Wu Wang

Bilirubin (BR) is a tetrapyrrolic compound stemming from heme catabolism with diverse physiological functions. It can be oxidized by H₂O₂ to form several degradation products, some of which have been detected in vivo and may contribute to the pathogenesis of certain diseases. However, the oxidative degradation of BR is complex and the conditions that BR degradation occurs pathophysiologically remain obscure. Neutrophils are known to generate large amounts of reactive oxygen species, including H₂O₂, upon activation and they are mobilized to inflammatory sites; therefore, we hypothesized that activated neutrophils could cause BR degradation, which could occur at inflammatory sites. In the present study, we investigated BR degradation by H₂O₂ and identified hematinic acid (BHP1) and a new product BHP2, whose structure was characterized as 2,5-diformyl-4-methyl-1H-pyrrole-3-propanoic acid. An LC-MS/MS method for the quantitation of the two compounds was then established. Using the LC-MS/MS method, we observed the concentration-dependent formation of BHP1 and BHP2 in mouse neutrophils incubated with 10 and 30 μM of BR with the yields being 16 ± 3.2 and 31 ± 5.9 pmol/10⁶ cells for BHP1, and 25 ± 4.4 and 71 ± 26 pmol/10⁶ cells for BHP2, respectively. After adding phorbol 12-myristate 13-acetate, a neutrophil agonist, to 30 μM of BR-treated cells, the BHP1 yield increased to 43 ± 6.6 pmol/10⁶ cells, whereas the BHP2 one decreased to 47 ± 9.2 pmol/10⁶ cells. The two products were also detected in hemorrhagic skins of mice with dermal inflammation and hemorrhage at levels of 4.5 ± 1.9 and 0.18 ± 0.10 nmol/g tissue, respectively, which were significantly higher than those in the non-hemorrhagic skins. BHP2 was neurotoxic starting at 0.10 μM but BHP1 was not, as assessed using Caenorhabditis elegans as the animal model. Neutrophil-mediated BR degradation may be a universally pathophysiological process in inflammation and can be particularly important under pathological conditions concerning hemorrhage.

Levoglucosan is a thermal decomposition product of cellulose in particulate matter. δ¹³C value of levoglucosan could be used in studying the combustion mechanisms and chemical pathways. In order to introduce a minimum number of carbon atoms, based on the stereostructure of levoglucosan, a two-step derivatization method with methylboronic acid and MSTFA was developed and carefully optimized. The recommended reaction temperature is 70°C; the reaction time is 60 min for MBA and 120 min for MSTFA derivatization; and the molar ratio of levoglucosan : MBA : MSTFA is 1 : 1: 100 and 1 : 1: 120 and the reagent volume ratio of MSTFA : pyridine is between 1 : 3 and 1 : 4. The developed method achieved excellent reproducibility and high accuracy. The differences in the carbon isotopic compositions of the target boronate trimethysilylated derivative between the measured and calculated ranged from 0.09 to 0.36‰. The standard deviation of measured δ¹³C value of levoglucosan was between 0.22 and 0.48‰. The method was applied to particle samples collected from the combustion of cellulose at four different temperatures. δ¹³C values of levoglucosan in particle samples generated from a self-made combustion setup suggesting that combustion temperature play a little role on isotope fractionation of levoglucosan, although ¹³C enriched in levoglucosan during the combustion process.

The stable carbon isotope compositions of 2-methyltetrols, biomarker compounds for secondary organic aerosols formed from isoprene in the atmosphere, have been determined by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). In this work, isoprene with various δ¹³C values was used to produce 2-methyltetrols via an oxidation reaction with hydrogen peroxide in sulfuric acid under direct sunlight. The target compounds with different stable carbon isotope compositions were then derivatized by methylboronic acid with a known δ¹³C value and measured by GC/C/IRMS. With δ¹³C values of 2-methyltetrols and methylboronic acid predetermined, isotopic fractionation is evaluated for the derivatization process. Through reduplicate δ¹³C measurements, the carbon isotope analysis achieved excellent reproducibility and high accuracy with an average error of 13C values range from −0.10 to 0.29‰, indicating that the derivatization process does not introduce isotopic fractionation. The δ¹³C values of 2-methyltetrols could be calculated on the basis of the stoichiometric mass balance equation among 2-methyltetrols, methylboronic acid, and methylboronate derivatives. Preliminary tests of 2-methyltetrols in PM_2.5 aerosols at two forested sites were conducted and revealed significant differences in their isotope compositions, implying possible application of the method in helping us understand the primary emission, photochemical reaction, or removal processes of isoprene in the atmosphere.

Isoprene is one of the most important non‐methane hydrocarbons (NMHCs) in the troposphere: it is a significant precursor of O ₃ and it affects the oxidative state of the atmosphere. The diastereoisomeric 2‐methyltetrols, 2‐methylthreitol and 2‐methylerythritol, are marker compounds of the photooxidation products of atmospheric isoprene. In order to obtain valuable information on the δ ¹³ C value of isoprene in the atmosphere, the stable carbon isotopic compositions of the 2‐methyltetrols in ambient aerosols were investigated. The 2‐methyltetrols were extracted from filter samples and derivatized with methylboronic acid, and the δ ¹³ C values of the methylboronate derivatives were determined by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). The δ ¹³ C values of the 2‐methyltetrols were then calculated through a simple mass balance equation between the 2‐methyltetrols, methylboronic acid and the methylboronates. The δ ¹³ C values of the 2‐methyltetrols in aerosol samples collected at the Changbai Mountain Nature Reserves in eastern China were found to be −24.66 ± 0.90‰ and −24.53 ± 1.08‰ for 2‐methylerythritol and 2‐methylthreitol, respectively. Based on the measured isotopic composition of the 2‐methyltetrols, the average δ ¹³ C value of atmospheric isoprene is inferred to be close to or slightly heavier than −24.66‰ at the collection site during the sampling period. Copyright © 2010 John Wiley & Sons, Ltd.

The stable carbon isotope compositions of tetrols, erythritol and threitol were determined by gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). Using four tetrols with various δ ¹³ C values derivatized by methylboronic acid, the carbon isotope analysis method achieved excellent reproducibility and high accuracy. There was no carbon isotopic fractionation during the derivatization processes. The differences in the carbon isotopic compositions of methylboronates between the measured and calculated ranged from −0.20 to 0.12‰, within the specification of the GC/C/IRMS system. It was demonstrated that δ ¹³ C values of tetrols could be calculated by a simple mass balance equation between tetrols, methylboronic acid, and methylboronates. The analogous 2‐methyltetrols, marker compounds of photooxidation products of atmospheric isoprene, should have similar behavior using the same derivatization reagent. This method may provide insight on sources and sinks of atmospheric isoprene. Copyright © 2009 John Wiley & Sons, Ltd.