Affinity purification coupled to mass spectrometry (AP-MS) is a powerful method to analyze protein-protein interactions (PPIs). The AP-MS approach provides an unbiased analysis of the entire protein complex and is useful to identify indirect interactors. However, reliable protein identification from the complex AP-MS experiments requires appropriate control of false identifications and rigorous statistical analysis. Another challenge that can arise from AP-MS analysis is to distinguish bona fide interacting proteins from the non-specifically bound endogenous proteins or the "background contaminants" that co-purified by the bait experiments. In this chapter, we will first describe the protocol for performing in-solution trypsinization for the samples from the AP experiment followed by LC-MS/MS analysis. We will then detail the MaxQuant workflow for protein identification and quantification for the PPI data derived from the AP-MS experiment. Finally, we describe the CRAPome interface to process the data by filtering against contaminant lists, score the interactions and visualize the protein interaction networks.
This work employed the model protein β-lactoglobulin (BLG) to investigate the contribution of microstructural changes to regulating the interaction patterns between protein and flavor compounds through employing computer simulation and multi-spectroscopic techniques. The formation of molten globule (MG) state-like protein during the conformational evolution of BLG, in response to ultrasonic (UC) and heat (HT) treatments, was revealed through multi-spectroscopic characterization. Differential MG structures were distinguished by variations in surface hydrophobicity and the microenvironment of tryptophan residues. Fluorescence quenching measurements indicated that the formation of MG enhanced the binding affinity of heptanal to protein. LC-MS/MS and NMR revealed the covalent bonding between heptanal and BLG formed by Michael addition and Schiff-base reactions, and MG-like BLG exhibited fewer chemical shift residues. Molecular docking and molecular dynamics simulation confirmed the synergistic involvement of hydrophobic interactions and hydrogen bonds in shaping BLG-heptanal complexes thus promoting the stability of BLG structures. These findings indicated that the production of BLG-heptanal complexes was driven synergistically by non-covalent and covalent bonds, and their interaction processes were influenced by processes-induced formation of MG potentially tuning the release and retention behaviors of flavor compounds.
The chemical composition of the essential oil of Lindera subumbelliflora (Lauraceae) was investigated for the first time. The essential oil was obtained by hydrodistillation and fully characterised by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). The antifungal activity of L. subumbelliflora essential oil was tested against Candida albicans and Streptococcus mutans using the broth microdilution assay, whereas the microbial biofilms were determined using a semi-quantitative static biofilm. A total of 28 components (99.6%) were successfully identified, which were characterised by β-eudesmol (14.6%), cis-α-bergamotene (11.0%), α-copaene (8.5%), dodecen-1-ol (8.5%), and (E)-nerolidol (8.3%). The essential oil exhibited activity against Candida albicans and Streptococcus mutans with MIC values of 250 and 500 µg/mL, respectively. The essential oil increased the biofilm of Candida albicans by 38.25%, however, decreased the biofilm of Streptococcus mutans by 47.89% when treated with 500 µg/mL. Thus, the essential oil has a promising application in dentistry via inhibition of the growth of Candida albicans and Streptococcus mutans. However, the antibiofilm activity of the essential oil is only applicable for cariogenic Streptococcus mutans.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry*
Parabens (PBs) are esters of p-hydroxybenzoic acid, and there are growing concerns due to their potential to disrupt endocrine function and their wide use as preservatives in foodstuffs, including beverages. The consumption of bottled and hand-shaken teas is gradually replacing traditional tea consumption through brewing. However, no study has reported PB concentrations in different types of teas or packaging and their associated health risks. Our aim was to determine the concentration of PBs (methyl- (MetPB), ethyl- (EthPB), propyl- (PropPB), butyl-paraben (ButPB)) in green, black, and oolong teas in two varieties of products (bottled and hand-shaken teas), using UPLC-MS/MS. Additionally, we estimated the health risks associated with tea consumption in the general adult population of Taiwan. A Monte Carlo simulation was applied to estimate the distribution of daily PB intake through bottled (n = 79) and hand-shaken (n = 71) tea consumption. Our findings revealed geometric mean concentrations in bottled green/black/oolong teas were 714.1/631.2/532.1 ng/L for MetPB, 95.2/ 30.5/14.9 ng/L for EthPB, 77.9/28.3/non-detected (ND) ng/L for PropPB, and 69.3/26.6/ND ng/L for ButPB. Hand-shaken green/black/oolong teas exhibited concentrations of 867.5/2258/1307 ng/L for MetPB, 28.5/28.8/14.5 ng/L for EthPB, 25.4/18.3/17.8 ng/L for PropPB, and 30.3/18.0/15.5 ng/L for ButPB. The median MetPB concentrations in hand-shaken black (2333 ng/L) and oolong teas (1215 ng/L) were significantly higher than those in bottled black (595.4 ng/L) and oolong teas (489.3 ng/L). Conversely, median concentrations of EthPB, PropPB, and ButPB in bottled teas were significantly higher than those in hand-shaken teas. MetPB was the predominant PB, constituting 73.2-91.9% in bottled teas and 85-94% in hand-shaken teas. Our results showed no health risks associated with bottled or hand-shaken tea consumption based on reference doses. However, the study highlights the importance of continued vigilance given the potential chronic exposure to PBs from various sources, necessitating ongoing concern despite the absence of immediate risks from tea consumption.
The aim of this study was to pyrolyze individual (oil palm shell, empty fruit bunch and sawdust) as well as blend biomass in a thermogravimetric mass spectrometry (TG-MS) from room temperature to 800 °C at constant heating rate of 15 °C/min. The results showed that the onset TG temperature for blend biomass shifted slightly to lower values. Activation energy values were also found to decrease slightly after blending the biomass. Interestingly, the MS spectra of selected gases (H2O CH4, H2O, C2H2, C2H4 or CO, CH2O, CH3OH, HCl, C3H6, CO2, HCOOH, and C6H12) evolved from blend biomass showed decreased in the intensity as compared to their individual biomass. Overall, the blend biomass showed synergy which provides ways to expand the possibility of utilizing multiple feedstocks in one thermo-chemical system.
The fruit and leaf of God's crown (Phaleria macrocarpa) have been traditionally used to treat a wide variety of diseases. However, the proteins of this tropical plant are still heavily understudied. Three protein extraction methods; phenol (Phe), trichloroacetic acid (TCA)-acetone-phenol (TCA-A-Phe), and ultrasonic (Ult) were compared on the fruit and leaf of P. macrocarpa. The Phe extraction method showed the highest percentage of recovered protein after the resolubilization process for both leaf (12.24%) and fruit (30.41%) based on protein yields of the leaf (6.15 mg/g) and fruit (36.98 mg/g). Phe and TCA-A-Phe extraction methods gave well-resolved bands over a wide range of molecular weights through sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Following liquid chromatography-tandem mass spectrometry analysis, proteins identified through the Phe extraction method were 30%-35% enzymatic proteins, including oxidoreductases, transferases, hydrolases, lyases, isomerases, and ligases that possess various biological functions. PRACTICAL APPLICATIONS: Every part of God's crown plant is traditionally consumed to treat various illnesses. While plant's benefits are well known and have led to a plethora of health products, the proteome remains mostly unknown. This study compares three protein extraction methods for the leaf and fruit of P. macrocarpa and identifies their proteins thru LC-MS/MS coupled with PEAKS. These method comparisons can be a guide for works on other plants as well. In addition, the proteomics data from this study may shed light on the functional properties of these plant parts and their products.
The carnivorous plants of genus Nepenthes produce unique pitchers containing secretory glands, which secrete proteins into the digestive fluid. We investigated protein profile in the pitcher fluid during the first three days of opening to understand carnivory trait of Nepenthes × ventrata. The proteome analysis of pitcher fluid from N. × ventrata was performed by label-free quantitative liquid chromatography mass spectrometry (nLC-MS/MSALL). Raw MS data have been deposited to the ProteomeXchange with identifier PXD007251. This dataset allows the identification and quantification of proteins from pitcher fluids to elucidate proteins involved in carnivory physiology of Nepenthes species.
Kajian ini adalah mengenai peranan sterol sebagai penunjuk bio-lipid untuk mengenal pasti variasi dan sumber bahan organik yang hadir bersama sedimen permukaan dari Kuala Selangor, Selangor. Kajian melibatkan kaedah pengekstrakan sterol daripada sampel sedimen dan seterusnya dianalisis menggunakan GC-MS untuk menentukan kehadiran sebatian tersebut. Sepuluh sebatian sterol dikenal pasti hadir dengan fitosterol merupakan sebatian dominan di kawasan kajian iaitu 79% daripada jumlah keseluruhan sterol. Ini diikuti oleh kolesterol serta sterol kumbahan masing-masing menyumbangkan 6% daripada jumlah keseluruhan sterol manakala selebihnya berada dalam julat 1-5%. Indeks Sumber Sterol (SSI) juga menunjukkan kandungan fitosterol yang tinggi walaupun hadir pada kadar yang berbeza di setiap stesen pensampelan. Penilaian pencemaran kumbahan menggunakan nisbah koprostanol/kolesterol, koprostanol/(koprostanol+kholestanol) serta epikoprostanol/koprostanol menunjukkan kawasan kajian tidak mengalami pencemaran kumbahan walaupun sterol daripada sumber kumbahan hadir di persekitarannya. Kesimpulannya sedimen permukaan di Kuala Selangor, Selangor mengandungi campuran sterol daripada pelbagai sumber yang hadir di persekitarannya dengan didominasi oleh fitosterol yang berasal daripada tumbuhan terestrial.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Microcystins (MCs) are secondary metabolites produced by cyanobacteria and have been well-documented in temperate and tropical regions. However, knowledge of the production of MCs in extremely cold environments is still in its infancy. Recently, examination of 100-year-old Antarctic cyanobacterial mats collected from Ross Island and the McMurdo Ice Shelf during Captain R.F. Scott's Discovery Expedition revealed that the presence of MCs in Antarctica is not a new phenomenon. Here, morphological and molecular phylogenetic analyses are used to identify a new microcystin-producing freshwater cyanobacterium, Anagnostidinema pseudacutissimum. The strain was isolated from a deep-frozen (-15 °C) sample collected from a red-brown cyanobacterial mat in a frozen pond at Cape Crozier (Ross Island, continental Antarctica) in 1984-1985. Amplification of the mcyE gene fragment involved in microcystin biosynthesis from A. pseudacutissimum confirmed that it is identical to the sequence from other known microcystin-producing cyanobacteria. Analysis of extracts from this A. pseudacutissimum strain by HPLC-MS/MS confirmed the presence of MC-LR and -YR at concentrations of 0.60 μg/L and MC-RR at concentrations of 0.20 μg/L. This is the first report of microcystin production from a species of Anagnostidinema from Antarctica.
A benzoylthiourea molecule namely 1,1-dibenzyl-3-(2-bromobenzoyl)thiourea (2BrBT) was synthesized and characterized
by C, H, N and S elemental, mass spectrometry and spectroscopic analyses (infrared, ultraviolet-visible and nuclear
magnetic resonance). The 2BrBT compound crystallized in a tetragonal system with the space group P43 and exhibits
an acentric crystalline packing due to the presence of intermolecular H-bonding network that forms a self-assembly
of 1D helical motif. The asymmetric delocalisation of electrons in the molecule retains its transparency throughout the
visible and near-infrared region and hence, essentially propagates the macroscopic helical motif in the solid state. The
highest-occupied and lowest-unoccupied molecular orbital (HOMO/LUMO) are mainly found on the thiourea moiety and
the benzoylthiourea fragment, respectively and shows an optical bandgap of 3.50 eV. The influence of its geometrical
characteristics to the optical properties of 2BrBT is established and discussed in view of nonlinear optical (NLO)
application.
Poor performance of wet scrubbers in rubber processing plants due to breakthrough of specific volatile organic compounds (VOCs) causes odour impact events. The performance of wet scrubbers in the rubber drying process to remove VOCs was investigated in order to determine the responsible odorants. VOC emissions originating at the inlet and outlet of wet scrubbers were quantified using gas chromatography-mass spectrometry/olfactometry (GC-MS/O). Critical VOCs were identified alongside seasonal and daily variations of those VOCs. Altogether, 80 VOCs were detected in rubber emissions with 16 classified as critical VOCs based on their chemical concentration, high odour activity value (OAV) and unpleasant odour. Volatile fatty acids (VFAs) were the dominant VOCs with seasonal variations affecting emission composition. Results demonstrated the ineffectiveness of the wet scrubbers to mitigate odorous VOCs whereas the removal of some VOCs could be improved based on their polarity and solubility. It was found that there is a correlation between the wet scrubber performance and VFAs concentration in the emissions. The findings demonstrated that combining quantitative and sensory analyses improved accuracy in identifying odorous VOCs, which can cause odour annoyance from rubber processing. A VOC identification framework was proposed using both analyses approaches.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Durian shell (DS) was pyrolyzed in a drop-type fixed-bed reactor to study the physicochemical properties of the products. The experiment was carried out with different particle sizes (up to 5mm) and reaction temperatures (250-650°C). The highest bio-oil yield was obtained at 650°C (57.45wt%) with DS size of 1-2mm. The elemental composition and higher heating value of the feedstock, bio-oil (650°C), and bio-char (650°C) were determined and compared. The compositions of product gases were determined via gas chromatography with thermal conductivity detector. The chemical composition of bio-oil was analyzed by gas chromatography-mass spectrometry. The bio-oil produced at lower temperature yields more alcohols, whereas the bio-oil produced at higher temperature contains more aromatics and carbonyls. Bio-oil has potential to be used as liquid fuel or fine chemical precursor after further upgrading. The results further showed the potential of bio-char as a solid fuel.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
A first search for pair production of dark matter candidates through vector boson fusion in proton-proton collisions at sqrt[s]=8 TeV is performed with the CMS detector. The vector boson fusion topology enhances missing transverse momentum, providing a way to probe supersymmetry, even in the case of a compressed mass spectrum. The data sample corresponds to an integrated luminosity of 18.5 fb^{-1}, recorded by the CMS experiment. The observed dijet mass spectrum is consistent with the standard model expectation. In an effective field theory, dark matter masses are explored as a function of contact interaction strength. The most stringent limit on bottom squark production with mass below 315 GeV is also reported, assuming a 5 GeV mass difference with respect to the lightest neutralino.
Spatial segmentation is a critical procedure in mass spectrometry imaging (MSI)-based biochemical analysis. However, the commonly used unsupervised MSI segmentation methods may lead to inappropriate segmentation results as the MSI data is characterized by high dimensionality and low signal-to-noise ratio. This process can be improved by the incorporation of precise prior knowledge, which is hard to obtain in most cases. In this study, we show that the incorporation of partial or coarse prior knowledge from different sources such as reference images or biological knowledge may also help to improve MSI segmentation results. Here, we propose a novel interactive segmentation strategy for MSI data called iSegMSI, which incorporates prior information in the form of scribble-regularization of the unsupervised model to fine-tune the segmentation results. By using two typical MSI data sets (including a whole-body mouse fetus and human thyroid cancer), the present results demonstrate the effectiveness of the iSegMSI strategy in improving the MSI segmentations. Specifically, the method can be used to subdivide a region into several subregions specified by the user-defined scribbles or to merge several subregions into a single region. Additionally, these fine-tuned results are highly tolerant to the imprecision of the scribbles. Our results suggest that the proposed iSegMSI method may be an effective preprocessing strategy to facilitate the analysis of MSI data.
Tongkat ali commonly known as Malaysian Ginseng (Eurycoma longifolia) is a herbal root worldwide available in nutraceuticals, either as a crude powder or capsules blended with other herbal products. Herein, a multiplexed metabolomics approach based on nuclear magnetic resonance (NMR) and solid-phase microextraction combined with gas chromatography-mass spectrometry (SPME-GC-MS) was applied for authentic tongkat ali extract vs some commercial products quality control analysis. NMR metabolite fingerprinting identified 15 major metabolites mostly ascribed to sugars, organic and fatty acids in addition to quassinoids and cinnamates. Following that, multivariate analysis as the non-supervised principal component analysis (PCA) and supervised orthogonal partial least squares-discriminant analysis (OPLS-DA) were applied revealing that differences were related to fatty acids and 13,21-dihydroeurycomanone being more enriched in authentic root. SPME-GC-MS aroma profiling led to the identification of 59 volatiles belonging mainly to alcohols, aldehydes/furans and sesquiterpene hydrocarbons. Results revealed that aroma of commercial products showed relatively different profiles being rich in vanillin, maltol, and methyl octanoate. Whereas E-cinnamaldehyde, endo-borneol, terpinen-4-ol, and benzaldehyde were more associated to the authentic product. The present study shed the light for the potential of metabolomics in authentication and standardization of tongkat ali and identification of its true flavor composition.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry
Natural rubber or latex from the Hevea brasiliensis is an important commodity in various economic sectors in today's modern society. Proteins have been detected in latex since the early twentieth century, and they are known to regulate various biological pathways within the H. brasiliensis trees such as the natural rubber biosynthesis, defence against pathogens, wound healing, and stress tolerance. However, the exact mechanisms of the pathways are still not clear. Proteomic analyses on latex have found various proteins and revealed how they fit into the mechanisms of the biological pathways. In the past three decades, there has been rapid latex protein identification due to the improvement of latex protein extraction methods, as well as the emergence of two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS). In this manuscript, we reviewed the methods of latex protein extraction that keeps on improving over the past three decades as well as the results of numerous latex protein identification and quantitation.
Metabolomics and proteomics offer significant advantages in understanding biological mechanisms at two hierarchical levels. However, conventional single omics analysis faces challenges due to the high demand for specimens and the complexity of intrinsic associations. To obtain comprehensive and accurate system biological information, we developed a multiomics analytical method called Windows Scanning Multiomics (WSM). In this method, we performed simultaneous extraction of metabolites and proteins from the same sample, resulting in a 10% increase in the coverage of the identified biomolecules. Both metabolomics and proteomics analyses were conducted by using ultrahigh-performance liquid chromatography mass spectrometry (UPLC-MS), eliminating the need for instrument conversions. Additionally, we designed an R-based program (WSM.R) to integrate mathematical and biological correlations between metabolites and proteins into a correlation network. The network created from simultaneously extracted biomolecules was more focused and comprehensive compared to those from separate extractions. Notably, we excluded six pairs of false-positive relationships between metabolites and proteins in the network established using simultaneously extracted biomolecules. In conclusion, this study introduces a novel approach for multiomics analysis and data processing that greatly aids in bioinformation mining from multiomics results. This method is poised to play an indispensable role in systems biology research.
Rice is one of the crops cultivated in Malaysia, and it is the main diet for most of the population. In this study, dispersive liquid-liquid micro-extraction (DLLME) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were used to develop, optimise and validate a reliable, easy-to-use and quick approach to detect aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1) and aflatoxin G2 (AFG2). The extraction recoveries in DLLME were enhanced by the addition of 5% salt, utilising chloroform as the extraction solvent and acetonitrile as the dispersive solvent. The DLLME parameters - the extraction solvent volume, salt concentration and dispersive solvent volume were optimised with Box-Behnken design (BBD) and response surface methodology (RSM). Under optimised experimental conditions, excellent linearity was obtained with a limit of detection (LOD) ranging from 0.125 to 0.25 ng g-1, a limit of quantitation (LOQ) ranging from 0.25 to 0.3 ng g-1 and a correlation value (R2) of 0.990. The matrix effects were between -11.1% and 19.9%, and recoveries ranged from 87.4% to 117.3%. The optimised and validated method was used effectively to assess aflatoxins contamination in 20 commercial rice samples collected from local supermarkets in Penang, Malaysia. AFB1 was detected at 0.41-0.43 ng g-1 in two rice samples, below the regulatory limit.
As the first in-person Asia Oceania Human Proteomics Organization (AOHUPO) congress since 2018, the 11th AOHUPO congress was an opportune time for the research community to reconnect and to renew friendships after the long period of restricted travel due to the global pandemic. Moreover, this congress was a great opportunity for the many AO regional proteomics and mass spectrometry scientists to meet in Singapore to exchange ideas and to present their latest findings. Cohosted by the Singapore Society for Mass Spectrometry and the Malaysian Proteomics Society and held in conjunction with the seventh Asia Oceania Agricultural Proteomics Organization Congress and Singapore Society for Mass Spectrometry 2023, the meeting featured both human and agricultural proteomics. Over five hundred scientists from the AO region converged on the MAX Atria @ Singapore EXPO, Changi, Singapore from May 8 to 10 for the main congress. The diverse program was made up of 64 invited speakers and panellists for seven plenary lectures, 27 concurrent symposia, precongress and postcongress workshops, and 174 poster presentations. The AOHUPO society were able to celebrate not only their 20th anniversary but also the outstanding academic research from biological and agricultural proteomics and related 'omics fields being conducted across the Asia-Oceania region.
Nowadays, microplastic pollution is one of the globally urgent concerns as a result of discharging plastic products into the atmosphere, aquatic and soil environments. Microplastics have average size of less than 5 mm, are non-biodegradable, accumulative, and highly persistent substances. Thousands of tons of microplastics are still accumulated in various environments, posing an enormous threat to human health and living creatures. Here, we review the occurrence and analytical methods, and impact of microplastics in the environments including soil, aquatic media, and atmosphere. Analytical methods including visual observation, Fourier-transform infrared spectroscopy, Raman spectroscopy, scanning electron microscopy, and pyrolysis-gas chromatography-mass spectrometry were evaluated. We elucidated the environmental and human health impacts of microplastics with emphasis on life malfunction, immune disruption, neurotoxicity, diseases and other tangible health risks. This review also found some shortages of analytical equivalence and/or standardization, inconsistence in sampling collection and limited knowledge of microplastic toxicity. It is hopeful that the present work not only affords a more insight into the potential dangers of microplastics on human health but also urges future researches to establish new standardizations in analytical methods.
Matched MeSH terms: Gas Chromatography-Mass Spectrometry