A review of methods for the chemical characterization of cannabis natural products.

Cannabis has garnered a great deal of new attention in the past couple of years in the United States due to the increasing instances of its legalization for recreational use and indications for medicinal benefit. Despite a growing number of laboratories focused on cannabis analysis, the separation science literature pertaining to the determination of cannabis natural products is still in its infancy despite the plant having been utilized by humans for nearly 30 000 years and it being now the most widely used drug worldwide. This is largely attributable to the restrictions associated with cannabis as it is characterized as a schedule 1 drug in the United States. Presented here are reviewed analytical methods for the determination of cannabinoids (primarily) and terpenes (secondarily), the primary natural products of interest in cannabis plants. Focus is placed foremost on analyses from plant extracts and the various instrumentation and techniques that are used, but some coverage is also given to analysis of cannabinoid metabolites found in biological fluids. The goal of this work is to provide a collection of relevant separation science information, upon which the field of cannabis analysis can continue to grow.

Determination of cannabinoids from a surrogate hops matrix using multiple reaction monitoring gas chromatography with triple quadrupole mass spectrometry.

Cannabinoids are the primary bioactive constituents of Cannabis sativa and Cannabis indica plants. In this work, gas chromatography in conjunction with triple quadrupole mass spectrometry in multiple reaction monitoring mode was explored for determination of cannabinoids from a surrogate hops matrix. Gas chromatography with mass spectrometry is a reasonable choice for the analysis of these compounds; however, such methods are susceptible to false positives for Δ9-tetrahydrocannabinol, due to decarboxylation of Δ9-tetrahydrocannabinolic acid, its acid precursor, in the hot injection port. To avoid this transformation, the carboxyl group of Δ9-tetrahydrocannabinolic acid was protected through a silylation reaction. Multiple reaction monitoring transitions for both unmodified and silylated cannabinoids were developed and the fragmentation pathways for the different species were assigned. Precision and accuracy were evaluated for cannabinoids spiked into hops at different levels. The developed methods provided good linearity (R2  > 0.99) for all the cannabinoids with a linear range from 0.15 to 20 mg/L, and with limits of detection in the orders of low- to mid-picogram on column. The recoveries for the cannabinoids were generally between 75 and 120%. Precisions (<6% coefficient of variation) were within acceptable ranges.

Applications of MALDI-TOF MS in environmental microbiology.

Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is an emerging technique for microbial identification, characterization, and typing. The single colony method can be used for obtaining a protein fingerprint or profile unique to each microorganism. This technique has been mainly used in the clinical field, but it also has significant potential in the environmental field. The applications of MALDI-TOF MS in environmental microbiology are discussed in this review. An overview on the use of MALDI-TOF MS for environmental proteomics and metabolomics is given as well as its use for bacterial strain typing and bioremediation research. A more detailed review on the use of this technique for the identification, differentiation, and categorization of environmental microorganisms is given. Some of the parameters that can influence the results and reproducibility of MALDI-TOF MS are also discussed.

Expression and in vitro functional analyses of recombinant Gam1 protein.

Gam1, an early gene product of an avian adenovirus, is essential for viral replication. Gam1 is the first viral protein found to globally inhibit cellular SUMOylation, a critical posttranslational modification that alters the function and cellular localization of proteins. The interaction details at the interface between Gam1 and its cellular targets remain unclear due to the lack of structural information. Although Gam1 has been previously characterized, the purity of the protein was not suitable for structural investigations. In the present study, the gene of Gam1 was cloned and expressed in various bacterial expression systems to obtain pure and soluble recombinant Gam1 protein for in vitro functional and structural studies. While Gam1 was insoluble in most expression systems tested, it became soluble when it was expressed as a fusion protein with trigger factor (TF), a ribosome associated bacterial chaperone, under the control of a cold shock promoter. Careful optimization indicates that both low temperature induction and the chaperone function of TF play critical roles in increasing Gam1 solubility. Soluble Gam1 was purified to homogeneity through sequential chromatography techniques. Monomeric Gam1 was obtained via size exclusion chromatography and analyzed by dynamic light scattering. The SUMOylation inhibitory function of the purified Gam1 was confirmed in an in vitro assay. These results have built the foundation for further structural investigations that will broaden our understanding of Gam1's roles in viral replication.

A Comprehensive Analysis of Groundwater Quality in The Barnett Shale Region.

The exploration of unconventional shale energy reserves and the extensive use of hydraulic fracturing during well stimulation have raised concerns about the potential effects of unconventional oil and gas extraction (UOG) on the environment. Most accounts of groundwater contamination have focused primarily on the compositional analysis of dissolved gases to address whether UOG activities have had deleterious effects on overlying aquifers. Here, we present an analysis of 550 groundwater samples collected from private and public supply water wells drawing from aquifers overlying the Barnett shale formation of Texas. We detected multiple volatile organic carbon compounds throughout the region, including various alcohols, the BTEX family of compounds, and several chlorinated compounds. These data do not necessarily identify UOG activities as the source of contamination; however, they do provide a strong impetus for further monitoring and analysis of groundwater quality in this region as many of the compounds we detected are known to be associated with UOG techniques.

An evaluation of water quality in private drinking water wells near natural gas extraction sites in the Barnett Shale formation.

Natural gas has become a leading source of alternative energy with the advent of techniques to economically extract gas reserves from deep shale formations. Here, we present an assessment of private well water quality in aquifers overlying the Barnett Shale formation of North Texas. We evaluated samples from 100 private drinking water wells using analytical chemistry techniques. Analyses revealed that arsenic, selenium, strontium and total dissolved solids (TDS) exceeded the Environmental Protection Agency's Drinking Water Maximum Contaminant Limit (MCL) in some samples from private water wells located within 3 km of active natural gas wells. Lower levels of arsenic, selenium, strontium, and barium were detected at reference sites outside the Barnett Shale region as well as sites within the Barnett Shale region located more than 3 km from active natural gas wells. Methanol and ethanol were also detected in 29% of samples. Samples exceeding MCL levels were randomly distributed within areas of active natural gas extraction, and the spatial patterns in our data suggest that elevated constituent levels could be due to a variety of factors including mobilization of natural constituents, hydrogeochemical changes from lowering of the water table, or industrial accidents such as faulty gas well casings.

Chaperonin-mediated folding of viral proteins.

Chaperonins are universally conserved molecular machines that facilitate the proper -folding of nascent and partially folded polypeptides into their respective three-dimensional structures. These multimeric protein complexes utilize the energy derived from ATP hydrolysis to fuel a protein-folding mechanism that consists of multiple rounds of substrate binding, encapsulation, and eventual expulsion back into the cytosolic environment. In this portion of the chapter, the structure and function of group I and group II chaperonins are discussed. Furthermore, the general mechanism of chaperonin-mediated protein folding is addressed in addition to illustrating how viral phages such as Lambda, T4, and RB49 exploit the host machinery for the proper folding of viral gene products. Lastly, the phiEL chaperonin from phage EL is revealed to be the first virally encoded chaperonin and is proposed to function independently of the host chaperonin machinery. The molecular architecture of the phiEL chaperonin, coupled with its unique functional abilities, renders its characterization a challenge and further highlights its novelty as a potentially whole new class of chaperonins.

Insights Into the Role of Heat Shock Protein 27 in the Development of Neurodegeneration.

Small heat shock protein 27 is a critically important chaperone, that plays a key role in several essential and varied physiological processes. These include thermotolerance, apoptosis, cytoskeletal dynamics, cell differentiation, protein folding, among others. Despite its relatively small size and intrinsically disordered termini, it forms large and polydisperse oligomers that are in equilibrium with dimers. This equilibrium is driven by transient interactions between the N-terminal region, the α-crystallin domain, and the C-terminal region. The continuous redistribution of binding partners results in a conformationally dynamic protein that allows it to adapt to different functions where substrate capture is required. However, the intrinsic disorder of the amino and carboxy terminal regions and subsequent conformational variability has made structural investigations challenging. Because heat shock protein 27 is critical for so many key cellular functions, it is not surprising that it also has been linked to human disease. Charcot-Marie-Tooth and distal hereditary motor neuropathy are examples of neurodegenerative disorders that arise from single point mutations in heat shock protein 27. The development of possible treatments, however, depends on our understanding of its normal function at the molecular level so we might be able to understand how mutations manifest as disease. This review will summarize recent reports describing investigations into the structurally elusive regions of Hsp27. Recent insights begin to provide the required context to explain the relationship between a mutation and the resulting loss or gain of function that leads to Charcot-Marie Tooth disease and distal hereditary motor neuropathy.

Reservoir optimized plunger lift technology reduces hydrocarbon emissions from aging gas wells.

Gas well liquification is a problematic process whereby liquids collect in the wellbore and near wellbore reservoir resulting in production impedance in aging gas wells. Removal of these liquids is traditionally performed through human operated blowdown events; however, this practice results in the release of hydrocarbon emissions into the atmosphere. The removal process, called 'deliquification', can also be accomplished through the utilization of various plunger lift technologies. These allow the extraction of retained fluids from the wellbore and near-wellbore reservoir; however, these technologies vary greatly with respect to automation, intelligence, and efficacy. Here we examined the rates of production loss and the frequency of emission events in mature natural gas wells equipped with various automated plunger lift technologies. Overall, 'intelligent' plunger lift systems that base their optimization on reservoir and wellbore conditions, as opposed to standardized or scheduled operations, performed the best exhibiting a 0.13% loss of production gas to atmospheric emissions compared to a 1.37% loss of production observed from wells without a plunger lift system. Additionally, wells equipped with a next generation reservoir optimized plunger lift demonstrated a reduced rate of production decline compared to those wells without a plunger lift technology (-0.066%/day and -0.242%/day, respectively). These data have widespread implications for the operational and environmental management of a consistently increasing count of aging natural gas production wells.

Characterizing anecdotal claims of groundwater contamination in shale energy basins.

The increased societal monitoring of unconventional oil and gas development (UD) has brought forth tremendous scrutiny over the environmental stewardship and subsequent public health impacts of surface and sub-surface activities. Concerns over groundwater quality in shale energy basins have prompted concerned citizens into monitoring UD activities for a series of qualitative parameters, and even coordinating sampling efforts for chemical analysis. Here we present a list of analytical parameters, hierarchically structured to guide concerned citizens through an efficient and cost-effective monitoring program. Utilizing this multi-step testing regime, we assessed groundwater quality from 36 private water wells involved in 19 anecdotal claims of alleged UD-related contamination across the Barnett, Eagle Ford, Haynesville, and Marcellus Shale formations in the United States. Our analytical findings aligned with the landowners' accounts of their situation in only 5 of the 36 collected samples, with several cases revealing environmental abnormalities that were unbeknownst to the landowners but likely unrelated to UD activities. These data are some of the first to assess the relationship between landowner perception and analytical determination in a cohort of highly variable anecdotal cases of alleged groundwater contamination, revealing a notable disconnect that is likely attributable to a myriad of societal and environmental factors. The analytical modalities presented here also serve as a step-wise method in a weight of evidence approach to assess the presence or absence of anthropogenic contamination under the most variable hydrogeological conditions.

A review of the analytical methods used for beer ingredient and finished product analysis and quality control.

Beer is an incredibly complex beverage containing more than 3000 different compounds, including carbohydrates, proteins, ions, microbes, organic acids, and polyphenols, among others. Beer becomes even more complex during storage, for over time it may undergo chemical changes that negatively affect the flavor, aroma, and appearance. Thus, it can be expected that maintaining the quality of beer throughout its lifetime is a difficult task. Since it is such a popular drink throughout the world, being familiar with proper analytical techniques for beer evaluation is useful for researchers and brewers. These techniques include, but are not limited to, gas chromatography, liquid chromatography, matrix assisted laser desorption/ionization, capillary electrophoresis, mass spectrometry, ultraviolet-visible spectroscopy, and flame ionization detection. This review aims to summarize the various ingredients and components of beer, discuss how they affect the finished product, and present some of the analytical methods used for quality control and understanding the formation of chemicals in beer during the brewing process.

Analysis of bacteria stress responses to contaminants derived from shale energy extraction.

In order to survive environmental changes, bacteria have stress responses, which protect them from adverse and variable conditions. Contamination can be a source of stress and bacterial responses can serve as an indicator of environmental abnormality. In this work, the biochemical effects of toxic compounds that stem from hydraulic fracturing were measured on the whole cell-derived fatty acid and protein compositions of Escherichia coli, Klebsiella oxytoca, Pseudomonas aeruginosa, Pseudomonas putida, Pseudomonas stutzeri, Aeromonas hydrophila, Bacillus cereus, and Bacillus subtilis. These microorganisms were exposed to elevated levels of benzene, ethanol, propanol, toluene, and salt. These were chosen to represent significant subsurface contamination or a surface spill. The fatty acid and protein profiles for the bacteria were analyzed using gas chromatography - vacuum ultraviolet spectroscopy and matrix-assisted laser desorption ionization time-of-flight mass spectrometry, respectively. Overall, different fatty acid and protein profiles were observed when the microorganisms were grown in the presence of the toxic compounds. The cells exhibited an increase in the saturated/unsaturated ratio and displayed the presence of branched and cyclopropane fatty acids when in the presence of common fracture fluid constituents to decrease membrane permeability which was confirmed by the analysis of produced water. This approach provides a potentially useful tool for environmental diagnosis, since proteins and fatty acids can act as a harbinger of ecological health.

Treatment modalities for the reuse of produced waste from oil and gas development.

Unconventional oil and gas development is achieved through a series of sub-processes, which utilize large amounts of water, proppant, and chemical additives to retrieve sequestered hydrocarbons from low permeability petroliferous strata. As a result, a large amount of wastewater is produced, which is traditionally disposed of via subsurface injection into non-productive stratum throughout the country. However, this method of waste management has been linked to the induction of seismic events in a number of regions across North America, calling into question the environmental stewardship and sustainability of subsurface waste disposal. Advancements in water treatment technologies have improved the efficacy and financial viability of produced water recycling for beneficial reuse in the oil and gas sector. This review will cover the various treatment options that are currently being utilized in shale energy basins to remove organic, inorganic, and biological constituents, as well as some emerging technologies that are designed to remove pertinent contaminants that would otherwise preclude the reuse of produced water for production well stimulation.

Matrix-effect-free determination of BTEX in variable soil compositions using room temperature ionic liquid co-solvents in static headspace gas chromatography mass spectrometry.

Concerns about the environmental impact of shale energy exploration (i.e., hydraulic fracturing and other well stimulation techniques) have risen due to its rapid expansion in the United States and other countries. Soils in shale basins engaged in unconventional oil and gas development can become contaminated by volatile organic compounds (VOCs), such as benzene, toluene, ethylbenzene, and xylenes (BTEX) through mishandling of chemical additives, products, and/or waste fluids. In this study, room temperature ionic liquids (RTILs) were evaluated as solvents in an effort to increase sensitivity and to reduce and normalize matrix effects associated with varying soil compositions during analysis. Headspace gas chromatography mass spectrometry (HS-GC-MS) experiments demonstrated that hydrophilic RTILs 1-ethyl-3-methylimidazolium ethyl sulfate [EMIM][ESO4], 1-ethyl-3-methylimidazolium diethyl phosphate [EMIM][DEP], and tris(2-hydroxyethyl) methylammonium methylsulfate [MTEOA][MeOSO3] normalized the response for BTEX compounds between 2 different soils, sandy loam and sandy clay loam. Furthermore, the optimization of the HS equilibration time to 30 min resulted in the reduction of matrix effects in certified reference soils of sand, clay, and loam textures. Limits of detection and limits of quantification were in the sub- to mid- pg g-1 level in soil. For determination at 1 g of certified BTEX reference soil, the relative standard deviation was within 10% and percent recoveries were above 80% for toluene, ethylbenzene, and xylenes. This method reduces the need to characterize and matrix-match soil texture for calibration purposes. It also reduces the analysis time and increases precision and accuracy for the quantification of BTEX in variable soil matrices relative to standard methods.

Characterization of bacterial diversity in contaminated groundwater using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Groundwater is a major source for drinking water in the United States, and therefore, its quality and quantity is of extreme importance. One major concern that has emerged is the possible contamination of groundwater due to the unconventional oil and gas extraction activities. As such, the impacts of exogenous contaminants on microbial ecology is an area to be explored to understand what are the chemical and physical conditions that allow the proliferation of pathogenic bacteria and to find alternatives for water treatment by identifying organic-degrading bacteria. In this work, we assess the interplay between groundwater quality and the microbiome in contaminated groundwaters rich in hydrocarbon gases, volatile organic and inorganic compounds, and various metals. Opportunistic pathogenic bacteria, such as Aeromonas hydrophila, Bacillus cereus, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia, were identified, increasing the risk for consumption of and exposure to these contaminated groundwaters. Additionally, antimicrobial tests revealed that many of the identified bacteria were resistant to different antibiotics. The MALDI-TOF MS results were successfully confirmed with 16S rRNA gene sequencing, proving the accuracy of this high-throughput method. Collectively, these data provide a seminal understanding of the microbial populations in contaminated groundwater overlying anthropogenic activities like unconventional oil and gas development.

Exploring the links between groundwater quality and bacterial communities near oil and gas extraction activities.

Bacterial communities in groundwater are very important as they maintain a balanced biogeochemical environment. When subjected to stressful environments, for example, due to anthropogenic contamination, bacterial communities and their dynamics change. Studying the responses of the groundwater microbiome in the face of environmental changes can add to our growing knowledge of microbial ecology, which can be utilized for the development of novel bioremediation strategies. High-throughput and simpler techniques that allow the real-time study of different microbiomes and their dynamics are necessary, especially when examining larger data sets. Matrix-assisted laser desorption-ionization (MALDI) time-of-flight mass spectrometry (TOF-MS) is a workhorse for the high-throughput identification of bacteria. In this work, groundwater samples were collected from a rural area in southern Texas, where agricultural activities and unconventional oil and gas development are the most prevalent anthropogenic activities. Bacterial communities were assessed using MALDI-TOF MS, with bacterial diversity and abundance being analyzed with the contexts of numerous organic and inorganic groundwater constituents. Mainly denitrifying and heterotrophic bacteria from the Phylum Proteobacteria were isolated. These microorganisms are able to either transform nitrate into gaseous forms of nitrogen or degrade organic compounds such as hydrocarbons. Overall, the bacterial communities varied significantly with respect to the compositional differences that were observed from the collected groundwater samples. Collectively, these data provide a baseline measurement of bacterial diversity in groundwater located near anthropogenic surface and subsurface activities.

Single-Ring Intermediates Are Essential for Some Chaperonins.

Chaperonins are macromolecular complexes found throughout all kingdoms of life that assist unfolded proteins reach a biologically active state. Historically, chaperonins have been classified into two groups based on sequence, subunit structure, and the requirement for a co-chaperonin. Here, we present a brief review of chaperonins that can form double- and single-ring conformational intermediates in their protein-folding catalytic pathway. To date, the bacteriophage encoded chaperonins ϕ-EL and OBP, human mitochondrial chaperonin and most recently, the bacterial groEL/ES systems, have been reported to form single-ring intermediates as part of their normal protein-folding activity. These double-ring chaperonins separate into single-ring intermediates that have the ability to independently fold a protein. We discuss the structural and functional features along with the biological relevance of single-ring intermediates in cellular protein folding. Of special interest are the ϕ-EL and OBP chaperonins which demonstrate features of both group I and II chaperonins in addition to their ability to function via single-ring intermediates.