Christine A. Hughey

Identification of acidic NSO compounds in crude oils of different geochemical origins by negative ion electrospray Fourier transform ion cyclotron resonance mass spectrometry

Abstract We present the selective ionization, resolution and identification of acidic NSO compounds in three crude oils of different geochemical origins by negative ion electrospray ionization (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS). Selective ionization by ESI affords direct detection of neutral nitrogen compounds and carboxylic acids in petroleum without pre-chromatographic isolation. Ultra-high resolution/mass accuracy allows detailed and positive identification of acidic NSO compounds in the crude oils. Observed compositional differences reflect known crude oil properties/histories. Collectively, ∼14,000 masses, spanning 18 different heteroatomic classes, are identified unequivocally, demonstrating the potential of ESI-FT-ICR MS for geochemical applications.

Effect of Polar Protic and Polar Aprotic Solvents on Negative-Ion Electrospray Ionization and Chromatographic Separation of Small Acidic Molecules

A comprehensive study investigated the effect of polar protic (methanol and water) and polar aprotic (acetonitrile and acetone) solvents on the chromatographic separation and negative-ion electrospray (ESI) response of 49 diverse small, acidic molecules. Flow injection experiments on a triple quadrupole were used to measure the response in neat solvents after optimization of source conditions and implementation of a rigorous quality control program (the later ensured that changes in analyte response were due to the analyte/solvent measured and not changes in instrument performance over time). In all solvents, compounds with electron-withdrawing groups and extended conjugation ionized best due to resonance and inductive effects. Ionization was greatest in methanol or water for all compounds that elicited a response, thus revealing that enhanced sensitivity and lower limits of detection are achieved with polar protic solvents. Response in acetone was equal to or slightly lower than response in acetonitrile in flow injection experiments; however, the water/acetonitrile and water/acetone mobile phases produced the better chromatographic separation. Water/methanol produced slightly less satisfactory separation but the greatest overall response. This increase in response was attributed to the protic nature of methanol and the elution of compounds in a higher organic mobile phase composition (retention times were ∼30% later in methanol). This work is intended to facilitate rational liquid chromatography/mass spectrometry method development for small molecule applications, including metabolomics.

Naphthenic acids as indicators of crude oil biodegradation in soil, based on semi-quantitative electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry.

Crude oil contaminated soil cores were collected from a basin that contained oily solids left from three decades of oil production. Hydrocarbon biomarker analyses revealed that the soil extracts were moderately biodegraded compared with the non-degraded source oil. The degree of biodegradation also decreased with core depth (7 cm to 1 m). These data were correlated to compositional changes observed in acidic NSO-compounds that were selectively ionized and mass resolved by negative ion electrospray Fourier transform ion cyclotron resonance mass spectrometry (ESI FT-ICR MS). Among the NSO-compounds ionized, the increase in naphthenic acid concentration (e.g., acyclic and alicyclic carboxylic acids) best correlated with the increase in biodegradation (e.g., from non-degraded to moderately degraded) as determined by the hydrocarbon biomarker analyses. The most biodegraded surface extracts (7 cm) exhibited an 80% increase in the abundance of acids relative to the source oil. Use of an internal standard allowed the semi-quantitative determination of the total naphthenic acid concentration, which decreased significantly (P < 0.05) with soil depth. Furthermore, the shift to higher double bond equivalents (DBEs), from acyclic to alicyclic acids, indicated that the increase in acids in the soil extracts was predominantly due to biotic processes. This work demonstrates the potential of ESI FT-ICR MS as a semi-quantitative tool to monitor the production of naphthenic acids during crude oil biotransformation in the environment.

Utilization of Electron Beam Irradiated Almond Skin Powder as a Natural Antioxidant in Ground Top Round Beef

Defatted Carmel variety almond skin powder (ASP) irradiated at 0, 10, 20, and 30 kGy was mixed with ground top round beef at 0.5% (w/w) and stored at 4 degrees C. Color stability, peroxide values (PV), conjugated dienes (CD), thiobarbituric acid reactive species (TBARS), and hexanal were determined periodically over a 2-wk period. The L values were not affected by time or treatment. Initially, the redness (a value) was higher for both the negative control with no ASP (a value of 21.83) and positive control with 0.01% BHT (a value of 22.33) compared to samples that contained the ASP (a values from 17.70 to 20.17) on day 1. This decrease in redness was attributed to the presence of the ASP. Similar to day 1, the a values were not significantly different between the 2 controls over the duration of the study. All the samples with ASP exhibited lower lipid oxidation when compared to the negative control, with greatest oxidation retardation observed at 20 and 30 kGy. Over the treatment period, a 13% to 85% reduction in PV and a 40% to 80% reduction in TBARS were observed in the sample with 30 kGy ASP compared to the negative control. Generally, PV and TBARS of samples with ASP decreased with an increase in irradiation dose. While a difference due to irradiation dose was not observed in CD and hexanal content, the values were significantly lower (P < 0.05) than the controls over time. This study demonstrates that almond skin power could be used to extend the shelf life of refrigerated ground raw beef.

Capillary liquid chromatography–mass spectrometry for the rapid identification and quantification of almond flavonoids

A rapid negative ion ESI high-performance capillary liquid chromatography-mass spectrometry method was developed to identify and quantify flavonoids (e.g., flavanols, flavonols, flavanones and glycosides). Fifteen standards and two varieties of almond skin extract powder (Carmel and Nonpareil) were used to demonstrate the chromatographic separation, reproducibility and accuracy of the method that employed a 150 mm x 0.3 mm ChromXP 3C18-EP-120 column. All standards eluted in less than 10 min, providing a 9-12x reduction in analysis time compared to existing methods (90-120 min). However, isomers (e.g., catechin/epicatechin and galactosides/glucosides) were not resolved and, therefore, identified and quantified collectively. RSDs for retention time and peak area reproducibility (mass spectrometry data) were <0.5% and <5.0%, respectively. Peak area reproducibility was greatly improved (from a RSD>10%) after the implementation of a low-flow metal needle in the ESI source. Quantitation by mass spectrometry also afforded a % error less than 5% for most compounds.

Nonlethal amphibian skin swabbing of cutaneous natural products for HPLC fingerprinting

Small organic molecules found on the skin of amphibians may help impart resistance to pathogens, such as the lethal fungus Batrachochytrium dendrobatidis. The study of these compounds has traditionally required euthanasia of the amphibian, followed by chemical extraction of excised skin. As an alternative method, we report the development and assessment of a non-lethal technique using foam-tipped swabs and HPLC analysis to directly isolate and characterize small molecules found on the skin of amphibians. This protocol was field-tested on Bioko Island, Equatorial Guinea with forty-seven frogs (representing 14 native species). Multiple species (particularly Afrixalus paradorsalis and Didynamipus sjostedti) carried sets of species-specific compounds (i.e., a chromatographic fingerprint). A principal coordinate analysis (PCO) of the commonly occurring compounds detected across all species revealed a significant relationship between chromatographic profile and species for all swab samples.

Extraction, identification, and quantification of flavonoids and phenolic acids in electron beam-irradiated almond skin powder.

The effect of electron beam irradiation doses from 0 to 30 kGy on extraction yield and phenolic compounds was evaluated in almond skin phenolic extracts (ASPE). Total soluble phenols and distribution of phenolic compounds from acidified methanol ASPE and 52% methanol ASPE were quantified using Folin-Ciocalteau method, liquid chromatography with diode array and fluorescence detection, and negative ion electrospray-mass spectrometry. Electron beam irradiation increased extraction yield by as much as 23%, with the greatest increase observed in the acidified methanol ASPE. Irradiated samples extracted with acidified methanol also exhibited an increase in extractable phenols (Folin-Ciocalteau) and total HPLC-resolved phenolics at all irradiation doses. Samples extracted with 52% methanol exhibited an increase at 10 and 20 kGy, but a 31% decrease at 30 kGy. An increase in aglycones respective to their glycosides was not observed with irradiation. Therefore, the increase in phenolics was attributed to release of phenolics from their cellular matrix.

Time of Day and Training Status Both Impact the Efficacy of Caffeine for Short Duration Cycling Performance

This project was designed to assess the effects of time of day and training status on the benefits of caffeine supplementation for cycling performance. Twenty male subjects (Age, 25 years; Peak oxygen consumption, 57 mL·kg−1·min−1) were divided into tertiles based on training levels, with top and bottom tertiles designated as ‘trained’ (n = 7) and ‘untrained’ (n = 7). Subjects completed two familiarization trials and four experimental trials consisting of a computer-simulated 3-km cycling time trial (TT). The trials were performed in randomized order for each combination of time of day (morning and evening) and treatment (6mg/kg of caffeine or placebo). Magnitude-based inferences were used to evaluate all treatment effects. For all subjects, caffeine enhanced TT performance in the morning (2.3% ± 1.7%, ‘very likely’) and evening (1.4% ± 1.1%, ‘likely’). Both untrained and trained subjects improved performance with caffeine supplementation in the morning (5.5% ± 4.3%, ‘likely’; 1.0% ± 1.7%, ‘likely’, respectively), but only untrained subjects rode faster in the evening (2.9% ± 2.6%, ‘likely’). Altogether, our observations indicate that trained athletes are more likely to derive ergogenic effects from caffeine in the morning than the evening. Further, untrained individuals appear to receive larger gains from caffeine in the evening than their trained counterparts.