Simulations of Test Reduction Using Pooled Heavy Metals Analysis in Cannabis.

BACKGROUND: Cannabis species have a propensity to bioaccumulate toxic heavy metals from their growth media. Increased testing for these metals is required to improve the safety of the legal medical and recreational cannabis industries. However, the current methods used for mandated heavy metals tests are not efficient for a large framework. As a result, there is limited testing capacity, high testing costs, and long wait times for results across North America. OBJECTIVE: This study aimed to demonstrate that pooling strategies can be used to increase the throughput in cannabis testing labs and reduce some of the strain on the industry. METHODS: This paper presents an algorithm to simulate different pooling strategies. The algorithm was applied to real world data sets collected from Washington and California state testing labs. RESULTS: Using a single pooling method, a pool size of three samples on average resulted in a 23.8% reduction in tests required for 100 samples for the Washington lab. For the California lab, pooling four samples on average resulted in a 54.1% reduction in tests required for 100 samples. CONCLUSION: The algorithms generated from the Washington and California lab data demonstrated that pooled testing strategies can be developed on a case-by-case method to reduce the time, effort, and costs associated with heavy metals tests. HIGHLIGHTS: The benefits of pooled testing will vary depending on the region and rate of contamination seen in each testing lab. Overall, our results demonstrate pooled testing has the potential to reduce the fiscal costs of testing through increased efficiency, allowing increased testing, leading to greater safety.

Metals in Cannabis Vaporizer Aerosols: Sources, Possible Mechanisms, and Exposure Profiles.

In recent years, cannabis vaporizer cartridges have increased in popularity and availability, and there are concerns regarding exposure to heavy-metal compounds from their use. The physical components of the cartridge devices themselves have been implicated as a potential source of metal exposure, but it is not known if these metals migrate into the inhalable vapor. This study analyzes the components of vaporizer cartridges for 10 different metals and also collects aerosol mixtures from 13 randomly purchased commercially available cannabis cartridges from Washington State to compare their elemental profiles. Results indicate that chromium, copper, nickel, as well as smaller amounts of lead, manganese, and tin migrate into the cannabis oil and inhaled vapor phase, resulting in a possible acute intake of an amount of inhaled metals above the regulatory standard of multiple governmental bodies. Noncartridge heating methods of cannabis flower and concentrate were compared, and results indicate that the heating device itself is a source of metal contamination. As safety and compliance testing regulations evolve, it will be important to include more than the standard As, Cd, Hg, and Pb to the list of regulated metals.

Strategies for Nonpolar Aerosol Collection and Heavy Metals Analysis of Inhaled Cannabis Products.

The rapid growth of inhalable cannabis concentrates raises questions about the safety of acute and chronic exposure to these aerosol mixtures. Due to the nonpolar nature of the aerosol mixture created from cannabis vapor cartridges, traditional aqueous-based capture methods used in e-cigarette or tobacco cigarette studies for analysis of metals are insufficient. Moreover, hydrophobic cannabis concentrates are not miscible with dilute aqueous acids and therefore not ideal for metal spiking unlike electronic nicotine delivery systems. This study describes a method of spiking nonaqueous matrices with aqueous metals standards to investigate aerosolization and recovery of the metals. It also compares various methods for nonpolar aerosol capture and subsequent analysis of 10 metals (As, Cd, Co, Cr, Cu, Hg, Mn, Ni, Pb, and Sn) in two model cannabis matrices, flower and concentrate. Spiked cannabis concentrates were vaped in commercially available cartridges, and their aerosol mixtures were investigated for recovery of heavy metals via ICP-MS. Spiked flower samples were also combusted to compare collection rates of the 10 metals. Results show that not all metals that are present in the concentrate or flower can be fully recovered in the aerosol capture processes at standard voltage settings or combustion temperatures. These studies also demonstrate the importance of a nonpolar solvent as part of the aerosol collection to increase the recovery of some metals. The high concentration of some metals seen in the concentrate suggests that the devices themselves are potential routes of exposure. The ICP-MS analysis method was further validated by evaluating different parameters including linearity, matrix effect, limit of detection, limit of quantitation, and repeatability.

Upstream adverse effects in risk assessment: a model of polychlorinated biphenyls, thyroid hormone disruption and neurological outcomes in humans.

BACKGROUND: Increasing data on early biological changes from chemical exposures requires new interpretation tools to support decision-making. OBJECTIVES: To test the possibility of applying a quantitative approach using human data linking chemical exposures and upstream biological perturbations to overt downstream outcomes. METHODS: Using polychlorinated biphenyl (PCB) exposures and maternal thyroid hormone (TH) perturbations as a case study, we model three relationships: (1) prenatal PCB exposures and TH changes, using free T(4) (FT(4)); (2) prenatal TH and childhood neurodevelopmental outcomes; and (3) prenatal PCB exposures and childhood neurodevelopmental outcomes (IQ). We surveyed the epidemiological literature; extracted relevant quantitative data; and developed models for each relationship, applying meta-analysis where appropriate. RESULTS: For relationship 1, a meta-analysis of 3 studies gives a coefficient of -0.27 pg/mL FT(4) per ln(sum of PCBs) (95% confidence interval [CI] -0.82 to 0.27). For relationship 2, regression coefficients from three studies of maternal FT(4) levels and cognitive scores ranged between 0.99 IQ points/(pg/mL FT(4)) (95% CI -0.31 to 2.2) and 7.6 points/(pg/mL FT(4)) (95% CI 1.2 to 16.3). For relationship 3, a meta-analysis of five studies produces a coefficient of -1.98 IQ points (95% CI -4.46 to 0.50) per unit increase in ln(sum of PCBs). Combining relationships 1 and 2 yields an estimate of -2.0 to -0.27 points of IQ per unit increase in ln(sum of PCBs). CONCLUSIONS: Combining analysis of chemical exposures and early biological perturbations (PCBs and FT(4)) with analysis of early biological perturbations and downstream overt effects (FT(4) and IQ) yields estimates within the range of studies of exposures and overt effects (PCBs and IQ). This is an example approach using upstream biological perturbations for effect prediction.

Adverse effects in risk assessment: modeling polychlorinated biphenyls and thyroid hormone disruption outcomes in animals and humans.

There is a growing need for quantitative approaches to extrapolate relationships between chemical exposures and early biological perturbations from animals to humans given increasing use of biological assays to evaluate toxicity pathways. We have developed such an approach using polychlorinated biphenyls (PCBs) and thyroid hormone (TH) disruption as a case study. We reviewed and identified experimental animal literature from which we developed a low-dose, linear model of PCB body burdens and decrements in free thyroxine (FT(4)) and total thyroxine (TT(4)), accounting for 33 PCB congeners; extrapolated the dose-response from animals to humans; and compared the animal dose-response to the dose-response of PCB body burdens and TH changes from eleven human epidemiological studies. We estimated a range of potencies for PCB congeners (over 4 orders of magnitude), with the strongest for PCB 126. Our approach to developing toxic equivalency models produced relative potencies similar to the toxicity equivalency factors (TEFs) from the World Health Organization (WHO). We generally found that the dose-response extrapolated from the animal studies tends to under-predict the dose-response estimated from human epidemiological studies. A quantitative approach to evaluating the relationship between chemical exposures and TH perturbations, based on animal data can be used to assess human health consequences of thyroid toxicity and inform decision-making.

Are oral contraceptives a significant contributor to the estrogenicity of drinking water?

Recent observed feminization of aquatic animals has raised concerns about estrogenic compounds in water supplies and the potential for these chemicals to reach drinking water. Public perception frequently attributes this feminization to oral contraceptives (OCs) in wastewater and raises concerns that exposure to OCs in drinking water may contribute to the recent rise in human reproductive problems. This paper reviews the literature regarding various sources of estrogens, in surface, source and drinking water, with an emphasis on the active molecule that comes from OCs. It includes discussion of the various agricultural, industrial, and municipal sources and outlines the contributions of estrogenic chemicals to the estrogenicity of waterways and estimates that the risk of exposure to synthetic estrogens in drinking water on human health is negligible. This paper also provides recommendations for strategies to better understand all the potential sources of estrogenic compounds in the environment and possibilities to reduce the levels of estrogenic chemicals in the water supply.

Discrete arrays of liquid-crystal-supported proteolipid monolayers as phantom cell surfaces.

The phospholipid bilayers of living cell membranes exist almost universally in a liquid state. This enables motion and spatial reorganization of membrane components on multiple length scales, which is an essential feature of many biological processes. There is great interest in the development of molecularly defined interfaces between synthetic materials and living cells. To this end, there is a need for solid substrate materials that can be derivatized with fluid, membrane-like interfaces. Herein, we describe array fabrication of discrete liquid-crystal areas supporting phospholipid monolayer membranes, and characterize the interactions with several different membrane surface proteins [avidin series, cholera toxin, green fluorescent protein (GFP), intercellular adhesion molecule (ICAM) and major histocompatibility complex (MHC)]. Three different linkage strategies (biotin, nickel chelating lipids complexing with histidine, and the choleratoxin binding unit (CTB) associating with G(M1) are evaluated. Additionally, experiments with live immunological T cells forming active synapses at the interface exhibit the specific nature of the surface.

Synthetic analogues of glycosylphosphatidylinositol-anchored proteins and their behavior in supported lipid bilayers.

Positioned at the C-terminus of many eukaryotic proteins, the glycosylphosphatidylinositol (GPI) anchor is a posttranslational modification that anchors the modified proteins in the outer leaflet of the plasma membrane. GPI-anchored proteins play vital roles in signal transduction, the vertebrate immune response, and the pathobiology of trypanosomal parasites. While many GPI-anchored proteins have been characterized, the biological functions of the GPI anchor have yet to be elucidated at a molecular level. We synthesized a series of GPI-protein analogues bearing modified anchor structures that were designed to dissect the contribution of various glycan components to the GPI-protein's membrane behavior. These anchor analogues were similar in length to native GPI anchors and included mimics of the native structure's three domains. A combination of expressed protein ligation and native chemical ligation was used to attach these analogues to the green fluorescent protein (GFP). These modified GFPs were incorporated in supported lipid bilayers, and their mobilities were analyzed using fluorescence correlation spectroscopy. The data from these experiments suggest that the GPI anchor is more than a simple membrane-anchoring device; it also may prevent transient interactions between the attached protein and the underlying lipid bilayer, thereby permitting rapid diffusion in the bilayer. The ability to generate chemically defined analogues of GPI-anchored proteins is an important step toward elucidating the molecular functions of this interesting post-translational modification.

Colorimetric bio-barcode amplification assay for cytokines.

The bio-barcode amplification assay has become a powerful tool in detecting tens to hundreds of biological targets such as proteins and nucleic acids in the entire sample. However, current bio-barcode detection schemes still require many experimental steps including microarrayer-based immobilization of oligonucleotides on a glass chip, silver enhancement of immobilized gold nanoparticles on a chip, and light-scattering measurement. Here, we report a colorimetric bio-barcode method that minimizes the above requirements while detecting 30 aM concentrations of cytokines (approximately 3 orders of magnitude more sensitive than conventional nonenzymatic cytokine detection assays). The assay is based on porous microparticles, which enable loading of a large number of barcode DNA per particle, and gold nanoparticle-based colorimetric barcode detection method.