High-performance Liquid Chromatography Measurements Research Articles

Concentrations of monosaccharides and their amino and alcohol derivatives in human preovulatory follicular fluid

The study purpose was to compare sugar and polyol concentrations in preovulatory ovarian follicular fluid (FF) with those in the circulation. Samples of FF and peripheral venous blood were obtained after an overnight fast from 14 women attending an IVF program. High performance liquid chromatography measurements of seven polyols, two aminohexoses and four hexoses were the main outcome measures. Glucose concentrations in FF and plasma were 2781.26 +/- 205.64 and 4431.25 +/- 65.17 microM, respectively (P < 0.001). Mannose concentration in FF was 38.99 +/- 3.33 microM, significantly lower than plasma concentration (55.38 +/- 2.29 microM; P < 0.001). A concentration gradient from plasma to FF was also significant for glycerol (99.41 +/- 8.47 versus 74.32 +/- 6.54 microM; P < 0.002), galactose (31.69 +/- 1.58 versus 26.73 +/- 1.93 microM; P < 0.01) and galactosamine (11.49 +/- 0.69 versus 6.38 +/- 0.59 microM; P < 0.001). The plasma-to-FF concentration difference was greatest for glucose (1649.99 +/- 204.09 microM). There was a significant correlation between plasma and FF concentrations for galactose and glycerol. This study supports a substantial utilization of glucose by the oocyte/granulosa cells complex, and documents a significant concentration gradient from plasma to FF for glycerol, mannose, galactose and galactosamine. These plasma-FF differences may reflect both utilization of these carbohydrates by the cells of the preovulatory ovarian follicle and/or transport characteristics of these cells.

Ethacrynic Acid Butyl-Ester Induces Apoptosis in Leukemia Cells through a Hydrogen Peroxide–Mediated Pathway Independent of GlutathioneS-Transferase P1-1 Inhibition

Ethacrynic acid (EA), a glutathione S-transferase inhibitor and diuretic agent, inhibits cell growth and induces apoptosis in cancer cells. To improve the activities, the structure of EA has been modified, and it has been shown that EA esters had an increased cell growth inhibitory ability compared with nonesterified analogue. EA butyl-ester (EABE) was synthesized, and its apoptosis induction ability was studied. The efficacy of EABE was compared with that of EA, and the mechanisms of action were studied in HL-60 leukemia cells. EABE exhibited greater cell growth inhibitory and apoptosis induction abilities than did EA. EABE-induced apoptosis in HL-60 cells correlated with increased levels of reactive oxygen species, the death receptor 5 (DR5), and caspase activation and decreased levels of the mitochondrial membrane potential. Pretreatment with antioxidants, either N-acetylcysteine or catalase, completely blocked EABE-induced apoptosis, H2O2 accumulation, and up-regulation of DR5 levels. RG19, a subclone of Raji cells stably transfected with a GSTpi expression vector, and K562 cells with high endogenous GSTP1-1 activity were less sensitive to EABE-induced apoptosis. EABE was more rapidly taken up than EA by HL-60 cells as determined by high-performance liquid chromatography (HPLC) measurements of intracellular concentrations. These results suggest that (a) H2O2 production is a mediator of EABE and EA-induced apoptosis; (b) GSTP1-1 plays a negative role in EABE and EA-induced apoptosis; and (c) the activity of EABE is greater than EA due to its more rapid entry into cells.

Chiral Discrimination at the Solid State of Methyl 2-(Diphenylmethylsulfinyl)acetate

The present paper reports a study on the chiral discrimination at the solid state of methyl 2-(diphenylmethylsulfinyl)acetate (hereafter, DMSAM) as a key intermediate in the synthesis of an acetamide derivative (modafinil). A partial solid solution (i.e., stable “racemic conglomerate” with partial miscibility at the solid state) between enantiomers was characterized by X-ray single crystal analyses and high performance liquid chromatography (HPLC). Binary and ternary phase diagrams of DMSAM were investigated by HPLC measurements, differential scanning calorimetry (DSC), discontinuous isoperibolic thermal analysis (DITA), and Raman spectroscopy. Finally, the distribution of the two enantiomers inside single crystals was studied by dissolution and Raman microspectroscopy.

Degradation processes in the cellulose/N-methylmorpholine-N-oxide system studied by HPLC and ESR. Radical formation/recombination kinetics under UV photolysis at 77 K

Degradation processes of N-methylmorpholine-N-oxide monohydrate (NMMO), cellulose and cellulose/NMMO solutions were studied by high performance liquid chromatography (HPLC) and electron spin resonance (ESR) spectroscopy. Kinetics of radical accumulation processes under UV (λ = 248 nm) excimer laser flash photolysis was investigated by ESR at 77 K. Beside radical products of cellulose generated and stabilized at low temperature, radicals in NMMO and cellulose/NMMO solutions were studied for the first time in those systems and attributed to nitroxide type radicals ∼CH2–NO•–CH2∼ and/or ∼CH2–NO•–CH3∼ at the first and methyl •CH3 and formyl •CHO radicals at the second step of the photo-induced reaction. Kinetic study of radicals revealed that formation and recombination rates of radical reaction depend on cellulose concentration in cellulose/NMMO solutions and additional ingredients, e.g., Fe(II) and propyl gallate. HPLC measurements showed that the concentrations of ring degradation products, e.g., aminoethanol and acetaldehyde, are determined by the composition of the cellulose/NMMO solution. Results based on HPLC are mainly maintained by ESR that supports the assumption concerning a radical initiated ring-opening of NMMO.

High performance liquid chromatographic measurement of iothalamate in human serum and urine for evaluation of glomerular filtration rate

A simple and sensitive HPLC-UV assay was developed for the measurement of iothalamate (IOT) in human serum and urine. Chromatographic separation was achieved using an embedded-carbamate-group bonded RP18 column and mobile phase consisting of 50 mM monobasic sodium phosphate and methanol (90:10, v/v) without the addition of ion-pair reagents. The assay demonstrated a high analytical reliability within the IOT concentration range of 1–150 μg/ml in serum and 25–1500 μg/ml in urine. The relative standard deviations (RSDs) for intra- and inter-day analysis were less than 5.1% in all cases. This method has been used for the evaluation of glomerular filtration rate (GFR) in subjects participating in a phase I clinical trial of a novel antimalarial medicine. The average baseline GFR was 100.41 ± 19.99 ml/min/1.73 m 2 in 119 healthy volunteers. The assay may also allow the simultaneous measurements of p-aminohippuric acid (PAH), N-acetyl PAH (aPAH), and IOT with some modification. PAH, IOT, aPAH, and β-hydroxyethyl-theophylline internal standard peaks appeared approximately at 2.5, 3.7, 5.9, and 11.8 min, respectively, in an isocratic run.

Synthesis of safflomide and its HPLC measurement in mouse plasma after oral administration

Safflomide ( N-caffeoyltryptamine) is a compound belonging to a group of phenylpropanoid amides found in plants. In this study, safflomide was chemically synthesized and confirmed by LC–MS, LC–MS/MS and NMR spectroscopic methods, and a high-performance liquid chromatography (HPLC) method was developed for quantifying safflomide in biological samples. The synthesis was simple, and the yield of safflomide was greater than 50%. Using the synthesized safflomide as a standard, HPLC separation was performed on a Nova-Pak C18 column using an isocratic buffer, and the separation was detected using a coulometric electrochemical detector. The detection of safflomide yielded an excellent peak resolution at the retention time of 21 min, and the lower limit of the detection was as little as 100 fmol. Using this HPLC method, the plasma concentrations of safflomide were determined in mouse blood, collected at 5, 10, 15, 20, 25, 30, and 35 min following its oral administrations (1 and 3 mg/30 g body weight). This HPLC method standardized with safflomide is the first reported method able to quantify the compound in standard and plasma samples with good detection limit and consistent reproducibility.

Insulin effects on myocardial function and bioenergetics in l-bupivacaine toxicity in the isolated rat heart

A positive effect of insulin-glucose-potassium infusion in severe bupivacaine-induced cardiovascular collapse has been described in vivo. It has been speculated that an antagonistic influence of insulin on sodium channel inhibition, transient outward potassium current, calcium-dependent adenosine triphosphatase or even improved myocardial energetics may be responsible for this effect. Using an isolated heart model, we therefore sought to further elucidate insulin effects in l-bupivacaine-induced myocardial depression. An isolated rat heart constant-pressure perfused, non-recirculating Langendorff preparation was used. Hearts were exposed to l-bupivacaine 5 microg mL(-1) and insulin 10 mIU mL(-1). Heart rate, systolic pressure, the first derivative of left ventricular pressure (+dP/dt), coronary flow, double product, PR and QRS intervals were recorded. Hearts were freeze-clamped and high-performance liquid chromatography measurement of the total adenine nucleotide pool was performed. l-Bupivacaine led to a significant decrease in heart rate, +dP/dt, systolic pressure, coronary flow and double product, and to an increase in PR and QRS. Insulin exerted a positive inotropic effect, significantly augmenting +dP/dt and systolic pressure in both l-bupivacaine-treated and control hearts. Heart rate, coronary flow, total adenine nucleotides, PR and QRS were not significantly changed by the insulin intervention. Insulin did not have a significant effect on total adenine nucleotides in controls and in l-bupivacaine-treated hearts. However, it does exert a positive inotropic action in bupivacaine-induced myocardial depression. We conclude that the positive effect of insulin application lies in positive inotropic action and not in changes in total adenine nucleotides.

Quantification of Liver Glucose Metabolism by Positron Emission Tomography: Validation Study in Pigs

The liver is inaccessible to organ balance measurements in humans. To validate [(18)F]fluorodeoxyglucose ([(18)F]FDG) positron emission tomography (PET) in the quantification of hepatic glucose uptake (HGU), we determined [(18)F]FDG modeling parameters, lumped constant (LC), and input functions (single arterial versus dual). Anesthetized pigs were studied during fasting (n = 6), physiologic (n = 4), and supraphysiologic (n = 4) hyperinsulinemia. PET was performed with C(15)O (blood pool) and [(18)F]FDG (glucose uptake). 6,6-Deuterated glucose ([(2)H]G) was coinjected with [(18)F]FDG and blood collected from the carotid artery and portal and hepatic veins to compute LC as ratio between tracers fractional extraction. HGU was estimated from PET images and ex vivo from high-performance liquid chromatography measurements of liver [(18)F]FDG versus [(18)F]FDG-6-phosphate and [(18)F]-glycogen. Endogenous glucose production was measured with [(2)H]G and hepatic blood flow by flowmeters. HGU was increased in hyperinsulinemia versus fasting (P < .05). Fractional extraction of [(18)F]FDG and [(2)H]G was similar (not significant), intercorrelated (r = 0.98, P < .0001), and equally higher during hyperinsulinemia than fasting (P <or= .05), with an LC of 0.98 +/- 0.10 and 1.18 +/- 0.26, respectively. [(18)F]FDG-PET modeling provided HGU values that did not differ from, and were correlated with, those from ex vivo measurements (r = 0.61, P <or= .02); proportional estimates of liver perfusion and endogenous glucose production were also obtained. Single and dual input functions produced strongly intercorrelated results (r > 0.95, P < .0001), with a modest underestimation of HGU by the former. [(18)F]FDG-PET-derived parameters provide accurate quantification of HGU and estimates of liver perfusion and glucose production. In the liver, LC of [(18)F]FDG is nearly unitary. Using a single arterial input introduces only a small error in estimation of HGU.

Guanine-specific DNA oxidation photosensitized by the tetraphenylporphyrinP(V) complex

Porphyrins have been studied as photosensitizers in photodynamic therapy. DNA is one of the most important targets of the sensitizer. In the present study, we have examined the photosensitized DNA damage caused by dihydroxo P ( V ) tetraphenylporphyrin ( P ( V ) TPP ), a cationic water-soluble porphyrin. P ( V ) TPP photosensitized guanine-specific damage to the DNA fragment. P ( V ) TPP induced severe photodamage to single-stranded rather than to double-stranded DNA. High performance liquid chromatography measurements confirmed the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-G), an oxidized product of 2'-deoxyguanosine, and showed that the content of 8-oxo-G in single-stranded DNA is larger than that in double-stranded DNA. The effects of reactive oxygen scavengers on DNA damage suggested the involvement of singlet oxygen (1O2). Photosensitized1O2formation was confirmed by near-infrared emission measurements. The results showed that1O2formation mainly contributes to the mechanism of DNA photodamage by P ( V ) TPP . Absorption spectrum measurements showed the interaction between P ( V ) TPP and DNA. This interaction is expected to enhance the1O2-mediated DNA damage since the lifetime of1O2in a cell is very short. On the other hand, P ( V ) TPP induced DNA damage at the consecutive guanines in double-stranded DNA. Because the consecutive guanines act as a hole trap, this DNA-damaging pattern suggests the partial involvement of photo-induced electron transfer. The fluorescence of P ( V ) TPP was quenched by DNA, supporting the electron transfer mechanism. However, DNA damage by electron transfer was not a main mechanism possibly due to reverse electron transfer. In conclusion, P ( V ) TPP binds to DNA and induces guanine-specific, photo-oxidation mainly via1O2generation.

Amino acid, ammonia and urea concentrations in human pre-ovulatory ovarian follicular fluid

This study aimed to determine amino acid (AA), ammonia and urea concentrations in human ovarian follicular fluid and to compare these concentrations with those in the circulation. Samples of pre-ovulatory follicular fluid and peripheral venous blood were obtained from 14 IVF patients. High-performance liquid chromatography (HPLC) measurements of 25 AAs were the main outcome measures. There was a significant gradient of most AAs from plasma to follicular fluid, with the exception of glutamate, which demonstrated a three-fold increase in follicular fluid concentration (70.0 +/- 3.80 microM) compared with plasma (23.18 +/- 2.20 microM; P < 0.001). The plasma-to-follicular fluid concentration difference for glutamine (81.83 +/- 9.2 microM) was greatest among all AAs. Among essential AAs, this difference was greatest for the branched-chain AAs, isoleucine, leucine and valine. Ammonia concentrations in follicular fluid and blood were 38.87 +/- 2.23 and 22.11 +/- 1.96 microM, respectively (P < 0.001). Urea concentration in follicular fluid was 3.37 +/- 0.18 mM, a value not significantly different from plasma concentration (3.36 +/- 0.22 mM; P = 0.911). These plasma-follicular fluid differences may reflect both the utilization of AAs and the transport characteristics of the follicular cells. There is accumulation of glutamate and ammonia in pre-ovulatory follicular fluid. The data for urea are consistent with transport by passive diffusion, with no evidence of an active urea cycle in the cells of the follicle.

Experimental calibration of lake-sediment spectral reflectance to chlorophyll a concentrations: methodology and paleolimnological validation

Chlorophyll a preserved in lake sediments reflects, in part, past primary production. This study assesses the spectral properties of sedimentary chlorophyll a using visible-near infrared reflectance (VNIR) spectroscopy, with the objective of establishing a new, non-destructive paleolimnological proxy. Reflectance spectra were determined from a dilution series (n = 10) involving incremental additions of pulverized modern algae to a lake sediment matrix of low organic content. This enabled an assessment of the development of sediment reflectance spectra in relation to different sediment chlorophyll a concentrations, and subsequent regression of spectral features against measured concentrations of chlorophyll a and derivatives obtained by high performance liquid chromatography (HPLC). The experiment demonstrates that ubiquitous troughs in sediment reflectance near 675 nm are attributable to chlorophyll a and derivative compounds. A significant correlation (r 2 = 0.98, P < 0.01) was obtained between the area of the reflectance trough in the 650–700 nm interval and summed concentrations of chlorophyll a, all derivative isomers, and degradational pheopigments. A simple linear inference model derived from this experiment was applied to a down-core sequence of VNIR spectra from a productive prairie lake (Alberta, Canada), where it produced inferred sediment chlorophyll a concentrations in concordance with HPLC measurements. Although a larger training set is desirable to further refine the inference model, the analyses reported here demonstrate that reflectance spectroscopy provides a rapid, semi-quantitative method for assessing the chlorophyll a content of lake sediments.

A new generation of anticancer, drug-loaded, colloidal vectors reverses multidrug resistance in glioma and reduces tumor progression in rats

By focusing on rat glioma, we elucidated whether new lipid nanocapsules (LNC) were able to improve anticancer hydrophobic drug bioavailability while also overcoming multidrug resistance. Blank LNCs and LNCs loaded with the antineoplastic agent paclitaxel were formulated by an emulsion inversion phase process. Expression of efflux pumps by rat glioma cells was assessed by reverse transcription-PCR, Western blot, and immunohistochemistry, and their activity was followed using the tracer (99)Tc(m)-methoxyisobutylisonitrile. Modalities of LNC action were addressed by using confocal microscopy detection of fluorescently labeled LNCs, fluorescence-activated cell sorting, high-performance liquid chromatography measurement of paclitaxel release, and analysis of tumor cell growth. This revealed an interaction between LNCs and efflux pumps that resulted in an inhibition of multidrug resistance in glioma cells, both in culture and in cell implants in animals. LNCs were able to target the intracellular compartment of glioma cells, a mechanism that was abrogated by using intracellular cholesterol inhibitors but not by clathrin-coated pit or caveolae uptake inhibitors. This result can be correlated to the LNC inhibitory effects on efflux pump activity that is itself known to be stimulated by intracellular cholesterol. In parallel, we showed that paclitaxel-loaded LNCs were active reservoirs from which paclitaxel could be released. Finally, we established that paclitaxel-loaded LNCs were more efficient than the commercially available paclitaxel formulation (Taxol) for clinical use, thus reducing tumor expansion in vitro and in vivo. Considering the physiologically compatible nature of LNC excipients, these data may represent an important step towards the development of new clinical therapeutic strategies against cancers.

The Ras/Raf-1/MEK1/ERK Signaling Pathway Coupled to Integrin Expression Mediates Cholinergic Regulation of Keratinocyte Directional Migration

The physiologic mechanisms that determine directionality of lateral migration are a subject of intense research. Galvanotropism in a direct current (DC) electric field represents a natural model of cell re-orientation toward the direction of future migration. Keratinocyte migration is regulated through both the nicotinic and muscarinic classes of acetylcholine (ACh) receptors. We sought to identify the signaling pathway mediating the cholinergic regulation of chemotaxis and galvanotropism. The pharmacologic and molecular modifiers of the Ras/Raf-1/MEK1/ERK signaling pathway altered both chemotaxis toward choline and galvanotropism toward the cathode in a similar way, indicating that the same signaling steps were involved. The galvanotropism was abrogated due to inhibition of ACh production by hemicholinium-3 and restored by exogenously added carbachol. The concentration gradients of ACh and choline toward the cathode in a DC field were established by high-performance liquid chromatographic measurements. This suggested that keratinocyte galvanotaxis is, in effect, chemotaxis toward the concentration gradient of ACh, which it creates in a DC field due to its highly positive charge. A time-course immunofluorescence study of the membrane redistribution of ACh receptors in keratinocytes exposed to a DC field revealed rapid relocation to and clustering at the leading edge of alpha7 nicotinic and M(1) muscarinic receptors. Their inactivation with selective antagonists or small interfering RNAs inhibited galvanotropism, which could be prevented by transfecting the cells with constitutively active MEK1. The end-point effect of the cooperative signaling downstream from alpha7 and M(1) through the MEK1/ERK was an up-regulated expression of alpha(2) and alpha(3) integrins, as judged from the results of real-time PCR and quantitative immunoblotting. Thus, alpha7 works together with M(1) to orient a keratinocyte toward direction of its future migration. Both alpha7 and M(1) apparently engage the Ras/Raf/MEK/ERK pathway to up-regulate expression of the "sedentary" integrins required for stabilization of the lamellipodium at the keratinocyte leading edge.

Simultaneous determination of DTPA, EDTA, and NTA by UV–visible spectrometry and HPLC

In this study, UV-visible spectrophotometry (UV-Vis) and high-performance liquid chromatography (HPLC) were used for simultaneous analysis of chelating agents diethylenetriamine pentaacetic acid (DTPA), ethylenediamine tetraacetic acid (EDTA), and nitrilotriacetic acid (NTA), as their metal chelates in dishwashing detergents, natural waters, and pulp mill water. The total amounts of the chelating agents in dishwashing detergents were verified by potentiometric titration with Fe(III) solution. Nickel(II) chelates were determined by UV-Vis and iron(III)chelates by HPLC and titration. Recoveries of DTPA, EDTA, and NTA from a standard mixture of analytes by UV-Vis were 107+/-7, 101+/-12 and 94+/-13%, respectively, and the recovery of the total amount of complexing agents was 99+/-4%. The limits of detection for DTPA, EDTA, and NTA were 667, 324, and 739 micromol L(-1), respectively. In HPLC measurements the optimized mobile phase contained 0.03 mol L(-1) sodium acetate, 0.002 mol L(-1) tetrabutylammonium bromide, and 5% methanol at pH 3.15 and the detection was by UV-Vis detection at 254 nm. All three complexing agents could be separated from each other in a simultaneous analysis in less than 5 min. The limits of detection were 0.34, 0.27, and 0.62 micromol L(-1) for DTPA, EDTA, and NTA, respectively. The total amounts of the analytes measured in the dishwashing detergents by the three techniques were found to be highly comparable (ANOVA: F=0.04, P=0.96). R(2) values were 0.99 for EDTA, 0.99 for NTA, and 0.99 for all the results when UV-Vis and HPLC determinations were compared using regression lines. The UV-Vis and HPLC methods were proved to be viable also for analyses of natural and pulp mill waters. The absence of matrix interferences was verified by the standard addition technique.

Comparison of Acetone and N,N-Dimethylformamide for Pigment Extraction in Turfgrass

The extraction and isolation of plant pigments are critical parts of the procedures used for pigment analyses by plant scientists. Various organic solvents have been used to liberate chlorophylls and carotenoids, including xanthophylls, from plant tissue. Acetone extraction is the most common extractant, but other solvents, such as N,N-dimethylformamide (DMF), have been used successfully. The objectives of this study were to compare the repeatability of acetone and DMF extraction for turfgrass pigment analysis and to investigate which extraction method is more appropriate for sample preparation before analysis using high-performance liquid chromatography (HPLC). Leaf tissue was collected from three plots of a mixed sward of creeping bentgrass (Agrostis stolonifera L.) and annual bluegrass (Poa annua L.) that were differentially treated in the field with magnesium (Mg), iron (Fe), and/or a plant growth regulator to encourage a range of leaf pigment concentrations. Variation among tissue samples, pigment extraction, and the HPLC process was used to compare the precision of acetone with DMF extraction. Results indicated that both acetone and DMF extraction procedures exhibited acceptable variation, but DMF extraction was more consistent at each procedural level. Procedural levels included variation among tissue samples (coefficient of variation = 11.2 < 12.1), variation among HPLC measurements (coefficient of variation = 2.1 < 2.9), and variation among extracted samples (coefficient of variation = 4.8 < 12.7). The DMF extraction also yielded 84% more neoxanthin, 53% more violaxanthin, 47% more lutein, 104% more chlorophyll b, 145% more chlorophyll a, and 14% more β-carotene than acetone. Overall, pigment extraction with DMF produced more quantitative and more reliable results than acetone extraction.

High-performance liquid chromatography measurement of hyperforin and its reduced derivatives in rodent plasma

A reverse-phase high-performance liquid chromatography method was developed for the determination of hyperforin and its reduced derivatives octahydrohyperforin and tetrahydrohyperforin in rodent plasma. The procedure includes solid-phase extraction from plasma using the Baker 3cc C8 cartridge, resolution on the Symmetry Shield RP8 column (150 mm × 4.6 mm, i.d. 3.5 μm) and UV absorbance detection at 300 nm. The assay was linear over a wide range, with an overall coefficient of variation less than 10% for all compounds. The precision and accuracy were within acceptable limits and the limit of quantitation was sufficient for studies preliminarily assessing the disposition of tetrahydrohyperforin and octahydrohyperforin in the mouse and rat.

Analysis of heat-processed corn foods for fumonisins and bound fumonisins

Thirty retail samples of heat-processed corn foods, i.e. corn flakes, corn-based breakfast cereals, tortilla chips and corn chips, were analysed for fumonisins — fumonisin B1 (FB1), fumonisin B2 (FB2) and hydrolysed FB1 (HFB1) — as well as for protein- and total-bound FB1. Bound (hidden) fumonisins cannot be detected by conventional analysis. Improved methods for the determination of bound FB1 were developed. The protein-bound FB1 was extracted with 1% sodium dodecylsulfate (SDS) solution. The SDS, which interfered with high-performance liquid chromatography (HPLC) analysis, was then separated from protein-bound FB1 by complexing with methylene blue followed by solvent extraction and hydrolysis with 2 N KOH. To measure total-bound FB1, the sample itself was hydrolysed with KOH. In both cases, clean-up was accomplished on an OASIS polymeric solid-phase extraction column and the bound fumonisins were determined by HPLC measurement of HFB1. Fourteen of 15 samples of corn flakes and other corn-based breakfast cereals analysed contained detectable levels of FB1 with a mean in positive samples of 67 ng g−1 (13–237 ng g−1). Two samples also had detectable levels of FB2 (21–23 ng g−1). Bound FB1 was found in all samples; the mean protein-bound FB1 measured was 58 ng g−1 (22–176 ng g−1) and the mean total-bound FB1 measured was 106 ng g−1 (28–418 ng g−1), reported as FB1 equivalents after correction for recoveries of HFB1. There was an average of about 1.3 times more FB1 in the bound form compared with extractable FB1, and this was about twice as much as protein-bound FB1. Seven of the 15 samples of alkali-processed corn-based foods, such as tortilla chips and corn chips, contained FB1 and three contained HFB1 with means in measurable positive samples of 78 (48–134) and 29 (13–47) ng g−1, respectively. Five of these alkali-processed corn foods contained bound FB1; the mean measurable protein-bound FB1 was 42 ng g−1 (39–46 ng g−1) and the mean measurable total-bound FB1 was 100 ng g−1(54–209 ng g−1). HFB1 derived from bound FB1 in selected samples was confirmed by HPLC with mass spectrometry (MS).

A Rapid and Accurate Method for Estimating Tomato Lycopene Content by Measuring Chromaticity Values of Fruit Purée

Lycopene is the red pigment and a major carotenoid in tomato (Lycopersicon esculentum Mill.) fruit. It is a potent natural antioxidant, and the focus of many tomato genetics and breeding programs. Crop improvement for increased fruit lycopene content requires a rapid and accurate method of lycopene quantification. Among the various available techniques, high-performance liquid chromatography (HPLC) can be accurate, however, it is laborious and requires skilled labor and the use of highly toxic solvents. Similarly, spectrophotometric methods, although easier than HPLC, also require time-consuming extractions and may not be as accurate as HPLC, as they often overestimate fruit lycopene content. Colorimetric estimation of fruit lycopene using chromaticity values has been proposed as an alternative rapid method. Previous studies that examined the utility of this technique, however, were confined to the evaluation of only one or few cultivars and, therefore, lacked broad applicability. The purpose of the present study was to examine the utility of chromaticity values for estimating lycopene and β-carotene contents in tomato across diverse genetic backgrounds. Measurements of the chromaticity values (L*, a*, b*, C*, h*) were taken on whole fruit and purée of 24 tomato genotypes and were compared with HPLC measurements of fruit lycopene and β-carotene. Examination of different regression models indicated that a model based on the transformed value a*4 from purée measurements explained up to 94.5% of the total variation in fruit lycopene content as measured by HPLC. When this model was applied to a second set of fruit harvested at a later date from the same 24 genotypes, it explained more than 90% of the total variation in lycopene, suggesting its reliability. The best estimation for β-carotene content was obtained by using the b* chromaticity value from whole fruit measurements or the transformed a*2 value from purée measurements. Neither model, however, could explain more than 55% of the variation in β-carotene content, suggesting that chromaticity values may not be appropriate for estimating tomato β-carotene content. The overall results indicated that fruit lycopene content could be measured simply and rather accurately across a wide range of tomato genotypes using chromaticity values taken on fruit purée.

Preferential solvation of the eutectic mixture of liquid crystals E7 in a polysiloxane

Blends of the nematic liquid crystal E7 and poly(methylphenylsiloxane) (PMPS) with molecular weight 120,000 g/mol are investigated by high performance liquid chromatography (HPLC) measurements. This study was prompted by observations made recently while analyzing the phase behavior of poly(siloxane)/E7 and poly(acrylate)/E7 systems. A remarkable increase of the nematic to isotropic transition temperature T NI was found when polymer was added to the liquid crystal. Surprisingly, the increase of T NI was enhanced with the polymer concentration up to 80 wt%, where it reached its highest value. This behavior could be interpreted by invoking a preferential solvation of the constituents of E7 towards the polymer. The present investigation provides an evidence of this phenomenon using HPLC data.

Rapid high-performance liquid chromatographic measurement of buspirone in human plasma after overdose.

For toxicological purposes, a rapid reversed-phase high-performance liquid chromatographic method was developed for the determination of the anxiolytic drug, buspirone, in human plasma. A liquid-liquid procedure was used to extract this compound from plasma in the presence of an internal standard, quinupramine. The analysis was performed on a Spherisorb S5 C8 analytical column with UV detection at 240 nm. No endogenous compounds were found to interfere. A linear response was observed over the concentration range 5-100 ng/mL. A good accuracy (bias < or =7.9%) was achieved for all quality controls, with intra-day and inter-day variation coefficients equal or less than 7.6%. The limit of quantification was 5 ng/mL. Stability of buspirone in plasma stored at different temperatures was checked. This rapid method (run time <12 min) was used to manage an acute poisoning involving buspirone.