Quantitation Experiments Research Articles

Quantitative Antioxidant Profiling Throughout Beer Brewing Followed by Discovery and Isolation of Precursors from Barley (Hordeum vulgare L.).

The application of high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) revealed the origin and evolution of antioxidants during the brewing process of hopped and unhopped reference beer. As tachioside (3-methoxy-4-hydroxyphenyl-β-d-glucopyranoside), arbutin (4-hydroxyphenyl-β-d-glucopyranoside), and hordatines clearly increased during the fermentation step, the raw material barley was investigated as a source of the corresponding precursors. Therefore, 4-hydroxyphenyl-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside, 4-hydroxy-3-methoxyphenyl-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside, 4-hydroxy-3-methoxyphenyl-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside, and 4-hydroxy-2-methoxyphenyl-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside were isolated from barley for the first time, and identified using liquid chromatography-mass spectrometry (LC-MS) and one-dimensional/two-dimensional-nuclear magnetic resonance (1D/2D-NMR) experiments. Moreover, hordatine glucosides A, B, and C were isolated and identified from barley, and hordatine C glucoside was characterized for the first time. A fermentation model followed by HPLC-MS/MS analysis substantiated the release of tachioside from 4-hydroxy-3-methoxyphenyl-β-d-glucopyranosyl-(1 → 6)-β-d-glucopyranoside by Saccharomyces cerevisiae. Quantitation experiments monitoring the content in wheat, barley, and different barley malt types demonstrated a wide range of concentrations, providing a basis for further comprehensive investigations to optimize the antioxidant yield in beer to contribute to improved flavor stability.

Abstract 7420: Optimizing proteomic data access and analysis in the cloud: Leveraging Terra's integration with the Proteomic Data Commons

Abstract The Proteomic Data Commons (PDC) hosts cancer proteomics data with the goal of making this data available to the public to support development of cancer diagnostics, treatment, and progression tracking. As a part of the Cancer Research Data Commons (CRDC), the Terra platform provides a cloud workbench for the PDC data. FireCloud is a Broad Institute project funded by NCI to empower cancer researchers to access data, run analysis tools and collaborate securely in the cloud. It is powered by Terra, a secure, scalable cloud-native platform developed by the Broad Institute, Microsoft, and Verily, an Alphabet company. It provides batch workflow execution, interactive analysis including data visualization, and ~2,900 publicly available tools with the ability to import more tools from Dockstore. The integration of PDC and Terra enables researchers to leverage the data navigation and file-level search capabilities on the PDC web browser (pdc.cancer.gov) and export selected data manifests to a Terra workspace. This integrated hand-off specifically allows metadata and cloud links to PDC data in Portable Format for Bioinformatics (PFB) to be transferred to Terra and used in analysis with cloud resources. A featured workspace for Terra-PDC integration includes tools for (i) downloading data files and relevant additional metadata; (ii) organizing the data and metadata for running the FragPipe workflow, a comprehensive collection of tools for reading and processing raw Mass Spec (MS) data; and (iii) implementing a pre-configured FragPipe workflow to process isobarically labeled MS data from Tandem Mass Tag (TMT) or Isobaric tags for relative and absolute quantitation (iTRAQ) experiments using information automatically imported from PDC via the PFB import. The pipelines can also be customized as needed to process any type of raw MS data the PDC supports. In addition, Terra connects to the NCI Genomic Data Commons (GDC) and hosts The Cancer Genome Atlas (TCGA) and Therapeutically Applicable Research to Generate Effective Treatments (TARGET) datasets allowing users to pull in other data types for co-analysis. As a whole, using the CRDC cloud resources, in conjunction with Terra, allows analysis of data on a massive scale, enabling multi-omic integration spanning the genomic and proteomic commons data, with easy sharing of data and tools. Citation Format: Emily LaPlante, Bingxing Huo, DR Mani, Ratna R. Thangudu. Optimizing proteomic data access and analysis in the cloud: Leveraging Terra's integration with the Proteomic Data Commons [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7420.

Quantitative LC-MS/MS. 1. Impact of Points across a Peak on the Accuracy and Precision of Peak Area Measurements.

The number of points across a chromatographic peak has long been recognized as a key determinant of the accuracy and precision of the measured peak area. In LC-MS-based quantitation experiments in drug discovery and development, the "rule-of-thumb" has been to use 15 or more points. This "rule" is based on the literature describing chromatographic methods where the goal was to achieve the lowest possible imprecision in the measurements, especially when unknown analytes are being detected. Restricting methods to the requirement of at least 15 points across a peak can be detrimental to the development methods that fully optimize the signal-to-noise ratio for the assay using longer dwell times and/or transition summing. This study aims to show that 7 points across the peak for peaks that are 9 s or less wide provide more than sufficient accuracy and precision for drug quantitation studies. Data from simulated Gaussian curves using a sampling interval of 7 points across the peak gave peak area calculations within 1% of the expected total peak area using the Trapezoidal and Riemann rules and 0.6% for the Simpson rule. Low and high concentration samples (n = 5) were assayed using three different LC methods on three different days on two different instruments (API5000 and API5500). The difference in peak area (%ΔPA) and relative standard deviation of the peak areas (%RSD) was less than ∼5%. No significant difference was observed from the data that were obtained from different sampling intervals, different peak widths, different days, different peak sizes, and different instruments. Three core analytical runs were performed on three different days. In each core run, the lower limit of quantitation (LLOQ, n = 5), low quality control (LQC, n = 5), middle quality control (MQC, n = 5), and high-quality control samples (HQC, n = 5) were processed and run simultaneously with a standard curve. The range of the intra- and interday accuracy and precision for 3 core runs was 98.0-105% and 0.9-3.0% for 7 data points and 97.5-105% and 0.8-4.3% for 17 data points, respectively. No significant difference was observed for the different sampling intervals. The results show a sampling interval of 7 points for peaks up to 9 s wide is sufficient to define a peak accurately and precisely for drug quantitation studies in drug discovery and development.

Quantitative analysis of polycyclic aromatic hydrocarbons using high-performance liquid chromatography-photodiode array-high-resolution mass spectrometric detection platform coupled to electrospray and atmospheric pressure photoionization sources.

Polycyclic aromatic hydrocarbons (PAHs) are common pollutants present in atmospheric aerosols and other environmental mixtures. They are of particular air quality and human health concerns as many of them are carcinogenic toxins. They also affect absorption of solar radiation by aerosols, therefore contributing to the radiative forcing of climate. For environmental chemistry studies, it is advantageous to quantify PAH components using the same analytical technics that are commonly applied to characterize a broad range of polar analytes present in the same environmental mixtures. Liquid chromatography coupled with photodiode array and high-resolution mass spectrometric detection (LC-PDA-HRMS) is a method of choice for comprehensive characterization of chemical composition and quantification of light absorption properties of individual organic compounds present in the environmental samples. However, quantification of non-polar PAHs by this method is poorly established because of their imperfect ionization in electrospray ionization (ESI) technique. This tutorial article provides a comprehensive evaluation of the quantitative analysis of 16 priority pollutant PAHs in a standard reference material using the LC-MS platform coupled with the ESI source. Results are further corroborated by the quantitation experiments using an atmospheric pressure photoionization (APPI) method, which is more sensitive for the PAH detection. The basic concepts and step-by-step practical guidance for the PAHs quantitative characterization are offered based on the systematic experiments, which include (1) Evaluation effects of different acidification levels by formic acid on the (+)ESI-MS detection of PAHs. (2) Comparison of detection limits in ESI+ versus APPI+ experiments. (3) Investigation of the PAH fragmentation patterns in MS2 experiments at different collision energies. (4) Calculation of wavelength dependent mass absorption coefficient (MACλ) of the standard mixture and its individual PAHs using LC-PDA data. (5) Assessment of the minimal injected mass required for accurate quantification of MACλ of the standard mixture and of a multi-component environmental sample.

Carbon nanodot-hybridized silica nanospheres assisted immunoassay for sensitive detection of Escherichia coli

Carbon dots (CDs) are intrinsically luminescent nanomaterials that have many potential applications in biosensing technologies due in part to their high photostability and fluorescent properties. However, attempts to integrate CDs into immunosorbent assays have been deterred by challenges preventing quantum yield augmentation and surface functionalization. To address these issues, we fabricated carbon dot-encapsulated silica nanospheres (CSNs) using a facile one-pot synthesis method. The enclosing silica matrix rendered extensive material stability to the nanospheres and abated quenching by nonradiative decay to cause considerable signal amplification. Nanosphere hybridization to antibodies permitted their use in solid-phase immunoassays as tracers. Here, we demonstrate the suitability of CSN-based immunosorbent assays as a point-of-care technique for the detection of foodborne pathogens through E. coli O157:H7 quantitation experiments. After optimization, detection limit of E. coli O157:H7 was determined to be 2.4 CFU mL−1. The estimated recoveries were in the range of 91.7–110.5% in spiked samples, which indicated that the developed method is capable for detecting E. coli O157:H7 in food samples. The nanosphere tracers described herein, and the methods used to create them, may be beneficial tools for the development of new pathogen biosensing strategies.

Targeted Quantitation Mode Comparison of Haloacetic Acids, Bromate, and Dalapon in Drinking Water Using Ion Chromatography Coupled to High-Resolution (Orbitrap) Mass Spectrometry

A highly selective, sensitive, and simple analytical method for identification and quantification of nine haloacetic acids, bromate, and dalapon has been developed. This method uses ion chromatography (IC) coupled with electrospray ionization-high-resolution mass spectrometry (IC-ESI-HRMS) to directly analyze water samples on a high capacity anion-exchange column, eliminating the need for sample pretreatment/derivatization. Our study compared the following three types of targeted quantitation experiments using a quadrupole–orbitrap hybrid mass spectrometer, full-scan MS with data-dependent tandem mass spectrometry (full MS/dd-MS2 with inclusion list), targeted selected ion monitoring (SIM) with data-dependent tandem mass spectrometry (t-SIM/dd-MS2), and parallel reaction monitoring (PRM). Sensitivity, linearity, accuracy, and precision were validated following the guidelines of U.S. EPA Method 557. Single laboratory lowest concentration minimum reporting levels (LCMRLs) for the analytes using three different acquisition modes ranged from 0.0011 to 0.18 μg/L. All three quantitation modes showed good linearity for the eleven analytes with coefficients of determination of 0.9981- 0.9993. This IC-ESI-HRMS method was successfully applied to the analysis of commercial bottled water, tap water from San Francisco Bay Area, and the same tap water that has been through a filtered drinking water faucet. Both t-SIM/dd-MS2 and PRM modes were sensitive to confirm the trace-level presence of all nine HAAs, bromate, and dalapon in the tap water sample. Full-scan HRMS data acquisition provided the benefits of simultaneous data collection for both targeted and non-targeted components, and thus, suitability for simultaneous quantification of an unlimited number of compounds. Data-dependent MS/MS (dd-MS2) product-ion spectra were used for confirmation. All three modes showed good quantitative performance and obtained similar values. Single laboratory precision and accuracy data are presented for three water matrices: reagent water, laboratory synthetic sample matrix (LSSM), and tap water. Single laboratory precision was 0.078- 8.04%, and accuracy was in the range 70–130% for the three MS modes.

Key Odor-Active Compounds in Raw Green and Red Toona sinensis (A. Juss.) Roem. and Their Changes during Blanching.

Application of aroma extract dilution analysis and headspace aroma dilution analysis revealed 51 odorants in raw green Toona sinensis and 54 odorants in raw red T. sinensis in the flavor dilution factor range of 8-4096. (E,E)-2,4-Decadienal, nonanal, 2,3,5-trimethylpyrazine, (E,Z)- and (Z,Z)-di-1-propenyl trisulfide, 2-methoxyphenol, and 4-ethylphenol were first identified as key odorants of T. sinensis. Clear differences between green and red T. sinensis in aroma profiles, flavor dilution factors, quantitative data, and odor activity values verified that (E,E)-, (E,Z)-, and (Z,Z)-di-1-propenyl disulfide, (E,E)-, (E,Z)- and (Z,Z)-di-1-propenyl trisulfide, cis- and trans-2-mercapto-3,4-dimethyl-2,3-dihydrothiophene, and dimethyl sulfide caused the distinct sulfury odor note of each variety. Further, hexanal, (E)-2-hexenal, (E)-2-hexen-1-ol, and (E,Z)-2,6-nonadienal led to the green odor note in green T. sinensis, while 2-methoxyphenol and 4-ethylphenol contributed to the intense phenolic aroma note in red T. sinensis. Quantitation experiments and triangle tests in blanched T. sinensis verified that the quick loss of the abovementioned sulfur-containing compounds, aldehydes, the alcohol (E)-2-hexen-1-ol, and phenols was responsible for the changes in the overall aroma profile during blanching.

Cancerous inhibitor of protein phosphatase 2A (CIP2A) modifies energy metabolism via 5' AMP-activated protein kinase signalling in malignant cells.

Cancerous inhibitor of protein phosphatase 2A (CIP2A) is an adverse biomarker across many malignancies. Using K562 cells engineered to have high or low CIP2A expression, we show that high CIP2A levels significantly bias cellular energy production towards oxidative phosphorylation (OXPHOS) rather than glycolysis. Mass spectrometric analysis of CIP2A interactors and isobaric tagging for relative and absolute protein quantitation (ITRAQ) experiments identified many associated proteins, several of which co-vary with CIP2A level. Many of these CIP2A associating and co-varying proteins are involved in energy metabolism including OXPHOS, or in 5' AMP-activated protein kinase (AMPK) signalling, and manipulating AMPK activity mimics the effects of low/high CIP2A on OXPHOS. These effects are dependent on the availability of nutrients, driven by metabolic changes caused by CIP2A. CIP2A level did not affect starvation-induced AMPK phosphorylation of Unc-51 autophagy activating kinase 1 (ULK-1) at Ser555, but autophagy activity correlated with an increase in AMPK activity, to suggest that some AMPK processes are uncoupled by CIP2A, likely via its inhibition of protein phosphatase 2A (PP2A). The data demonstrate that AMPK mediates this novel CIP2A effect on energy generation in malignant cells.

Characterization of the key aroma compounds in white bread by aroma extract dilution analysis, quantitation, and sensory evaluation experiments

The aroma compounds in four brands of white bread were characterized by aroma extraction dilution analysis, quantitation, aroma recombination, and omission tests. A total of 54 aroma compounds were detected. Among them, methional had the highest flavor dilution (FD) factor of 729, followed by (E,E)‐2,4‐decadienal (FD = 243), furaneol (FD = 243), and δ‐decalactone (FD = 243). Thirty‐nine compounds (FD ≥ 9) were quantitated then analyzed by odor activity values (OAVs), 31 of which had OAVs ≥ 1. Four recombination models showed a similarity of 90.35% to the corresponding white bread samples. Omission tests confirmed that (E,E)‐2,4‐decadienal (OAV = 1,229), 4‐methyl‐5‐thiazoleethanol (OAV = 274), δ‐decalactone (OAV = 70), furaneol (OAV = 40), 2‐phenylethanol (OAV = 20), and methional (OAV = 12) were the key aroma compounds in white bread. Additionally, the alcohols and acids also significantly (p < 0.001) contributed to the sour, sweet, and flour‐like notes of white bread. PRACTICAL APPLICATIONS: In this work, the key aroma compounds of white bread were confirmed. The content of aromas was significantly different among four brands of white bread with different recipes and processing techniques. These results provide an important theoretical basis for the improvement of quality control and processing of white bread.

Kaempferol 3- O-(2‴- O-Sinapoyl-β-sophoroside) Causes the Undesired Bitter Taste of Canola/Rapeseed Protein Isolates.

By means of activity-guided fractionation using taste dilution analysis, liquid chromatography-tandem mass spectrometry (LC-MS/MS), liquid chromatography-time-of-flight mass spectrometry, one-/two-dimensional nuclear magnetic resonance spectroscopy, LC-MS/MS quantitation, dose-over-threshold considerations, and sensory spiking experiments, kaempferol 3- O-(2‴- O-sinapoyl-β-sophoroside), exhibiting a bitter taste above the low threshold concentration of 3.4 μmol/L, was found for the first time as the key molecule contributing to the unpleasant bitter taste of rapeseed (canola) protein isolates. This finding opens new avenues for a biorefinery approach targeting an off-taste removal.

Comprehensive Single-Shot Proteomics with FAIMS on a Hybrid Orbitrap Mass Spectrometer.

Liquid chromatography (LC) prefractionation is often implemented to increase proteomic coverage; however, while effective, this approach is laborious, requires considerable sample amount, and can be cumbersome. We describe how interfacing a recently described high-field asymmetric waveform ion mobility spectrometry (FAIMS) device between a nanoelectrospray ionization (nanoESI) emitter and an Orbitrap hybrid mass spectrometer (MS) enables the collection of single-shot proteomic data with comparable depth to that of conventional two-dimensional LC approaches. This next generation FAIMS device incorporates improved ion sampling at the ESI-FAIMS interface, increased electric field strength, and a helium-free ion transport gas. With fast internal compensation voltage (CV) stepping (25 ms/transition), multiple unique gas-phase fractions may be analyzed simultaneously over the course of an MS analysis. We have comprehensively demonstrated how this device performs for bottom-up proteomics experiments as well as characterized the effects of peptide charge state, mass loading, analysis time, and additional variables. We also offer recommendations for the number of CVs and which CVs to use for different lengths of experiments. Internal CV stepping experiments increase protein identifications from a single-shot experiment to >8000, from over 100 000 peptide identifications in as little as 5 h. In single-shot 4 h label-free quantitation (LFQ) experiments of a human cell line, we quantified 7818 proteins with FAIMS using intra-analysis CV switching compared to 6809 without FAIMS. Single-shot FAIMS results also compare favorably with LC fractionation experiments. A 6 h single-shot FAIMS experiment generates 8007 protein identifications, while four fractions analyzed for 1.5 h each produce 7776 protein identifications.

Abstract 5536: Simultaneous characterization and quantification of immune cell subpopulations and PD-L1 expressing CTCs in peripheral blood of cancer patients

Abstract Background: Expression of PD-L1 on tumor and immune markers in tumor tissue is associated with improved response to PD-1 and PD-L1 checkpoint inhibitors. However, each alone has limited predictive utility. Multimodal characterization of both the tumor and host immune system is an unmet medical need for the improved prediction of response to immunotherapy. Metastatic lesions are likely to be undersampled and require a liquid biopsy, given tumor heterogeneity and evolution and temporal changes in the host immune system. We sought to examine expression of PD-L1 on circulating tumor cells (CTCs) as well as characterize rare immune cell populations with a noninvasive liquid biopsy. Examining dynamic biomarker changes in longitudinal samples could enable the development of novel diagnostic tools for response prediction and pharmacodynamics studies related to immunotherapy. Methods: Blood samples from lung cancer patients were collected and shipped to Epic Sciences. Contrived samples were also developed, using cancer cell lines spiked into healthy donor (HD) blood. Red blood cells were lysed and nucleated cells were plated onto glass slides. Slides were stained with DAPI and immune cocktails, then imaged. Targets included pan-CK, CD45, PD-L1, CD4, CD8, Ki-67, and TIM-3. Approximately 3 million nucleated cells per slide were examined through advanced digital pathology pipelines to detect and quantify changes in T-cell populations and to assess circulating tumor burden. Results: Epic Sciences' rare cell detection platform has an analytically validated limit of detection of 2 cells/mL of blood. Immuno-panels were developed to profile leukocyte subpopulations and CTCs from a single blood sample and feasibility was demonstrated in cell lines, HD and lung disease patient blood. Quantitation experiments showed that immunomagnetically purified CD8+ and CD4+ cells spiked in 3:1, 1:1, and 1:3 ratios were detected as 3.1:1, 1:1.1, and 1:3.1. Purified CD4 cells were also spiked into healthy donor WBCs at target ratios of 1-10%. Percentages of detected spike-in CD4 cells were linearly correlated with the target ratios with correlation coefficient (r) = 0.96. 22.8% (18/79) lung cancer patients had PD-L1+ CTCs detected, and the majority (91%) of patients had PD-L1+ leukocyte populations. Conclusions: The low limit of detection of the Epic Sciences CTC platform coupled with the ability to archive patient blood samples allowed for retrospective, precise quantification of leukocyte subpopulations and PD-L1 expression on CTCs. Development of a liquid biopsy-based platform that can simultaneously measure immune biomarkers in CTCs as well as on leukocytes will allow for real-time assessment and monitoring of response to immune checkpoint inhibitors. Citation Format: Rachel Krupa, David Lu, Adam Jendrisak, Angel Rodriguez, Nadia Ebrahim, Robin Richardson, Sean Nisperos, Ryon Graf, Jiyun Byun, Yipeng Wang, Mark Landers, Ryan Dittamore. Simultaneous characterization and quantification of immune cell subpopulations and PD-L1 expressing CTCs in peripheral blood of cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5536.

Characterization and comparison of key aroma compounds in raw and dry porcini mushroom (Boletus edulis) by aroma extract dilution analysis, quantitation and aroma recombination experiments

A study was carried out to determine and compare the key aroma compounds in raw and dry porcini mushroom (Boletus edulis). The volatile fractions were prepared by solvent-assisted flavor evaporation (SAFE), and aroma extract dilution analysis (AEDA) combined with gas chromatography-mass spectrometry (GC–MS) was employed to identify the odorants. Selected aroma compounds were quantitated and odor activity values (OAVs) were calculated revealing OAVs ≥ 1 for 12 compounds in raw porcini, among which 1-octen-3-one showed the highest OAV. In addition to compounds with eight carbon atoms, 3-methylbutanal, (E,E)-2,4-decadienal and (E,E)-2,4-nonadienal were also responsible for the unique aroma profile. In dry mushroom OAVs ≥ 1 were obtained for 20 odorants. Among them, 3-(methylthio)propanal, 1-octen-3-one and pyrazines were determined as predominant odorants. Overall, drying increased complexity of volatile compounds, thus significantly changing the aroma profile of porcini, providing more desirable roasted and seasoning-like flavor and less grass-like and earthy notes.

Non-specific Digestion Artifacts of Bovine Trypsin Exemplified with Surrogate Peptides for Endogenous Protein Quantitation

In bottom-up MS-based proteomic workflows, peptides are used as surrogates of proteins either for identification or quantification. Trypsin is the most preferred and widely used protease to generate these surrogate peptides for peptide based quantitative targeted proteomics. During one of such surrogate based protein quantitations, one of the experiments to understand the stability of the selected surrogate peptides during the process of digestion, yielded very low recovery (2%) compared to an equimolar amount of the pure peptide. This prompted us to investigate the reasons using high-resolution mass spectrometry (Q-Exactive plus) with data dependent MS2 acquisition (ddMS2) and parallel reaction monitoring (PRM) experiments. Investigations revealed non-specific cleavage of the peptide bond predominantly after phenylalanine (F) and tyrosine (Y) amino acid residues of the surrogate peptide standard by trypsin. PRM data showed high abundance of these truncated peptides compared to full length peptide, thus explaining the low recovery of peptide standards during the digestion process. Further investigations suggested a possible role of pseudo-trypsin (with chymotrypsin like specificity), which is likely formed from autolysis of trypsin during the course of digestion. Sequence grade trypsin (TPCK treated and reductively methylated) did not show any such non-specific cleavage of the peptides. Our findings, therefore, demonstrate the importance of choosing the appropriate trypsin for protein quantitation experiments. The propensity of trypsin that can form pseudotrypsin can produce non-specific digestion artifacts that can under predict the protein concentrations in surrogate peptide based quantification methods.

Evaluation of Key Aroma Compounds in Processed Prawns (Whiteleg Shrimp) by Quantitation and Aroma Recombination Experiments.

In our previous study on the aroma compounds of heated prawn meat, the main odorants in blanched (BPM) and fried prawn meat (FPM), respectively, were characterized by means of gas chromatography-olfactometry and aroma extract dilution analysis. In this follow-up study, these aroma compounds were quantified by means of stable isotope dilution assays, and odor activity values (OAV; ratio of concentration to odor detection threshold) were calculated. Results revealed 2-acetyl-1-pyrroline and (Z)-1,5-octadien-3-one as the most potent odor-active compounds in both prawn samples. In FPM, as compared to BPM, higher OAVs were determined for 2-acetyl-1-pyrroline, 2-acetyl-2-thiazoline, 3-methylbutanal, 3-(methylthio)propanal, phenylacetaldehyde, 3-hydroxy-4,5-dimethyl-2(5H)-furanone, 4-hydroxy-2,5-dimethyl-3(2H)-furanone, 2,3-diethyl-5-methylpyrazine, and trimethylpyrazine. Aroma recombination experiments corroborated that the overall aroma of the blanched as well as the fried prawn meat, respectively, could well be mimicked by the set of key odorants quantitated in this study.

Insights into the Key Compounds of Durian (Durio zibethinus L. 'Monthong') Pulp Odor by Odorant Quantitation and Aroma Simulation Experiments.

Sixteen compounds, previously identified as potent odorants by application of an aroma extract dilution analysis and the gas chromatography-olfactometry analysis of static headspace samples, were quantitated in the pulp of durians, variety Monthong, and odor activity values (OAVs) were calculated by dividing the concentrations obtained by the odor thresholds of the compounds in water. In combination with data recently reported for hydrogen sulfide and short-chain alkanethiols, OAVs > 1 were obtained for 19 compounds, among which ethyl (2S)-2-methylbutanoate (fruity; OAV 1700000), ethanethiol (rotten onion; OAV 480000), and 1-(ethylsulfanyl)ethane-1-thiol (roasted onion; OAV 250000) were the most potent, followed by methanethiol (rotten, cabbage; OAV 45000), ethane-1,1-dithiol (sulfury, durian; OAV 23000), and ethyl 2-methylpropanoate (fruity; OAV 22000). Aroma simulation and omission experiments revealed that the overall odor of durian pulp could be mimicked by only two compounds, namely, ethyl (2S)-2-methylbutanoate and 1-(ethylsulfanyl)ethane-1-thiol, when combined in their natural concentrations.

Odour-active compounds in the traditional Armenian soup seasoning herb Heracleum transcaucasicum

The volatile fraction isolated from dried shoots of Heracleum transcaucasicum Manden (syn. Heracleum pastinacifolium C. Koch) by solvent extraction and solvent-assisted flavour evaporation was screened for odour-active compounds by application of an aroma extract dilution analysis. In the flavour dilution (FD) factor range of 1–2048, 51 odorants were detected, among which soup seasoning-like smelling sotolon (3-hydroxy-4,5-dimethyl-2(5H)-furanone) showed the highest FD factor (2048). Its characteristic odour, which closely matched the overall aroma of the dried shoots, in combination with its high FD factor and the absence of any further odour-active compound exhibiting a similar smell, suggested that sotolon constitutes the character impact compound in the aroma of H. transcaucasicum. Stable isotope dilution quantitation using (13C2)sotolon as internal standard revealed a concentration of ~12 mg/kg sotolon in dried H. transcaucasicum shoots, which is several orders of magnitude higher than its odour detection threshold. Further quantitation experiments revealed that during traditional culinary use, sotolon is transferred from H. transcaucasicum shoots in odour-active amounts to Armenian karshm soup.

Harbin: a quantitation PCR analysis tool

To enable analysis and comparisons of different relative quantitation experiments, a web-browser application called Harbin was created that uses a quantile-based scoring system for the comparison of samples at different time points and between experiments. Harbin uses the standard curve method for relative quantitation to calculate concentration ratios (CRs). To evaluate if different datasets can be combined the Harbin quantile bootstrap test is proposed. This test is more sensitive in detecting distributional differences between data sets than the Kolmogorov-Smirnov test. The utility of the test is demonstrated in a comparison of three grapevine leafroll associated virus 3 (GLRaV-3) RT-qPCR data sets. The quantile-based scoring system of CRs will enable the monitoring of virus titre or gene expression over different time points and be useful in other genomic applications where the combining of data sets are required.

Dual visible-light active components containing self-doped Bi2O2CO3/g-C3N4 2D-2D heterojunction with enhanced visible-light-driven photocatalytic activity

The (CO3)2− self-doped Bi2O2CO3/g-C3N4 (C-BOC/g-C3N4) unique 2D-2D heterostructure has been developed via a mixed-calcination method. This heterostructure is confirmed by X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), SEM-mapping, and diffuse reflection spectroscopy (DRS). Due to the charge interaction, it is interesting to observe that C-BOC nanosheets are assembled on g-C3N4 in a surface–surface coupling way, which enables an intimate interfacial interaction between the two components. The photocatalytic activity is evaluated by photodegradation of Rhodamine B (RhB) under visible light (λ>420nm). It is found that all the C-BOC/g-C3N4 composites showed highly enhanced photocatalytic activity. Electrochemical impedance spectra (EIS) measurement revealed that the largely promoted charge transfer derived from the well matchable band structure and intimate interfacial interactions between the two visible-light active components is responsible for the photoactivity enhancement. Besides, the active species trapping and O2− quantitation experiments are conducted to disclose the photocatalytic mechanism.

Insights into the Key Aroma Compounds in Mango (Mangifera indica L. 'Haden') Fruits by Stable Isotope Dilution Quantitation and Aroma Simulation Experiments.

Thirty-four aroma-active compounds, previously identified with high flavor dilution factors by application of an aroma extract dilution analysis, were quantified in tree-ripened fruits of mango (Mangifera indica L. 'Haden'). From the results, the odor activity value (OAV) was calculated for each compound as the ratio of its concentration in the mangoes to its odor threshold in water. OAVs > 1 were obtained for 24 compounds, among which ethyl 2-methylbutanoate (fruity; OAV 2100), (3E,5Z)-undeca-1,3,5-triene (pineapple-like; OAV 1900), ethyl 3-methylbutanoate (fruity; OAV 1600), and ethyl butanoate (fruity; OAV 980) were the most potent, followed by (2E,6Z)-nona-2,6-dienal (cucumber-like), ethyl 2-methylpropanoate (fruity), (E)-β-damascenone (cooked apple-like), ethyl hexanoate (fruity), 4-hydroxy-2,5-dimethyl-3(2H)-furanone (caramel-like), 3-methylbut-2-ene-1-thiol (sulfurous), γ-decalactone (peach-like), β-myrcene (terpeny), (3Z)-hex-3-enal (green), 4-methyl-4-sulfanylpentan-2-one (tropical fruit-like), and ethyl octanoate (fruity). Aroma simulation and omission experiments revealed that these 15 compounds, when combined in a model mixture in their natural concentrations, were able to mimic the aroma of the fruits.