Determination of sialic acids in the serum of cancer patients by capillary electrophoresis

A novel method for the determination of N-acetylneuraminic acid (NANA) and N-glycolylneuraminic acid (NGNA) was developed by using high-performance capillary electrophoresis (HPCE) with UV detection at 195 nm. NANA and NGNA were separated directly and analyzed without pre- or postcolumn derivation. The detection limit of NANA is 9.6 x 10(-6) mol L-1 and for mass 3.879 x 10(-14) mol (39 fmol). This method was applied for the determination of NANA in 30 normal human and 72 cancer patients. The results demonstrated that NANA in the sera of cancer patients increased significantly as compared with the normal human (P < 0.001). The new method is simple and sensitive, and is suitable for basic research and clinical application to malignant tumors.

Simple derivatization method for sensitive determination of fatty acids with fluorescence detection by high-performance liquid chromatography using 9-(2-hydroxyethyl)-carbazole as derivatization reagent

A simple and sensitive method for the determination of short and long-chain fatty acids using high-performance liquid chromatography with fluorimetric detection has been developed. The fatty acids were derivatized to their corresponding esters with 9-(2-hydroxyethyl)-carbazole (HEC) in acetonitrile at 60 degreesC with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride as a coupling agent in the presence of 4-dimethylaminopyridine (DMAP). A mixture of esters of C-1-C-20 fatty acids was completely separated within 38 min in conjunction with a gradient elution on a reversed-phase C-18 column. The maximum fluorescence emission for the derivatized fatty acids is at 365 nm (lambda (ex) 335 nm). Studies on derivatization conditions indicate that fatty acids react proceeded rapidly and smoothly with HEC in the presence of EDC and DMAP in acetonitrile to give the corresponding sensitively fluorescent derivatives. The application of this method to the analysis of long chain fatty acids in plasma is also investigated. The LC separation shows good selectivity and reproducibility for fatty acids derivatives. The R.S.D. (n = 6) for each fatty acid derivative are <4%. The detection limits are at 45-68 fmol levels for C-14-C-20 fatty acids and even lower levels for acids. (C) 2001 Elsevier Science B.V. All rights reserved.

Grignard Reaction - Synthesis of Substituted Benzoic Acids

A convenient preparation of substituted benzoic acids from Grignard additions to solid carbon dioxide. Students create a library of carboxylic acids by using differentially substituted, commercially available aryl bromides, which can be used as the starting materials for a multistep synthesis. This is a modification and improvement of a very popular undergraduate organic chemistry experiment.

Diet-induced obesity alters the differentiation potential of stem cells isolated from bone marrow, adipose tissue and infrapatellar fat pad: the effects of free fatty acids.

INTRODUCTION: Obesity is a major risk factor for several musculoskeletal conditions that are characterized by an imbalance of tissue remodeling. Adult stem cells are closely associated with the remodeling and potential repair of several mesodermally derived tissues such as fat, bone and cartilage. We hypothesized that obesity would alter the frequency, proliferation, multipotency and immunophenotype of adult stem cells from a variety of tissues. MATERIALS AND METHODS: Bone marrow-derived mesenchymal stem cells (MSCs), subcutaneous adipose-derived stem cells (sqASCs) and infrapatellar fat pad-derived stem cells (IFP cells) were isolated from lean and high-fat diet-induced obese mice, and their cellular properties were examined. To test the hypothesis that changes in stem cell properties were due to the increased systemic levels of free fatty acids (FFAs), we further investigated the effects of FFAs on lean stem cells in vitro. RESULTS: Obese mice showed a trend toward increased prevalence of MSCs and sqASCs in the stromal tissues. While no significant differences in cell proliferation were observed in vitro, the differentiation potential of all types of stem cells was altered by obesity. MSCs from obese mice demonstrated decreased adipogenic, osteogenic and chondrogenic potential. Obese sqASCs and IFP cells showed increased adipogenic and osteogenic differentiation, but decreased chondrogenic ability. Obese MSCs also showed decreased CD105 and increased platelet-derived growth factor receptor α expression, consistent with decreased chondrogenic potential. FFA treatment of lean stem cells significantly altered their multipotency but did not completely recapitulate the properties of obese stem cells. CONCLUSIONS: These findings support the hypothesis that obesity alters the properties of adult stem cells in a manner that depends on the cell source. These effects may be regulated in part by increased levels of FFAs, but may involve other obesity-associated cytokines. These findings contribute to our understanding of mesenchymal tissue remodeling with obesity, as well as the development of autologous stem cell therapies for obese patients.

Preexisting compensatory amino acids compromise fitness costs of a HIV-1 T cell escape mutation.

BACKGROUND: Fitness costs and slower disease progression are associated with a cytolytic T lymphocyte (CTL) escape mutation T242N in Gag in HIV-1-infected individuals carrying HLA-B*57/5801 alleles. However, the impact of different context in diverse HIV-1 strains on the fitness costs due to the T242N mutation has not been well characterized. To better understand the extent of fitness costs of the T242N mutation and the repair of fitness loss through compensatory amino acids, we investigated its fitness impact in different transmitted/founder (T/F) viruses. RESULTS: The T242N mutation resulted in various levels of fitness loss in four different T/F viruses. However, the fitness costs were significantly compromised by preexisting compensatory amino acids in (Isoleucine at position 247) or outside (glutamine at position 219) the CTL epitope. Moreover, the transmitted T242N escape mutant in subject CH131 was as fit as the revertant N242T mutant and the elimination of the compensatory amino acid I247 in the T/F viral genome resulted in significant fitness cost, suggesting the fitness loss caused by the T242N mutation had been fully repaired in the donor at transmission. Analysis of the global circulating HIV-1 sequences in the Los Alamos HIV Sequence Database showed a high prevalence of compensatory amino acids for the T242N mutation and other T cell escape mutations. CONCLUSIONS: Our results show that the preexisting compensatory amino acids in the majority of circulating HIV-1 strains could significantly compromise the fitness loss due to CTL escape mutations and thus increase challenges for T cell based vaccines.

CPAG: software for leveraging pleiotropy in GWAS to reveal similarity between human traits links plasma fatty acids and intestinal inflammation.

Meta-analyses of genome-wide association studies (GWAS) have demonstrated that the same genetic variants can be associated with multiple diseases and other complex traits. We present software called CPAG (Cross-Phenotype Analysis of GWAS) to look for similarities between 700 traits, build trees with informative clusters, and highlight underlying pathways. Clusters are consistent with pre-defined groups and literature-based validation but also reveal novel connections. We report similarity between plasma palmitoleic acid and Crohn's disease and find that specific fatty acids exacerbate enterocolitis in zebrafish. CPAG will become increasingly powerful as more genetic variants are uncovered, leading to a deeper understanding of complex traits. CPAG is freely available at www.sourceforge.net/projects/CPAG/.