Using stable isotopes to inform water resource management in forested and agricultural ecosystems.

Present and future climatic trends are expected to markedly alter water fluxes and stores in the hydrologic cycle. In addition, water demand continues to grow due to increased human use and a growing population. Sustainably managing water resources requires a thorough understanding of water storage and flow in natural, agricultural, and urban ecosystems. Measurements of stable isotopes of water (hydrogen and oxygen) in the water cycle (atmosphere, soils, plants, surface water, and groundwater) can provide information on the transport pathways, sourcing, dynamics, ages, and storage pools of water that is difficult to obtain with other techniques. However, the potential of these techniques for practical questions has not been fully exploited yet. Here, we outline the benefits and limitations of potential applications of stable isotope methods useful to water managers, farmers, and other stakeholders. We also describe several case studies demonstrating how stable isotopes of water can support water management decision-making. Finally, we propose a workflow that guides users through a sequence of decisions required to apply stable isotope methods to examples of water management issues. We call for ongoing dialogue and a stronger connection between water management stakeholders and water stable isotope practitioners to identify the most pressing issues and develop best-practice guidelines to apply these techniques.

Extensive residence in a second language environment modifies perceptual strategies for suprasegmental categorization.

Languages differ in the importance of acoustic dimensions for speech categorization. This poses a potential challenge for second language (L2) learners, and the extent to which adult L2 learners can acquire new perceptual strategies for speech categorization remains unclear. This study investigated the effects of extensive English L2 immersion on speech perception strategies and dimension-selective-attention ability in native Mandarin speakers. Experienced first language (L1) Mandarin speakers (length of U.K. residence > 3 years) demonstrated more native-like weighting of cues to L2 suprasegmental categorization relative to inexperienced Mandarin speakers (length of residence < 1 year), weighting duration more highly. However, both the experienced and the inexperienced Mandarin speakers continued to weight duration less highly and pitch more highly during musical beat categorization and struggled to ignore pitch and selectively attend to amplitude in speech, relative to native English speakers. These results suggest that adult L2 experience can lead to retuning of perceptual strategies in specific contexts, but global acoustic salience is more resistant to change. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

Training auditory processing promotes second language speech acquisition.

Recent evidence suggests that domain-general auditory processing (sensitivity to the spectro-temporal characteristics of sounds) helps determine individual differences in L2 speech acquisition outcomes. The current study examined the extent to which focused training could enhance auditory processing ability, and whether this had a concomitant impact on L2 vowel proficiency. A total of 98 Japanese learners of English were divided into four groups: (1) Auditory-Only (F2 discrimination training); (2) Phonetic-Only (English [æ] and [ʌ] identification training); (3) Auditory-Phonetic (a combination of auditory and phonetic training); and (4) Control training. The results showed that the Phonetic-Only group improved only their English [æ] and [ʌ] identification, while the Auditory-Only and Auditory-Phonetic groups enhanced both auditory and phonetic skills. The results suggest that a learner's auditory acuity to key, domain-general acoustic cues (F2 = 1200-1600 Hz) promotes the acquisition of knowledge about speech categories (English [æ] vs. [ʌ]). (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Amino Acid Fingerprinting of Authentic Nonfat Dry Milk and Skim Milk Powder and Effects of Spiking with Selected Potential Adulterants.

A collaborative study was undertaken in which five international laboratories participated to determine amino acid fingerprints in 39 authentic nonfat dry milk (NFDM)/skim milk powder (SMP) samples. A rapid method of amino acid analysis involving microwave-assisted hydrolysis followed by ultra-high performance liquid chromatography-ultraviolet detection (UHPLC-UV) was used for quantitation of amino acids and to calculate their distribution. The performance of this rapid method of analysis was evaluated and was used to determine the amino acid fingerprint of authentic milk powders. The distribution of different amino acids and their predictable upper and lower tolerance limits in authentic NFDM/SMP samples were established as a reference. Amino acid fingerprints of NFDM/SMP were compared with selected proteins and nitrogen rich compounds (proteins from pea, soy, rice, wheat, whey, and fish gelatin) which can be potential economically motivated adulterants (EMA). The amino acid fingerprints of NFDM/SMP were found to be affected by spiking with pea, soy, rice, whey, fish gelatin and arginine among the investigated adulterants but not by wheat protein and melamine. The study results establish an amino acid fingerprint of authentic NFDM/SMP and demonstrate the utility of this method as a tool in verifying the authenticity of milk powders and detecting their adulteration.

Auditory precision hypothesis-L2: Dimension-specific relationships between auditory processing and second language segmental learning.

Growing evidence suggests a broad relationship between individual differences in auditory processing ability and the rate and ultimate attainment of language acquisition throughout the lifespan, including post-pubertal second language (L2) speech learning. However, little is known about how the precision of processing of specific auditory dimensions relates to the acquisition of specific L2 segmental contrasts. In the context of 100 late Japanese-English bilinguals with diverse profiles of classroom and immersion experience, the current study set out to investigate the link between the perception of several auditory dimensions (F3 frequency, F2 frequency, and duration) in non-verbal sounds and English [r]-[l] perception and production proficiency. Whereas participants' biographical factors (the presence/absence of immersion) accounted for a large amount of variance in the success of learning this contrast, the outcomes were also tied to their acuity to the most reliable, new auditory cues (F3 variation) and the less reliable but already-familiar cues (F2 variation). This finding suggests that individuals can vary in terms of how they perceive, utilize, and make the most of information conveyed by specific acoustic dimensions. When perceiving more naturalistic spoken input, where speech contrasts can be distinguished via a combination of numerous cues, some can attain a high-level of L2 speech proficiency by using nativelike and/or non-nativelike strategies in a complementary fashion.

Development and validation of a hydrophilic interaction chromatography method coupled with a charged aerosol detector for quantitative analysis of nonchromophoric α-hydroxyamines, organic impurities of metoprolol.

The European Pharmacopeia (EP) metoprolol impurities M and N are polar, nonchromophoric α-hydroxyamines, which are poorly retained in a conventional reversed-phase chromatographic system and are invisible for UV detection. Impurities M and N are currently analyzed by TLC methods in the EP as specified impurities and in the United States Pharmacopeia-National Formulary (USP-NF) as unspecified impurities. In order to modernize the USP monographs of metoprolol drug substances and related drug products, a hydrophilic interaction chromatography (HILIC) method coupled with a charged aerosol detector (CAD) was explored for the analysis of the two impurities. A comprehensive column screening that covers a variety of HILIC stationary phases (underivatized silica, amide, diol, amino, zwitterionic, polysuccinimide, cyclodextrin, and mixed-mode) and optimization of HPLC conditions led to the identification of a Halo Penta HILIC column (4.6 × 150 mm, 5 µm) and a mobile phase comprising 85% acetonitrile and 15% ammonium formate buffer (100 mM, pH 3.2). Efficient separations of metoprolol, succinic acid, and EP metoprolol impurities M and N were achieved within a short time frame (<8 min). The HILIC-CAD method was subsequently validated per USP validation guidelines with respect to specificity, robustness, linearity, accuracy, and precision, and could be incorporated into the current USP-NF monographs to replace the outdated TLC methods. Furthermore, the developed method was successfully applied to determine organic impurities in metoprolol drug substance (metoprolol succinate) and drug products (metoprolol tartrate injection and metoprolol succinate extended release tablets).

Characterization of the deoxyguanosine-lysine cross-link of methylglyoxal.

Methylglyoxal is a mutagenic bis-electrophile that is produced endogenously from carbohydrate precursors. Methylglyoxal has been reported to induce DNA-protein cross-links (DPCs) in vitro and in cultured cells. Previous work suggests that these cross-links are formed between guanine and either lysine or cysteine side chains. However, the chemical nature of the methylglyoxal induced DPC have not been determined. We have examined the reaction of methylglyoxal, deoxyguanosine (dGuo), and Nα-acetyllysine (AcLys) and determined the structure of the cross-link to be the N2-ethyl-1-carboxamide with the lysine side chain amino group (1). The cross-link was identified by mass spectrometry and the structure confirmed by comparison to a synthetic sample. Further, the cross-link between methylglyoxal, dGuo, and a peptide (AcAVAGKAGAR) was also characterized. The mechanism of cross-link formation is likely to involve an Amadori rearrangement.

Metabolomics evaluation of the effects of green tea extract on acetaminophen-induced hepatotoxicity in mice.

Green tea has been purported to have beneficial health effects including protective effects against oxidative stress. Acetaminophen (APAP) is a widely used analgesic drug that can cause acute liver injury in overdose situations. These studies explored the effects of green tea extract (GTE) on APAP-induced hepatotoxicity in liver tissue extracts using ultra performance liquid chromatography/quadrupole time-of-flight mass spectrometry and nuclear magnetic resonance spectroscopy. Mice were orally administered GTE, APAP or GTE and APAP under three scenarios. APAP alone caused a high degree of hepatocyte necrosis associated with increases in serum transaminases and alterations in multiple metabolic pathways. The time of GTE oral administration relative to APAP either protected against or potentiated the APAP-induced hepatotoxicity. Dose dependent decreases in histopathology scores and serum transaminases were noted when GTE was administered prior to APAP; whereas, the opposite occurred when GTE was administered after APAP. Similarly, metabolites altered by APAP alone were less changed when GTE was given prior to APAP. Significantly altered pathways included fatty acid metabolism, glycerophospholipid metabolism, glutathione metabolism, and energy pathways. These studies demonstrate the complex interaction between GTE and APAP and the need to employ novel analytical strategies to understand the effects of dietary supplements on pharmaceutical compounds.

Synthesis of the four stereoisomers of 2,3-epoxy-4-hydroxynonanal and their reactivity with deoxyguanosine.

2,3-Epoxy-4-hydroxynonanal (EHN) is a potential product of lipid peroxidation that gives rise to genotoxic etheno adducts. We have synthesized all four stereoisomers of EHN and individually reacted them with 2'-deoxyguanosine. In addition to 1,N(2)-etheno-2'-deoxyguanosine, 12 stereoisomeric products were isolated and characterized by (1)H NMR and circular dichroism spectroscopy. The stereochemical assignments were consistent with selective NOE spectra, vicinal coupling constants, and molecular mechanics calculations. Reversed-phase HPLC conditions were developed that could separate most of the adduct mixture.

Comparison of the in vitro replication of the 7-(2-oxoheptyl)-1,N2-etheno-2'-deoxyguanosine and 1,N2-etheno-2'-deoxyguanosine lesions by Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4).

Oligonucleotides were synthesized containing the 7-(2-oxoheptyl)-etheno-dGuo adduct, which is derived from the reaction of dGuo and the lipid peroxidation product 4-oxo-2-nonenal. The in vitro replication of 7-(2-oxoheptyl)-etheno-dGuo by the model Y-family polymerase Sulfolobus solfataricus P2 DNA Polymerase IV (Dpo4) was examined in two sequences. The extension products were sequenced using an improved LC-ESI-MS/MS protocol developed in our laboratories, and the results were compared to that of the 1,N(2)-etheno-dGuo adduct in the same sequence contexts. Both etheno adducts were highly miscoding when situated in 5'-TXG-3' local sequence contexts with <4% of the extension products being derived from error-free bypass. The major extension products resulted from the misinsertion of Ade opposite the adduct and a one-base deletion. The major extension products from replication of the etheno lesions in a 5'-CXG-3' local sequence context were the result of misinsertion of Ade, a one-base deletion, and error-free bypass. Other minor extension products were also identified. The 7-(2-oxoheptyl)-etheno-dGuo lesion resulted in a larger frequency of misinsertion of Ade, whereas the 1,N(2)-etheno-dGuo gave more of the one-base deletion product. Conformational studies of duplex DNA containing the 7-(2-oxoheptyl)-etheno-dGuo in a 5'-TXG-3' sequence context by NMR indicated the presence of a pH-dependent conformational transition, likely involving the glycosyl bond at the adducted guanosine; the pK(a) for this transition was lower than that observed for the 1,N(2)-epsilon-dGuo lesion. However, the 7-(2-oxoheptyl)-etheno-dGuo lesion, the complementary Cyt, and both flanking base pairs remained disordered at all pH values, which is attributed to the presence of the hydrophobic heptyl group of the 7-(2-oxoheptyl)-etheno-dGuo lesion. The altered pK(a) value and the structural disorder at the 7-(2-oxoheptyl)-etheno-dGuo lesion site, as compared to the same sequence containing the 1,N(2)-etheno-dGuo, may contribute to higher frequency of misinsertion of Ade.

Oxidation and glycolytic cleavage of etheno and propano DNA base adducts.

Non-invasive strategies for the analysis of endogenous DNA damage are of interest for the purpose of monitoring genomic exposure to biologically produced chemicals. We have focused our research on the biological processing of DNA adducts and how this may impact the observed products in biological matrixes. Preliminary research has revealed that pyrimidopurinone DNA adducts are subject to enzymatic oxidation in vitro and in vivo and that base adducts are better substrates for oxidation than the corresponding 2'-deoxynucleosides. We tested the possibility that structurally similar exocyclic base adducts may be good candidates for enzymatic oxidation in vitro. We investigated the in vitro oxidation of several endogenously occurring etheno adducts [1,N(2)-epsilon-guanine (1,N(2)-epsilon-Gua), N(2),3-epsilon-Gua, heptanone-1,N(2)-epsilon-Gua, 1,N(6)-epsilon-adenine (1,N(6)-epsilon-Ade), and 3,N(4)-epsilon-cytosine (3,N(4)-epsilon-Cyt)] and their corresponding 2'-deoxynucleosides. Both 1,N(2)-epsilon-Gua and heptanone-1,N(2)-epsilon-Gua were substrates for enzymatic oxidation in rat liver cytosol; heteronuclear NMR experiments revealed that oxidation occurred on the imidazole ring of each substrate. In contrast, the partially or fully saturated pyrimidopurinone analogues [i.e., 5,6-dihydro-M(1)G and 1,N(2)-propanoguanine (PGua)] and their 2'-deoxynucleoside derivatives were not oxidized. The 2'-deoxynucleoside adducts, 1,N(2)-epsilon-dG and 1,N(6)-epsilon-dA, underwent glycolytic cleavage in rat liver cytosol. Together, these data suggest that multiple exocyclic adducts undergo oxidation and glycolytic cleavage in vitro in rat liver cytosol, in some instances in succession. These multiple pathways of biotransformation produce an array of products. Thus, the biotransformation of exocyclic adducts may lead to an additional class of biomarkers suitable for use in animal and human studies.

Mechanism of 1,N2-etheno-2'-deoxyguanosine formation from epoxyaldehydes.

Background levels of etheno adducts have been attributed to the reaction of DNA with 2,3-epoxyaldehydes, a proposed product of lipid peroxidation. We have examined the reaction of (2R,3S)-epoxyhexanal with dGuo to give 7-(1S-hydroxybutyl)-1,N(2)-etheno-dGuo. We observed that the stereochemistry of the side chain scrambled over time. This process provided insight into the mechanism for the formation of 1,N(2)-etheno-dGuo from 4,5-epoxy-2-decenal [Lee, S. H., et al.(2002) Chem. Res. Toxicol. 15, 300-304]. The mechanistic proposal predicts that 2-octenal is a by-product of the reaction. The reaction of 4,5-epoxy-2-decenal was reinvestigated, and the 2-octenal adduct of dGuo was identified as a product of this reaction in support of the mechanistic proposal. Also observed are products that appear to be derived from 2,3-epoxyoctanal, which can be formed through Schiff base formation of 4,5-epoxy-2-decenal with the dGuo followed by hydration of the double bond and retro-aldol reaction.