Determination of umbilical cord and maternal plasma concentrations of fentanyl by using novel spectrophotometric and chemiluminescence enzyme immunoassays

Two novel enzyme immunoassays of fentanyl have been developed using Horseradish Peroxidase (HRP) as an enzyme, 3,3′,5,5′tetramethylbenzidine (TMB) and luminol as its colorimetric and its chemiluminescence substrate, respectively. A fentanyl polyclonal antibody was used as a capture antibody for fentanyl and fentanyl-bovine serum albumin (BSA) conjugate. The latter was first biotinylated and then bound by streptavidin labeled with HRP, resulting in the development of two novel competitive immunoassays. The detection limits were 0.045 and 0.0048 ng ml⁻¹ for spectrophotometric and chemiluminescence detection, respectively, and were much lower than existing HRP-fentanyl based kits. Intra- and inter-assay CVs were 2.6-4.5 and 5.4-11.2%, respectively, at concentrations of 0.050-5.000 ng ml⁻¹ for the colorimetric assay, whilst for the chemiluminescent assay intra- and inter-assay CVs were 3.7-6.2 and 6.2-12.3%, respectively, for the linear range of the assay at concentrations of 0.008-0.800 ng ml⁻¹. The methods in this study were developed in order to measure maternal and neonatal fentanyl plasma samples during cesarean section, after the application of a novel subarachnoid analgesia technique. The 28 maternal and neonatal plasma samples were measured by both assays, providing data for subarachnoid administration of fentanyl that had never been presented before.     

Diameter-selective solubilization of single-walled carbon nanotubes by reversible cyclic peptides

We have utilized reversible cyclic peptides (RCPs)-peptides containing alternating l- and d-amino acids with N- and C-termini derivatized with thiol-containing groups allowing reversible peptide cyclization-to solubilize and noncovalently functionalize carbon single-walled nanotubes (SWNTs) in aqueous solution. Solubilization occurs through wrapping of RCPs around the circumference of a SWNT, followed by the formation of head-to-tail covalent bonds, yielding closed rings on the nanotubes. By controlling the length of the RCPs, we have demonstrated limited diameter-selective solubilization of the SWNTs as revealed by UV/vis/NIR and Raman spectroscopies, as well as atomic force microscopy.     

Determination of the pesticides considered as endocrine-disrupting compounds (EDCs) by solid-phase extraction followed by gas chromatography with electron capture and mass spectrometric detection

An SPE method followed by GC-electron capture detection (ECD) with confirmation by MS for the trace determination of four pesticides considered as endocrine-disrupting compounds (EDCs) in natural waters and sediments has been developed. Target analytes, fenarimol, fenvalerate, pendimethalin, and vinclozolin, belong to different chemical groups and are used mainly in agriculture. In the present study, analysis employs an offline SPE step for the extraction of the target analytes from natural waters. Sonication and subsequent SPE clean-up was used for extraction and purification of the sediment samples which were finally treated with activated copper powder. The type of SPE disk, eluents as well as solution parameters including pH value, and concentrations of salts and humic substances were examined for the efficiency of the method. The recoveries of all pesticides were in relatively high levels, ranging from 75 to 97% for waters and 71 to 84% for sediment samples. Both methods were applied to real water and sediment samples and the presence of the tested compounds was investigated.     

Heteroarm Star‐Like Micelles Formed from Polystyrene‐block‐poly(2‐vinyl pyridine)‐block‐poly(methyl methacrylate) ABC Triblock Copolymers in Toluene

Polystyrene‐block‐poly(2‐vinyl pyridine)‐block‐poly(methyl methacrylate) ABC triblock copolymers were synthesized by sequential living anionic polymerization. Their solution properties were investigated in toluene, which is a bad solvent for the middle block. Spherical micelles are formed, which consist of a poly(2‐vinyl pyridine) dense core bearing polystyrene and poly(methyl methacrylate) soluble chains at the corona. These micelles exhibit the architecture of heteroarm star copolymers obtained by “living” polymerization methods. The aggregation numbers strongly depend on the length of the insoluble P2VP middle block, thus remarkably affecting the size of the micelles.     

Experimental Design and Optimization of Recovering Bioactive Compounds from Chlorella vulgaris through Conventional Extraction

Microalgae contain an abundance of valuable bioactive compounds such as chlorophylls, carotenoids, and phenolics and, consequently, present great commercial interest. The aim of this work is the study and optimization of recovering the aforementioned components from the microalgae species Chlorella vulgaris through conventional extraction in a laboratory-scale apparatus using a “green” mixture of ethanol/water 90/10 v/v. The effect of three operational conditions—namely, temperature (30–60 °C), duration (6–24 h) and solvent-to-biomass ratio (20–90 mL_solv/g_biom), was examined regarding the extracts’ yield (gravimetrically), antioxidant activity, phenolic, chlorophyll, and carotenoid contents (spectrophotometric assays), as well as concentration in key carotenoids, i.e., astaxanthin, lutein, and β-carotene (reversed-phase–high-performance liquid chromatography (RP–HPLC)). For this purpose, a face-centered central composite design (FC-CCD) was employed. Data analysis resulted in the optimal extraction conditions of 30 °C, for 24 h with 37 mL_solv/g_biom and validation of the predicted models led to 15.39% w/w yield, 52.58 mg_extr/mg_DPPH (IC50) antioxidant activity, total phenolic, chlorophyll, and carotenoid content of 18.23, 53.47 and 9.92 mg/g_extr, respectively, and the total sum of key carotenoids equal to 4.12 mg/g_extr. The experimental data and predicted results were considered comparable, and consequently, the corresponding regression models were sufficiently reliable for prediction.     

Comparative Interaction Studies of Quercetin with 2-Hydroxyl-propyl-β-cyclodextrin and 2,6-Methylated-β-cyclodextrin

Quercetin (QUE) is a well-known natural product that can exert beneficial properties on human health. However, due to its low solubility its bioavailability is limited. In the present study, we examine whether its formulation with two cyclodextrins (CDs) may enhance its pharmacological profile. Comparative interaction studies of quercetin with 2-hydroxyl-propyl-β-cyclodextrin (2HP-β-CD) and 2,6-methylated cyclodextrin (2,6Me-β-CD) were performed using NMR spectroscopy, DFT calculations, and in silico molecular dynamics (MD) simulations. Using T1 relaxation experiments and 2D DOSY it was illustrated that both cyclodextrin vehicles can host quercetin. Quantum mechanical calculations showed the formation of hydrogen bonds between QUE with 2HP-β-CD and 2,6Μe-β-CD. Six hydrogen bonds are formed ranging between 2 to 2.8 Å with 2HP-β-CD and four hydrogen bonds within 2.8 Å with 2,6Μe-β-CD. Calculations of absolute binding free energies show that quercetin binds favorably to both 2,6Me-β-CD and 2HP-β-CD. MM/GBSA results show equally favorable binding of quercetin in the two CDs. Fluorescence spectroscopy shows moderate binding of quercetin in 2HP-β-CD (520 M⁻¹) and 2,6Me-β-CD (770 M⁻¹). Thus, we propose that both formulations (2HP-β-CD:quercetin, 2,6Me-β-CD:quercetin) could be further explored and exploited as small molecule carriers in biological studies.     

Micellar and surface properties of a poly(methyl methacrylate)-block-poly(N-isopropylacrylamide) copolymer in aqueous solution

Critical micelle concentrations (cmc) of aqueous solutions of poly(methyl methacrylate)-block-poly(N-isopropylacrylamide) were determined at several temperatures by surface tensiometry. Below the lower critical solution temperature (LCST), the low Delta mic H 0 determined can be assigned to the PMMA block being tightly coiled in the dispersed molecular state, so that the unfavorable interactions of hydrophobic entities with water are minimized. Above the LCST the cmc value was found to increase; an anomalous behavior that can be directly related to the micelle-globule transition of the hydrophilic block. Interestingly, above the LCST the surface tension of relatively concentrated solutions was found to depend weakly on temperature not following the usual strong decrease with temperature expected for aqueous solutions.     

Portable, low-cost NMR with laser-lathe lithography produced microcoils

Nuclear Magnetic Resonance (NMR) is unsurpassed in its ability to non-destructively probe chemical identity. Portable, low-cost NMR sensors would enable on-site identification of potentially hazardous substances, as well as the study of samples in a variety of industrial applications. Recent developments in RF microcoil construction (i.e. coils much smaller than the standard 5mm NMR RF coils), have dramatically increased NMR sensitivity and decreased the limits-of-detection (LOD). We are using advances in laser pantographic microfabrication techniques, unique to LLNL, to produce RF microcoils for field deployable, high sensitivity NMR-based detectors. This same fabrication technique can be used to produce imaging coils for MRI as well as for standard hardware shimming or "ex-situ" shimming of field inhomogeneities typically associated with inexpensive magnets. This paper describes a portable NMR system based on the use of a 2 kg hand-held permanent magnet, laser-fabricated microcoils, and a compact spectrometer. The main limitations for such a system are the low resolution and sensitivity associated with the low field values and quality of small permanent magnets, as well as the lack of large amounts of sample of interest in most cases. The focus of the paper is on the setting up of this system, initial results, sensitivity measurements, discussion of the limitations and future plans. The results, even though preliminary, are promising and provide the foundation for developing a portable, inexpensive NMR system for chemical analysis. Such a system will be ideal for chemical identification of trace substances on site.