Near-Ultraviolet Circular Dichroism and Two-Dimensional Spectroscopy of Polypeptides

A fully quantitative theory of the relationship between protein conformation and optical spectroscopy would facilitate deeper insights into biophysical and simulation studies of protein dynamics and folding. In contrast to intense bands in the far-ultraviolet, near-UV bands are much weaker and have been challenging to compute theoretically. We report some advances in the accuracy of calculations in the near-UV, which were realised through the consideration of the vibrational structure of the electronic transitions of aromatic side chains.     

Validated Stability-Indicating GC-MS Method for Characterization of Forced Degradation Products of Trans-Caffeic Acid and Trans-Ferulic Acid

When dealing with simple phenols such as caffeic acid (CA) and ferulic acid (FA), found in a variety of plants, it is very important to have control over the most important factors that accelerate their degradation reactions. This is the first report in which the stabilities of these two compounds have been systematically tested by exposure to various different factors. Forced degradation studies were performed on pure standards (trans-CA and trans-FA), dissolved in different solvents and exposed to different oxidative, photolytic and thermal stress conditions. Additionally, a rapid, sensitive, and selective stability-indicating gas chromatographic-mass spectrometric method was developed and validated for determination of trans-CA and trans-FA in the presence of their degradation products. Cis-CA and cis-FA were confirmed as the only degradation products in all the experiments performed. All the compounds were perfectly separated by gas chromatography (GC) and identified using mass spectrometry (MS), a method that additionally elucidated their structures. In general, more protic solvents, higher temperatures, UV radiation and longer storage times led to more significant degradation (isomerization) of both trans-isomers. The most progressive isomerization of both compounds (up to 43%) was observed when the polar solutions were exposed to daylight at room temperature for 1 month. The method was validated for linearity, precision as repeatability, limit of detection (LOD) and limit of quantitation (LOQ). The method was confirmed as linear over tested concentration ranges from 1−100 mg L⁻¹ (r²s were above 0.999). The LOD and LOQ for trans-FA were 0.15 mg L⁻¹ and 0.50 mg L⁻¹, respectively. The LOD and LOQ for trans-CA were 0.23 mg L⁻¹ and 0.77 mg L⁻¹, respectively.     

An improved LC–MS/MS method for determination of docetaxel and its application to population pharmacokinetic study in Chinese cancer patients

Because of its unpredictable side effects and efficacy, the anticancer drug docetaxel (DTX) requires improved characterisation of its pharmacokinetic profiles through population pharmacokinetic studies. A sensitive and rugged LC–MS/MS method for the detection of DTX in human plasma was developed and optimised using paclitaxel as an internal standard (IS). The plasma samples underwent rapid extraction using hybrid solid-phase extraction-protein precipitation. The analyte and IS were separated with an isocratic system on a Zorbax Eclipse Plus C₁₈ column using water containing 0.05% acetic acid along with 20 μM of sodium acetate and methanol (30/70, v/v) as the mobile phase. Quantification was performed using a triple quadrupole mass spectrometer through multiple reaction monitoring in positive mode, using the m/z 830.3 → 548.8 and m/z 876.3 → 307.7 transitions for DTX and paclitaxel, respectively. The range of the calibration curve was 1–500 ng/mL for DTX, and the linear correlation coefficient was >0.99. The accuracies ranged from −4.6 to 4.2%, and the precision was no higher than 7.0% for the analytes. No significant matrix effect was observed. Both DTX and the IS showed considerable recovery. This method was finally applied to the establishment of a population pharmacokinetic model to optimise the clinical use of DTX.     

Identification of metabolic markers in patients with type 2 Diabetes by Ultrafast gas chromatography coupled to electronic nose. A pilot study

Metabolomics is a potential tool for the discovery of new biomarkers in the early diagnosis of diseases. An ultra-fast gas chromatography system equipped to an electronic nose detector (FGC eNose) was used to identify the metabolomic profile of Volatile Organic Compounds (VOCs) in type 2 diabetes (T2D) urine from Mexican population. A cross-sectional, comparative, and clinical study with translational approach was performed. We recruited twenty T2D patients and twenty-one healthy subjects. Urine samples were taken and analyzed by FGC eNose. Eighty-eight compounds were identified through Kovats's indexes. A natural variation of 30% between the metabolites, expressed by study groups, was observed in Principal Component 1 and 2 with a significant difference (p < 0.001). The model, performed through a Canonical Analysis of Principal coordinated (CAP), allowed a correct classification of 84.6% between healthy and T2D patients, with a 15.4% error. The metabolites 2-propenal, 2-propanol, butane- 2,3-dione and 2-methylpropanal, were increased in patients with T2D, and they were strongly correlated with discrimination between clinically healthy people and T2D patients. This study identified metabolites in urine through FGC eNose that can be used as biomarkers in the identification of T2D patients. However, more studies are needed for its implementation in clinical practice.     

A DNA Switch for Detecting Single Nucleotide Polymorphism within a Long DNA Sequence Under Denaturing Conditions

DNA detection is usually conducted under nondenaturing conditions to favor the formation of Watson–Crick base-paring interactions. However, although such a setting is excellent for distinguishing a single-nucleotide polymorphism (SNP) within short DNA sequences (15–25 nucleotides), it does not offer a good solution to SNP detection within much longer sequences. Here we report on a new detection method capable of detecting SNP in a DNA sequence containing 35–90 nucleotides. This is achieved through incorporating into the recognition DNA sequence a previously discovered DNA molecule that forms a stable G-quadruplex in the presence of 7 molar urea, a known condition for denaturing DNA structures. The systems are configured to produce both colorimetric and fluorescent signals upon target binding.