Development and Validation of a LC-MS/MS Technique for the Analysis of Short Chain Fatty Acids in Tissues and Biological Fluids without Derivatisation Using Isotope Labelled Internal Standards

The gut microbiota is critical to the maintenance of physiological homeostasis and as such is implicated in a range of diseases such as colon cancer, ulcerative colitis, diabetes, cardiovascular diseases, and neurodegenerative diseases. Short chain fatty acids (SCFAs) are key metabolites produced by the gut microbiota from the fermentation of dietary fibre. Here we present a novel, sensitive, and direct LC-MS/MS technique using isotopically labelled internal standards without derivatisation for the analysis of SCFAs in different biological matrices. The technique has significant advantages over the current widely used techniques based on sample derivatization and GC-MS analysis, including fast and simple sample preparation and short LC runtime (10 min). The technique is specific and sensitive for the quantification of acetate, butyrate, isobutyrate, isovalerate, lactate, propionate and valerate. The limits of detection were all 0.001 mM except for acetate which was 0.003 mM. The calibration curves for all the analytes were linear with correlation coefficients r² > 0.998. The intra- and inter-day precisions in three levels of known concentrations were <12% and <20%, respectively. The quantification accuracy ranged from 92% to 120%. The technique reported here offers a valuable analytical tool for use in studies of SCFA production in the gut and their distribution to host tissues.     

火焰原子吸收光谱法测定鸭肾中六种元素

选择采用ＨＮＯ３＋Ｈ２Ｏ２（２＋１消化法对鸭肾进行消化处理,用火焰原子吸收光谱法测定鸭肾中铜、锌、铅、铁、钙、镉等六种元素,实验表明该法有耗样少,精确度高,快速等优点,结果令人满意,该法具有一定的实用意义。     

B cell-associated immune profiles in patients with end-stage renal disease (ESRD)

Most of the previous studies on immune dysregulation in end-stage renal disease (ESRD) have focused on T cell immunity. We investigated B cell subpopulations in ESRD patients and the effect of hemodialysis (HD) on B cell-associated immune profiles in these patients. Forty-four ESRD [maintenance HD patients (n = 27) and pre-dialysis patients (n = 17)] and 27 healthy volunteers were included in this study. We determined the percentage of B cell subtypes, such as mature and immature B cells, memory B cells, and interleukin (IL)-10+ cells, as well as B cell-producing cytokines (IL-10, IL-4 and IL-21) by florescent activated cell sorting (FACS). B cell-associated gene expression was examined using real-time PCR and B cell producing cytokines (IL-10, IL-4 and IL-21) were determined using an enzyme- linked immunosorbent assay (ELISA). The percentage of total B cells and mature B cells did not differ significantly among the three groups. The percentages of memory B cells were significantly higher in the pre-dialysis group than in the HD group (P < 0.01), but the percentage of immature B cells was significantly lower in the pre-dialysis group than in the other groups. The percentages of IL-10-expressing cells that were CD19+ or immature B cells did not differ significantly (P > 0.05) between the two subgroups within the ESRD group, but the serum IL-10 concentration was significantly lower in the pre-dialysis group (P < 0.01). The results of this study demonstrate significantly altered B cell-associated immunity. Specifically, an imbalance of immature and memory B cells in ESRD patients was observed, with this finding predominating in pre-dialysis patients.     

A preliminary study of blood-oxygen-level-dependent MRI in patients with chronic kidney disease

Background

Blood-oxygen-level-dependent (BOLD) magnetic resonance imaging (MRI) can provide regional measurements of oxygen content using deoxyhemoglobin paramagnetic characteristics. The apparent relaxation rate or R2*(=1/T2*) can be determined from the slope of log (intensity) versus echo time and is directly proportional to the tissue content of deoxyhemoglobin. Thus, as the level of deoxyhemoglobin increases, T2* will decrease, leading to an increase in R2*. Chronic kidney disease (CKD) can affect oxygenation levels in renal parenchyma, which influences the clinical course of the disease. The goal of this study was to detect and assess renal oxygenation levels in CKD using BOLD MRI.

Methods

Fifteen healthy subjects and 11 patients with CKD underwent a renal scan using multigradient-recalled-echo sequence with eight echoes. R2* (1/s) of the renal cortex and medulla was measured on BOLD images. Of the 11 patients, nine had biopsy-proven chronic glomerulonephritis, and two had a similar diagnosis based on clinical symptoms and investigations.

Results

Mean medullary R2* (MR2*) and cortex R2* (CR2*) levels were significantly higher in patients (22 kidneys, MR2*=24.79±4.84 s⁻¹, CR2*=18.97±2.72 s⁻¹) than in controls (30 kidneys, MR2*=19.98±1.19 s⁻¹, CR2*=16.03±1.23 s⁻¹) (P<.01), and MR2* was increased more than CR2*. Medullary to cortical R2* ratios (MCR2*) of patients were significantly increased when compared with those of controls (P<.01). In the patient group, estimated glomerular filtration rate levels were greater than or equal to 60 ml/min/1.73 m² in six patients (12 kidneys), whose MR2* and CR2* were also significantly higher than those of controls (P<.01). Serum creatinine levels were normal in seven patients (14 kidneys), whose MR2*, CR2* and MCR2* were also higher than those of controls (P<.01).

Conclusions

BOLD MRI can be used to evaluate changes in renal oxygenation in CKD, suggesting that it has the potential to be an excellent noninvasive tool for the evaluation of renal function.     

液相色谱-串联质谱法检测动物肝肾组织中3种雌激素

建立了动物肝、肾组织中雌酮、17β-雌二醇、雌三醇药物残留检测的液相色谱-串联质谱(LC-MS/MS)分析方法。样品以叔丁基甲醚提取,于45 ℃下氮吹至干,用正己烷-二氯甲烷(6:4, v/v)重新溶解,过硅胶固相萃取小柱净化,氮吹至干,用1 mL乙腈-水(7:3, v/v)定容后测定,用Poroshell 120 EC-C18柱分离,以乙腈和水作为流动相进行梯度洗脱,电喷雾负离子模式电离,多反应监测模式检测,外标法定量。该方法对3种雌激素的线性范围均为1.0～20.0 μg/kg,相关系数大于0.99,定量限为1.0 μg/kg。在不同基质中,1.0、2.0、10.0 μg/kg 3个添加水平的平均回收率范围为70.2%～114%,相对标准偏差范围为2.01%～14.5%。该方法具有快速简便、净化效果好、灵敏度高等特点,适用于动物肝、肾组织中雌激素的检测。     

不同模拟体系中草酸钙结晶的比较研究

泌尿系结石是一种世界范围内的常见病及多发病 ,如深圳市的尿结石患病率高达 4.87%[1] ,其发病率呈上升趋势 .泌尿系结石在手术后复发率高 ,尤其是对结石的预防 ,目前尚无十分理想的方法 ,至今其形成机制未完全阐明 ,80 %以上的尿结石患者病因不清 [2 ] .草酸钙 (Ca C2 O4 )是泌尿系结石的主要成分 .体内 Ca C2 O4 结石的形成与热力学 (过饱和度 )和动力学 (成核、生长和聚集 )因素相关 .虽然在尿液中发现有二水草酸钙 (COD)晶体 ,但是热力学稳定的一水草酸钙 (COM)是尿石中最普遍的晶型[3 ] .在大多数的 Ca C2 O4 结石中 ,COM的发…     

Determination of bile acids in pig liver, pig kidney and bovine liver by gas chromatography-chemical ionization tandem mass spectrometry with total ion chromatograms and extraction ion chromatograms

An effective method has been developed for quantitative determination of six bile acids including lithocholic acid (LCA), deoxycholic acid (DCA), chenodeoxycholic acid (CDCA), hydodeoxycholic acid (HDCA), cholic acid (CA) and ursodeoxycholic acid (UDCA) in biological tissues including pig liver, pig kidney and bovine liver by gas chromatography-chemical ionization/tandem mass spectrometry (GC-CI/MS/MS). Camphor-10-sulphonic acid (CSA) was proposed as effective catalyst for bile acid derivatization. Reactions were accelerated ultrasonically. The effects of different catalysts and reaction times on derivatization efficiency were evaluated and optimized. Bile acids were determined as methyl ester-trimethylsilyl ether and methyl ester-acetate derivatives. The efficiency of trimethylsilylation and acetylation was evaluated. Trimethylsilylation was done with N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) as the trimethylsilyl donating reagent in a ultrasonic bath for 20 min. Acetylation was done in pyridine with acetic anhydride at 40-45°C for 4 h. The former reaction was faster than the latter. Thus, trimethylsilylation was employed for the quantitative analysis. Negligible interferences from sterols in biological matrices were observed when the biological samples were treated with solid phase extraction before GC-CI/MS/MS. The linearity, reproducibility, detection limit and recovery were evaluated under the optimized conditions. Satisfactory results were obtained when bile acid derivatives of LCA, CDCA, HDCA, and UDCA were determined with total ion chromatograms (TIC) while DCA and CA were determined with extracted ion chromatograms (EIC), respectively. The detection limits (S/N=3) for six bile acids in biological tissues were ranging from 0.40 to 1.6 ng/mL and the recoveries indicated that the proposed method was feasible for the determination of trace bile acids in the biological samples studied. The experimental results for the animal tissues purchased from five different markets were compared. Interestingly, all of the six bile acids were present in pig liver while only the dihydroxy bile acids, DCA, CDCA and HDCA were found in pig kidney. In addition to DCA and CDCA, trihydroxy bile acid, CA, are the major bile acids in bovine liver.     

Rosmarinic Acid Prevents Cisplatin-Induced Liver and Kidney Injury by Inhibiting Inflammatory Responses and Enhancing Total Antioxidant Capacity,Thereby Activating the Nrf2 Signaling Pathway

Drug-induced liver and kidney damage is an emergent clinical issue that should be addressed. Rosmarinic acid (RA) has obvious anti-inflammatory and antioxidant effects, so we evaluated the anti-inflammatory and antioxidant effects of RA pretreatment on serum and liver and kidney tissues of cisplatin (CP)-treated mice and explored the possible mechanisms. The results showed that RA pretreatment effectively downregulated the serum, liver, and kidney levels of ALT, AST, BUN, and CRE and the inflammatory factors IL-1β, IL-6, and TNF-α, and simultaneously enhanced the total antioxidant capacity of the liver and kidney. RA pretreatment significantly reduced the levels of MPO, MDA, and NO in liver and kidney tissue, inhibited the mRNA expression of IL-1β, IL-6, and TNF-α in liver and kidney tissue, activated the Nrf2 signaling pathway, and upregulated the mRNA expression of downstream target genes. Our findings show that RA could effectively prevent and alleviate acute liver and kidney injury caused by CP.     

Nutritional and Therapeutic Properties of Fermented Camel Milk Fortified with Red Chenopodium quinoa Flour on Hypercholesterolemia Rats

Quinoa is a nutrient-dense food that lowers chronic disease risk. This study evaluated the physicochemical and sensory qualities of fermented camel milk with 1, 2, 3, and 4% quinoa. The results showed that improvement in camel’s milk increased the total solids, protein, ash, fiber, phenolic content, and antioxidant activity more effectively. Fermented camel milk with 3% of quinoa flour exhibited the highest sensory characteristics compared to other treatments. Fermented camel milk enriched with 3% red quinoa flour was studied in obese rats. Forty male Wistar rats were separated into five groups: the first group served as a normal control, while groups 2–4 were fed a high-fat, high-cholesterol (HF)-diet and given 2 mL/day of fermented milk and quinoa aqueous extract. Blood glucose, malondialdehyde (MDA), low-density lipoprotein (LDL), cholesterol, triglyceride, aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP), creatinine, and urea levels decreased dramatically in comparison to the positive control group, while high-density lipoprotein (HDL), albumin, and total protein concentrations increased significantly. Fortified fermented camel milk decreased the number of giant adipocytes while increasing the number of tiny adipocytes in the body. The results showed that the liver and renal functions of hypercholesterolemic rats were enhanced by consuming fermented milk and quinoa. These results demonstrated the ability of quinoa and camel milk to protect rats from oxidative stress and hyperlipidemia. Further studies are needed to clarify the mechanisms behind the metabolic effects of fermented camel milk and quinoa.     

补肾调肝方对特发性少弱精症患者精液锌影响的临床观察

临床观察了60例特发性少弱精症患者使用补肾调肝方治疗后精液质量、精液微量元素锌的改变。60例患者精液密度、精液活力在治疗前后分别为(59.40±54.95)×10~6/mL、(85.48±75.58)×10~6/mL(P<0.05);(38.78±28.11)％、(47.10±28.74)％(P<0.05)。治疗前后精液锌含量分别为(1.79±1.20)mmol·L~(-1)、(1.96±1.07)mmol·L~(-1)。经计算,t值等于2.397,大于t_(0.05)(60),即P<0.05,治疗前后锌元素的差异有显著性意义。