摘要
目的
1、本研究选择狗建立利多卡因蛛网膜下腔麻醉致死的动物模型,观察狗蛛网膜下腔麻醉致死过程的生命体征变化和死后组织脏器的病理改变。
2、建立血、脑脊液和脏器组织中利多卡因的薄层色谱扫描检测方法。
3、研究利多卡因在蛛网膜下腔麻醉致死狗体内的分布情况,为麻醉意外致死案件法医学鉴定的检材采取、检测、结果分析和死因判定提供科学依据。
方法
1、动物模型:狗10只,实验组每只狗行腰椎(L_(2-3))穿刺,置管于蛛网膜下腔,5分钟内经管缓慢注入利多卡因12.67mg/kg致死。
2、生命体征记录方法:BL-生物机能实验系统全程记录从给药开始到动物死亡的心电、血压和呼吸等主要生命体征变化。
3、样品采集:当心电、血压和呼吸全部消失时,迅速解剖动物,采取心脏、肝脏、肾脏、脾脏、肺脏、大脑、注射部位肌肉、注射部位20cm以外肌肉、心血、尿液、胆汁、侧
山西医科大学硕士学位论文
脑室脑脊液、脊髓腔脑脊液和不同节段的脊髓(包括延髓、
颈髓、上胸部脊髓、胸部脊髓和腰部脊髓)等组织,冷冻保
存。
4、病理观察:采取心脏、肝脏、’肾脏、脾脏、肺脏、大
脑、脊髓等组织,4%甲醛固定,石蜡包埋,切片,HE染色,
光镜观察。
5、利多卡因检测方法:样品碱化处理后,乙醚提取,薄
层扫描检测,根据利多卡因的Rf值结合光谱扫描图定性,外
标两点法或标准曲线法定量。
结果
1、实验组狗心电消失的平均时间为23.8士16.4分钟
(7一42分钟),血压的平均时间为16.4士n.5分钟(7一35分
钟),呼吸的平均时间为18.6士16.0分钟(10一47分钟)。
2、病理改变:各脏器呈现明显的急死表现,明显淤血、
水肿,心肌有纤维断裂,肝细胞出现空泡变性,脑和脊髓及
小脑神经细胞和血管周隙增宽等改变。
3、利多卡因薄层色谱检测线性范围1一200 pg加,各组
织中利多卡因最低检测限0.1一0.5 pg加,绝对回收率为
79.7%一90.2%,标准曲线相关系数均大于0.996。
4、蛛网膜下腔麻醉致死狗体内利多卡因平均含量从高到
低依次为:脊髓腔中脑脊液(485.6士51.5雌/ml)、侧脑室中
脑脊液(464.4士51.6 pg/ml)、上胸段脊髓(353.8士44.0 pg
/g)、腰段脊髓(353.0士60.1 pg/g)、注射部位脊髓(339.8
山西医科大学硕士学位论文
士52.1 pg/g)、胸段脊髓(237.0士36.3 pg/g)、颈段脊髓
(235.1士35.2 pg/g)、延髓(226.8士35.2 pg/g)、注射部位
肌肉(107.6士11.6pg/g)、脑(44.9士11.spg/g)、血液(40.3
士6.5 pg/ml)、胆汁(38.0士9.9 ug/ml)、尿液(37.5士9.5
pg/ml)、肺(28.4士6.8 pg/g)、肝( 26.3士6.7 pg/g)、肾(24.6
士7 .3 pg/g)、心(23.1士5.5 pg/g)、脾(22.2士4.6 pg/g)、
非注射部位肌肉(13.5士13.7 pg/g)。脑脊液与血液、脑组
织中利多卡因含量之比为12.4士2.7和10.8士4.5,各节段脊
髓与血液、脑组织中利多卡因含量比为5.7士0.9一9.0士2.6
和5 .1士3.1一7.9士5.2。
结论
1、本研究所建立的利多卡因蛛网膜下腔麻醉致死狗动物
模型的主要生命体征变化和死后病理变化与临床上利多卡因
麻醉意外死亡时的情况相似。此模型可应用于麻醉意外的法
医学研究。
2、实验狗麻醉致死过程中的主要生命体征变化和死后病
理变化以及利多卡因在其体内的分布数据可应用于利多卡因
麻醉意外的法医学鉴定。
3、本研究建立的各组织利多卡因检测方法具有简便易
行、快速有效、经济可靠的特点,可应用于麻醉意外致死案
件各组织中利多卡因的检测,尤其适合在基层公、检、法、
司系统法医检测中推广。
AIM: 1. To establish a lidocaine subarachnoid anesthesia death model with dogs, observe the vital sign and pathology of dogs after a lethal dose of lidocaine being injected into subarachnoid space. 2. To set up a thin layer chromatography (TLC) scanning analysis of lidocaine in tissue samples. 3. To study the distribution of lidocaine in dogs died of subarachnoid anesthesia, provide the scientific evidences for forensic medicine identification of the anesthesia accident death case.
METHOD: 1. Model: After having been placed a tube in the subarachnoid space, ten dogs were given a lethal dose of lidocaine ( 12.67 mg/kg ) by a slow injection in 5 minutes. 2. Record of vital signs: By an experimental system of biological function, the vital signs alteration of the experiment dogs, such as electro cardiac, respiration and blood pressure, had been recorded. 3. Collection and preservation of samples: As soon as the three vital signs disappeared, the dogs were anatomized, and the heart, liver, kidney, spleen, lung, brain, muscle in the injection location and no injection location, the heart blood, urine, bile, cerebrospinal fluid (CSF) in the lateral
ventricle and spinal subarachnoid space, spinal cord (medulla oblongata, cervical cord, the upper beast spinal cord, breast spinal cord and waist spinal cord) were taken out, some of which were preserved at -20 for qualitative and quantitative analysis, and the others were fixed with 4% formaldehyde for the pathology observation. 4. Phathology: The fixed tissues were cut into sections and stained with a HE staining for a light microscopic examination. 5. Detection: The samples were extracted with ethyl ether. The lidocaine in the sample was qualitated by Rf and a TLC scanning spectrometry, quantitated by a TLC scanning chromatography and a two points external standard method or a standard curve method.
RESULTS: 1. The electro cardiac, blood pressure and respiration of the dogs disappeared within 23.8 16.4 (7~42)min, 16.4 11.5 (7-35) min and 18.6 16.0 (10-47) minafterthe lidocaine was given. The dogs showed the pathology of rapid death cases. The vital sign and pathology of dogs were mostly similar to the signs of an anesthesia accident. 2. The concentrations detected in spinal subarachnoid space CSF, lateral ventricle CSF, the upper beast spinal cord, waist spinal cord, injection location spinal cord, breast spinal cord, cervical spinal cord, medulla oblongata, injection location muscle, brain, blood, bile, urine, lung, liver, kidney, hearts, spleen and no injection location muscle were 485.6 51.5 ug/ml ,464.4 51.6 ug/ml,
353.8 44.0 g/g, 353.0 60.1 ug/g, 339.8 52.1 ug/g, 237.0 36.3 ug/g, 235.1 35.2 ug/g, 226.8 35.2 ug/g, 107.6 11.6 ug/g, 44.9 11.5 ug/g, 40.3 6.5 ug/ml, 38.0 9.8 Ug/ml, 37.5 9.5 ug /ml, 28.4 6.8ug/g, 26.3 6.7ug/g, 24.6 7.3ug/g, 23.1 5.5 ug/g, 22.2 4.6ug/g, 13.5 13.7ug/g. Ratios of lidocaine in CSF to in blood and in brain were 12.4 2.7 and 10.8 4.5. Ratios of lidocaine in spinal cord to in blood and in brain were 5.7 0.9-9.0 2.6 5.1 3.1-7.9 5.2. 3. The lidocaine detection curve was linear over the range of 1-200 g/ml, the correlation coefficients were bigger than 0.996. The detection limit in tissues were 0.1-0.5 g /ml. The absolute extraction recoveries of lidocaine in tissues were 79.7%~90.2%.
CONCLUSION: The animal model established in this paper can be used for the forensic medicine study on lidocaine anesthesia accident death. The vital signs alteration data and distribution data of lidocaine in dogs died of subarachnoid anesthesia can be used for the forensic medicine identification of the anesthesia accident death case. The TLC scanning analysis can be used for identification and determination of lidocaine in tissue samples.
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