In vivo study on the protection of indole‐3‐carbinol (I3C) against the mouse acute alcoholic liver injury by micro‐Raman spectroscopy

Micro‐Raman spectroscopy (MRS) was utilized for the first time to evaluate the effect of indole‐3‐carbinol (I3C) on acute alcoholic liver injury in vivo. In situ Raman analysis of tissue sections provided distinct spectra that can be used to distinguish alcoholic liver injury as well as ethanol‐induced liver fibrosis from the normal state. Sixteen mice with liver diseases including acute liver injury and chronic liver fibrosis, and eight mice with normal liver tissues, and eight remedial mice were studied employing the Raman spectroscopic technique in conjunction with biomedical assays. The biochemical changes in mouse liver tissue when liver injury/fibrosis occurs such as the loss of reduced glutathione (GSH), and the increase of collagen (α‐helix protein) were observed by MRS. The intensity ratio of two Raman peaks (I₁₄₅₀/I₆₆₆) and in combination with statistical analysis of the entire Raman spectrum was found capable of classifying liver tissues with different pathological features. Raman spectroscopy therefore is an important candidate for a nondestructive in vivo screening of the effect of drug treatment on liver disease, which potentially decreases the time‐consuming clinical trials. Copyright © 2008 John Wiley & Sons, Ltd.     

Respiratory motion-corrected proton magnetic resonance spectroscopy of the liver

Purpose

To develop a post-processing, respiratory-motion correction algorithm for magnetic resonance spectroscopy (MRS) of the liver and to determine the incidence and impact of respiratory motion in liver MRS.

Materials and Methods

One hundred thirty-two subjects (27 healthy, 31 with nonalcoholic fatty liver disease and 74 HIV-infected with or without hepatitis C) were scanned with free breathing MRS at 1.5 T. Two spectral time series were acquired on an 8-ml single voxel using TR/TE=2500 ms/30 ms and (1) water suppression, 128 acquisitions, and (2) no water suppression, 8 acquisitions. Individual spectra were phased and frequency aligned to correct for intrahepatic motion. Next, water peaks more than 50% different from the median water peak area were identified and removed, and remaining spectra averaged to correct for presumed extrahepatic motion. Total CH₂+CH₃ lipids to unsuppressed water ratios were compared before and after corrections.

Results

Intrahepatic-motion correction increased the signal to noise ratio (S/N) in all cases (median=11-fold). Presumed extrahepatic motion was present in 41% (54/132) of the subjects. Its correction altered the lipids/water magnitude (magnitude change: median=2.6%, maximum=290%, and was >5% in 25% of these subjects). The incidence and effect of respiratory motion on lipids/water magnitude were similar among the three groups.

Conclusion

Respiratory-motion correction of free breathing liver MRS greatly increased the S/N and, in a significant number of subjects, changed the lipids/water ratios, relevant for monitoring subjects.     

Three-dimensional contrast-enhanced ultrasound of the liver: Experience of 92 cases

Three-dimensional contrast-enhanced ultrasound (3D-CEUS) is a combination of three-dimensional ultrasound (3DUS) and contrast-enhanced ultrasound (CEUS). To evaluate the feasibility of 3D-CEUS in liver imaging, investigate possible influencing factors to its image quality, and evaluate the influence of 3D-CEUS to clinical outcome, low acoustic power (mechanical index, 0.08-0.13) 3D-CEUS was carried out in 102 focal liver lesions in 92 patients by using the LOGIQ 9 ultrasound scanner and a volume transducer (frequency range, 2-5 MHz; focusing ability, 2-25 cm in depth; azimuth aperture 5.9 cm). The lesions were classified into two groups: group 1 (n = 51) for characterization and group 2 (n = 51) for local treatment response evaluation. The factors that influenced the image quality of 3D-CEUS were analyzed. The image quality and usefulness of 3D-CEUS between the two groups were compared by using the χ²-test. The results showed that the lesion diameter, location, and scanning route had no significant influence on the image quality in both groups, whereas interfering factors damaged the image quality in group 1. In group 1, during arterial phase, high image quality was more frequently found in hyperenhanced and hypo- or non-enhanced lesions compared with isoenhanced lesions. In group 2, interfering factor and local treatment response had no obvious influence on the image quality. The visualization rate of high image quality was 94.1% (48/51) in group 2 vs. 72.6% (37/51) in group 1 (P = 0.012). The investigators found that 3D-CEUS improved confidence but made no change in diagnosis in 19 (37.3%) of 51 lesions in group 1, whereas 41 (80.4%) of 51 lesions in group 2 (P = 0.000). 3D-CEUS tends to obtain better image quality and lead to higher diagnostic confidence in the lesions for local treatment response evaluation, and perhaps is more useful in this aspect in future clinical settings.     

猪肝脏和肌肉组织中β-兴奋剂克伦特罗的高效液相色谱电喷雾质谱分析研究

建立了猪肉和肝组织中克伦特罗的高效液相色谱电喷雾质谱(HPLC—ESI／MS)分析方法。在最佳色谱条件下克伦特罗的保留时间为16min,样品空白无干扰。定量分析的线性范围为2—100mg／L;最低检出限为0．75mg／L;方法回收率为95％-98％;RSD小于2．0％。利用ESI／MS／MS对β-兴奋剂克伦特罗进行了质谱解析,选择特征离子峰m／z277、259和203作为准确定性的依据。对实际生物样品猪瘦肉和肝进行检测结果表明,当样品中克伦特罗残留含量较高时,可以利用紫外检测数据对其进行准确定量。当残留含量低于最低检出限时,可以根据HPLC／ESI／MS结果中有无特征峰出现,给出较准确的定性结果。     

冷原子荧光法测定鲨鱼肝脏中的痕量汞

研究了冷原子荧光法测定鲨鱼肝脏中痕量汞的工作条件。样品风干后在2mol/L HNO3-4mol/L HCI体系中以V2O5为催化剂消解1h,上清液中的汞被SnCl2还原后于253.7nm用冷原子荧光法测定,在以10％SnCl2为还原剂时,线性范围0～0.2ng/mL,r=0.9997,检出限为0.05ng/mL,相对标准偏差2.5％,平均回收率94.0％～109.5％,该法简便、快速、准确。     

Covalent conjugation of tetrameric bovine liver catalase to liposome membranes for stabilization of the enzyme tertiary and quaternary structures

Tetrameric bovine liver catalase (BLC) is unstable because of its dissociation into subunits at low enzyme concentrations and the conformational change of the subunits at high temperatures. In this work, for stabilization of BLC, the enzyme was covalently conjugated with liposome membranes composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), cholesterol and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-glutaryl (NGPE). The NGPE, which was responsible for the BLC/membrane coupling, was altered from 0.05 to 0.2 in its liposomal mole fraction f_G. The catalase-conjugated liposome (CCL) with f_G of 0.15 showed the maximum number of the conjugated BLC molecules of 28 per liposome. The reactivity of CCLs to H₂O₂ was as high as that of free BLC at 25 °C in Tris–HCl buffer of pH 7.4. Among the CCLs, the catalyst with f_G of 0.15 was the most stable at 55 °C in its enzyme activity in the buffer because the appropriate number of BLC/liposome covalent bonding prevented the dissociation-induced enzyme deactivation. Furthermore, the CCL showed much higher stability at 55 °C than the free BLC/enzyme-free liposome mixture and free BLC at the low BLC concentration of 340 ng/mL. This was because BLC in the CCL was located in the vicinity of the host membrane regardless of the catalyst concentration, which could induce the effective stabilization effect of the membrane on the enzyme tertiary structure as indicated by the intrinsic tryptophan fluorescence analysis. The results obtained demonstrate the high structural stability of BLC in the CCL system, which was derived from the covalent bonding and interaction between BLC and liposomes.     

日本血吸虫病肉芽肿的电镜观察

应用长爪沙鼠建立日本血吸虫病动物模型, 观察实验性日本血吸虫病肝脏肉芽肿的超微结构改变. 采用腹部贴片法感染血吸虫尾蚴, 每只沙鼠感染(120±2)条尾蚴, 于5周后解剖, 取出肝脏做病理组织学和透射电镜观察. 结果发现, 肝脏切片H.E.(铁苏木素伊红)染色后可观察到虫卵肉芽肿产生, 透射电镜观察, 可见超微结构改变. 肝脏肉芽肿的超微结构的病变可以清晰地显示肉芽肿的发生机制, 为实验性研究血吸虫病的防治奠定了基础.     

两次液液萃取-气相色谱-质谱联用法测定动物肝脏中左旋咪唑残留

建立了液液萃取-气相色谱-质谱联用法测定动物组织中残留左旋咪唑的方法。在碱性溶液中将左旋咪唑盐酸盐转化为左旋咪唑,以乙酸乙酯进行提取;分别以HCl水溶液、氢氧化钾-二氯甲烷体系进行两次液液萃取净化,依次消除提取液中的脂溶性杂质和水溶性杂质,最后进入气相色谱-质谱系统,在选择离子监测模式下,以m/z 148、176、204为定性离子,m/z 204为定量离子进行结构确证和定量检测。结果表明: 左旋咪唑含量在0.25～3.0 mg/L范围内方法的线性关系良好(相关系数为0.999);定量限为5 μg/kg,低于当前国际最低限量标准;在鸡肝、鸭肝、兔肝和猪肝样品中的加标回收率在76%～106%范围内,相对标准偏差(RSD)小于9%。该法简便、稳定性好,无需对样品进行复杂的预处理即可实现对动物肝脏中左旋咪唑残留的快速准确测定。     

分子印迹固相萃取/高效液相色谱法测定猪肝中盐酸克伦特罗的方法研究

建立了分子印迹固相萃取/高效液相色谱法测定猪肝中盐酸克伦特罗残留量的方法。以ASB C18色谱柱(250 mm×4.60 mm,5μm)分离,流动相为甲醇-水(45∶55,p H 3.0),紫外检测波长243 nm,进样量10μL,流速0.6 m L/min,柱温30℃。分子印迹固相萃取条件为:上样溶剂为双蒸水,上样流速为0.6 m L/min;洗脱剂为1%三氟乙酸的甲醇溶液。上述优化条件下,该方法在0.432~4.32μg/m L范围内线性关系良好,相关系数(r)为0.999 0。加标回收率为84.7%~92.0%,相对标准偏差(RSD)为1.5%~1.8%。方法的准确度和精密度满足残留分析的要求。