conducted under high temperature conditions, the rabbits were immediately placed at 37°C after being killed, and some enzymes in the corpses were still active. Therefore, EtG and EtS were produced in body fluids and organs after ethanol gavage within a short period of time after death. In addition, there are literature reports that UGTs and SULTs also exist in Escherichia coli

[21] . Under high temperature conditions, corpses are accelerated to decay, and microorganisms may convert ethanol into EtG and EtS under the action of their own UGTs and SULTs. UGTs and SULTs are tissue-specific in the body [16-17]. The enzymes that produce EtG are mainly two subtypes, UGT1A1 and UGT2B7 [5]. UGTIA1 mRNA was observed in the liver and small intestine, but not in the colon, kidneys, and brain

[18] . The main enzyme that produces EtS is SULT1A1

[19] . Riley et al.'s research showed that SULT1A1 is mainly expressed in the liver, small intestine, and large intestine of rabbits

[20] . Figure 2 , 3 It is shown in the literature that EtS is mostly produced in organs, while EtG is mostly produced in body fluids, which also confirms the above research. Based on the above results, the reason why EtG and EtS are produced in body fluids and tissues after ethanol gavage in postmortem rabbits may be due to the combined action of enzymes that have not been inactivated and microorganisms produced by corruption.

[0049] Therefore, in the actual case investigation process, if the ratio of ethanol concentrations in cardiac blood and vitreous humor is much greater than 1, it indicates that ethanol may have been infused after death. In addition, in addition to detecting EtG and EtS in cardiac blood, lower limb muscles and testicles should also be tested to assist in determining whether alcohol was consumed. Since rabbits are small in size and are significantly affected by ethanol diffusion, large animals such as miniature pigs or monkeys are used in subsequent experiments to simulate experiments and observe the production of EtG and EtS in body fluids and tissues.

[0050] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

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Claims

1. An identification mark for accurately distinguishing death from drinking and posthumous drinking, characterized by: The identification markers are the ratio of ethanol concentration in cardiac blood to vitreous humor, ethanol in lower limb muscles and testicles, and EtG and EtS in lower limb muscles and testicles.

2. According to claim 1, an identification mark for accurately distinguishing death from drinking and posthumous drinking, characterized in that: The identification markers are the ethanol concentration ratio in cadaver heart blood and vitreous humor samples, ethanol in lower limb muscle and testis, and EtG and EtS in lower limb muscle and testis.

3. A method for accurately distinguishing death due to drinking and posthumous alcohol infusion using the identification mark according to claim 1 or 2, characterized in that: If the ratio of the ethanol concentration in the heart blood to the vitreous humor of the corpse sample is <1, the sample drank alcohol before death; If the ratio of ethanol concentration in the heart blood and vitreous humor samples of the corpse sample is ≥ 1, and ethanol in the lower limb muscles and testicles is positive, the sample was drunk before death; If the ethanol in the lower limb muscles and testicles of the corpse sample is negative, and the EtG and EtS in the lower limb muscles and testicles are positive, the sample was consumed alcohol before death; If the ratio of ethanol concentration in the heart blood and vitreous humor sample of the corpse sample is ≧1, ethanol in the lower limb muscles and testicles is negative, EtG and EtS in the lower limb muscles and testicles are negative, and the sample was administered with alcohol after death.

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