Folic acid deficiency increases sensitivity to DNA damage by glucose and methylglyoxal.

Type 2 diabetes (T2D) is associated with elevated frequencies of micronuclei (MNi) and other DNA damage biomarkers. Interestingly, individuals with T2D are more likely to be deficient in micronutrients (folic acid, pyridoxal-phosphate, cobalamin) that play key roles in one-carbon metabolism and maintaining genomic integrity. Furthermore, it has recently been shown that deficiencies in these nutrients, in particular folic acid leaves cells susceptible to glucose-induced DNA damage. Therefore, we sought to investigate if the B lymphoblastoid WIL2-NS cell line cultured under folic acid-deficient conditions was more sensitive to DNA damage induced by glucose, or the reactive glycolytic byproduct methylglyoxal (MGO) and subsequent advanced glycation endproduct formation. Here, we show that only WIL2-NS cultured under folic acid-deficient conditions (23 nmol/l) experience an increase in MNi frequency when exposed to high concentrations of glucose (45 mmol/l) or MGO (100 µmol/l). Furthermore, we showed aminoguanidine, a well-validated MGO and free radical scavenger was able to prevent further MNi formation in folic acid-deficient cells exposed to high glucose, which may be due to a reduction in MGO-induced oxidative stress. Interestingly, we also observed an increase in MGO and other dicarbonyl stress biomarkers in folic acid-deficient cells, irrespective of glucose concentrations. Overall, our evidence shows that folic acid-deficient WIL2-NS cells are more susceptible to glucose and/or MGO-induced MNi formation. These results suggest that individuals with T2D experiencing hyperglycemia and folic acid deficiency may be at higher risk of chromosomal instability.

The role of a Mediterranean diet and physical activity in decreasing age-related inflammation through modulation of the gut microbiota composition.

Chronic inflammation is known to be a predominant factor in the development of many age-related conditions including CVD, type II diabetes and neurodegenerative diseases. Previous studies have demonstrated that during the ageing process there is an increase in inflammatory biomarkers, which may be partially brought about by detrimental changes in the gut microbiota. The Mediterranean diet (MedDiet) and physical activity (PA) are protective against inflammation and chronic disease, and emerging evidence has shown that these effects may be partially mediated through favourable changes in the gut microbiota. In this review, we have evaluated the published literature on the effect of a MedDiet and PA on the gut microbiota. We also discuss the relationship between the gut microbiota and inflammation with a focus on healthy ageing. While inconsistent study designs make forming definitive conclusions challenging, the current evidence suggests that both a MedDiet and PA are capable of modifying the gut microbiota in a way that is beneficial to host health. For example, the increases in the relative abundance of SCFA producing bacteria that are considered to possess anti-inflammatory properties. Modification of the gut microbiota through a MedDiet and PA presents as a potential method to attenuate age-related increases in inflammation, and additional studies utilising older individuals are needed to fill the knowledge gaps existing in current literature.

Methylglyoxal Impairs Sister Chromatid Separation in Lymphocytes.

The accurate segregation of sister chromatids is complex, and errors that arise throughout this process can drive chromosomal instability and tumorigenesis. We recently showed that methylglyoxal (MGO), a glycolytic by-product, can cause chromosome missegregation events in lymphocytes. However, the underlying mechanisms of this were not explored. Therefore, in this study, we utilised shotgun proteomics to identify MGO-modified proteins, and label-free quantitation to measure changes in protein abundance following exposure to MGO. We identified numerous mitotic proteins that were modified by MGO, including those involved in the separation and cohesion of sister chromatids. Furthermore, the protein abundance of Securin, an inhibitor of sister chromatid separation, was increased following treatment with MGO. Cytological examination of chromosome spreads showed MGO prevented sister chromatid separation, which was associated with the formation of complex nuclear anomalies. Therefore, results from this study suggest MGO may drive chromosomal instability by preventing sister chromatid separation.

Proteomic Analysis of Methylglyoxal Modifications Reveals Susceptibility of Glycolytic Enzymes to Dicarbonyl Stress.

Methylglyoxal (MGO) is a highly reactive cellular metabolite that glycates lysine and arginine residues to form post-translational modifications known as advanced glycation end products. Because of their low abundance and low stoichiometry, few studies have reported their occurrence and site-specific locations in proteins. Proteomic analysis of WIL2-NS B lymphoblastoid cells in the absence and presence of exogenous MGO was conducted to investigate the extent of MGO modifications. We found over 500 MGO modified proteins, revealing an over-representation of these modifications on many glycolytic enzymes, as well as ribosomal and spliceosome proteins. Moreover, MGO modifications were observed on the active site residues of glycolytic enzymes that could alter their activity. We similarly observed modification of glycolytic enzymes across several epithelial cell lines and peripheral blood lymphocytes, with modification of fructose bisphosphate aldolase being observed in all samples. These results indicate that glycolytic proteins could be particularly prone to the formation of MGO adducts.

Genotoxicity of advanced glycation end products in vitro is influenced by their preparation temperature, purification and cell exposure time.

Advanced glycation end products (AGEs) are formed via non-enzymatic reactions between amino groups of proteins and the carbonyl groups of reducing sugars. Previous studies have shown that highly glycated albumin prepared using a glucose-bovine serum albumin (Glu-BSA) model system incubated at 60°C for 6 weeks induces genotoxicity in WIL2-NS cells at 9 days of exposure measured by the cytokinesis-block micronucleus cytome (CBMNcyt) assay. However, this AGE model system is not physiologically relevant as normal body temperature is 37°C and the degree of glycation may exceed the extent of albumin modification in vivo. We hypothesised that the incubation temperature and purification method used in these studies may cause changes to the chemical profile of the glycated albumin and may influence the extent of genotoxicity observed at 3, 6 and 9 days of exposure. We prepared AGEs generated using Glu-BSA model systems incubated at 60°C or 37°C purified using trichloroacetic acid (TCA) precipitation or ultrafiltration (UF) and compared their chemical profile (glycation, oxidation, and aggregation) and genotoxicity in WIL2-NS cells using the CBMNcyt assay after 3, 6 and 9 days of exposure. The number of micronuclei (MNi) was significantly higher for cells treated with Glu-BSA incubated at 60°C and purified via TCA (12 ± 1 MNi/1000 binucleated cells) compared to Glu-BSA incubated at 37°C and purified using UF (6 ± 1 MNi/1000 binucleated cells) after 9 days (P < 0.0001). The increase in genotoxicity observed could be explained by a higher level of protein glycation, oxidation, and aggregation of the Glu-BSA model system incubated at 60°C relative to 37°C. This study highlighted that the incubation temperature, purification method and cell exposure time are important variables to consider when generating AGEs in vitro and will enable future studies to better reflect in vivo situations of albumin glycation.

Can a digital slide scanner and viewing technique assist the visual scoring for the cytokinesis-block micronucleus cytome assay?

The cytokinesis-block micronucleus cytome (CBMNcyt) assay is a comprehensive method to measure DNA damage, cytostasis and cytotoxicity caused by nutritional, radiation and chemical factors. A slide imaging technique has been identified as a new method to assist with the visual scoring of cells for the CBMNcyt assay. A NanoZoomer S60 Digital Pathology slide scanner was used to view WIL2-NS cells treated with hydrogen peroxide (H2O2) and measure CBMNcyt assay biomarkers using a high-definition desktop computer screen. The H2O2-treated WIL2-NS cells were also scored visually using a standard light microscope, and the two visual scoring methods were compared. Good agreement was found between the scoring methods for all DNA damage indices (micronuclei, nucleoplasmic bridges and nuclear buds) and nuclear division index with correlation R values ranging from 0.438 to 0.789, P < 0.05. Apoptotic and necrotic cell frequency was lower for the NanoZoomer scoring method, but necrotic frequency correlated well with the direct visual microscope method (R = 0.703, P < 0.0001). Considerable advantages of the NanoZoomer scoring method compared to direct visual microscopy includes reduced scoring time, improved ergonomics and a reduction in scorer fatigue. This study indicates that a digital slide scanning and viewing technique may assist with visual scoring for the CBMNcyt assay and provides similar results to conventional direct visual scoring.

Antioxidant and Antiglycation Activities of Syzygium paniculatum Gaertn and Inhibition of Digestive Enzymes Relevant to Type 2 Diabetes Mellitus.

Advanced glycation end-products (AGEs) may be a contributing factor in the development of diabetes-specific vascular pathologies that affect the retina, glomerulus and peripheral nerves. In this study, Australian native food plant species Syzygium paniculatum was investigated for activities relevant to Type 2 diabetes mellitus including inhibition of α-amylase, α-glucosidase and protein glycation. A methanolic extract of the leaves showed the strongest α-amylase inhibition (IC50 = 14.29 ± 0.82 µg/mL, p < 0.05) when compared with other extracts. For inhibition of α-glucosidase, the strongest inhibition was shown for the water, methanolic and acetone extracts of leaves with IC50 values ranging from 4.73 ± 0.96 to 7.26 ± 0.92 µg/mL. In the BSA-glucose model, fruit and leaf extracts inhibited formation of protein-bound fluorescent AGEs with IC50 values ranging between 11.82 ± 0.71 and 96.80 ± 13.41 µg/mL. Pearson's correlation analysis showed that the AGE inhibition significantly correlated with DPPH (rp = -0.8964, p < 0.05) and ABTS (rp = -0.8326, p < 0.05). α-amylase inhibitory activities strongly correlated with DPPH (rp = -0.8964, p < 0.001). α-glucosidase inhibition strongly correlated with TPC (rp = -0.9243, p < 0.05), FRAP (rp = -0.9502, p < 0.01), DPPH (rp = -0.9317, p < 0.01) and ABTS (rp = -0.9486, p < 0.01). This study provides a strong rationale for further investigation aimed at isolating and identifying the active compounds responsible for the observed effects on targets relevant to diabetes.

Isolation and Structural Characterization of Echinocystic Acid Triterpenoid Saponins from the Australian Medicinal and Food Plant Acacia ligulata.

The Australian plant Acacia ligulata has a number of traditional food and medicinal uses by Australian Aboriginal people, although no bioactive compounds have previously been isolated from this species. Bioassay-guided fractionation of an ethanolic extract of the mature pods of A. ligulata led to the isolation of the two new echinocystic acid triterpenoid saponins, ligulatasides A (1) and B (2), which differ in the fine structure of their glycan substituents. Their structures were elucidated on the basis of 1D and 2D NMR, GC-MS, LC-MS/MS, and saccharide linkage analysis. These are the first isolated compounds from A. ligulata and the first fully elucidated structures of triterpenoid saponins from Acacia sensu stricto having echinocystic acid reported as the aglycone. Compounds 1 and 2 were evaluated for cytotoxic activity against a human melanoma cancer cell line (SK-MEL28) and a diploid fibroblast cell line (HFF), but showed only weak activity.

In vitro metabolism of the anti-inflammatory clerodane diterpenoid polyandric acid A and its hydrolysis product by human liver microsomes and recombinant cytochrome P450 and UDP-glucuronosyltransferase enzymes.

1. The metabolism of the anti-inflammatory diterpenoid polyandric acid A (PAA), a constituent of the Australian Aboriginal medicinal plant Dodonaea polyandra, and its de-esterified alcohol metabolite, hydrolysed polyandric acid A (PAAH) was studied in vitro using human liver microsomes (HLM) and recombinant UDP-glucuronosyltransferase (UGT) and cytochrome P450 (CYP) enzymes. 2. Hydrolysis of PAA to yield PAAH occurred upon incubation with HLM. Further incubations of PAAH with HLM in the presence of UGT and CYP cofactors resulted in significant depletion, with UGT-mediated depletion as the major pathway. 3. Reaction phenotyping utilising selective enzyme inhibitors and recombinant human UGT and CYP enzymes revealed UGT2B7 and UGT1A1, and CYP2C9 and CYP3A4 as the major enzymes involved in the metabolism of PAAH. 4. Analysis of incubations of PAAH with UDP-glucuronic acid-supplemented HLM and recombinant enzymes by UPLC/MS/MS identified three glucuronide metabolites. The metabolites were further characterised by ß-glucuronidase and mild alkaline hydrolysis. The acyl glucuronide of PAAH was shown to be the major metabolite. 5. This study demonstrates the in vitro metabolism of PAA and PAAH and represents the first systematic study of the metabolism of an active constituent of an Australian Aboriginal medicinal plant.

In vitro inhibitory activities of selected Australian medicinal plant extracts against protein glycation, angiotensin converting enzyme (ACE) and digestive enzymes linked to type II diabetes.

BACKGROUND: There is a need to develop potential new therapies for the management of diabetes and hypertension. Australian medicinal plants collected from the Kuuku I'yu (Northern Kaanju) homelands, Cape York Peninsula, Queensland, Australia were investigated to determine their therapeutic potential. Extracts were tested for inhibition of protein glycation and key enzymes relevant to the management of hyperglycaemia and hypertension. The inhibitory activities were further correlated with the antioxidant activities. METHODS: Extracts of five selected plant species were investigated: Petalostigma pubescens, Petalostigma banksii, Memecylon pauciflorum, Millettia pinnata and Grewia mesomischa. Enzyme inhibitory activity of the plant extracts was assessed against α-amylase, α-glucosidase and angiotensin converting enzyme (ACE). Antiglycation activity was determined using glucose-induced protein glycation models and formation of protein-bound fluorescent advanced glycation endproducts (AGEs). Antioxidant activity was determined by measuring the scavenging effect of plant extracts against 1, 1-diphenyl-2-picryl hydrazyl (DPPH) and using the ferric reducing anti-oxidant potential assay (FRAP). Total phenolic and flavonoid contents were also determined. RESULTS: Extracts of the leaves of Petalostigma banksii and P. pubescens showed the strongest inhibition of α-amylase with IC50 values of 166.50 ± 5.50 µg/mL and 160.20 ± 27.92 µg/mL, respectively. The P. pubescens leaf extract was also the strongest inhibitor of α-glucosidase with an IC50 of 167.83 ± 23.82 µg/mL. Testing for the antiglycation potential of the extracts, measured as inhibition of formation of protein-bound fluorescent AGEs, showed that P. banksii root and fruit extracts had IC50 values of 34.49 ± 4.31 µg/mL and 47.72 ± 1.65 µg/mL, respectively, which were significantly lower (p < 0.05) than other extracts. The inhibitory effect on α-amylase, α-glucosidase and the antiglycation potential of the extracts did not correlate with the total phenolic, total flavonoid, FRAP or DPPH. For ACE inhibition, IC50 values ranged between 266.27 ± 6.91 to 695.17 ± 15.38 µg/mL. CONCLUSIONS: The tested Australian medicinal plant extracts inhibit glucose-induced fluorescent AGEs, α-amylase, α-glucosidase and ACE with extracts of Petalostigma species showing the most promising activity. These medicinal plants could potentially be further developed as therapeutic agents in the treatment of hyperglycaemia and hypertension.

Development and Evaluation of a Topical Anti-Inflammatory Preparation Containing Dodonaea polyandra Extract.

PURPOSE: We have previously reported that the Australian Northern Kaanju (Kuuku I'yu) medicinal plant Dodonaea polyandra has anti-inflammatory activity. This is attributed largely to the presence of clerodane diterpenoids contained within the leaf resin. We envisaged developing a topical preparation to treat indications relating to skin inflammation. However, it was unknown whether the resin could be incorporated into a suitable dosage form while retaining the therapeutic value demonstrated in previous work. Therefore, the following study was undertaken to assess parameters of safety and efficacy for a prototype formulation containing the leaf resin extracted from D. polyandra. METHODS: Using the assessment criteria of optimum appearance, tactile feeling, spreadability and odour, 78 different formulations were developed. Formulation stability was assessed using a centrifugal test with preparations displaying phase separation further modified or re-formulated. A prototype formulation containing 5% w/w plant resin was selected and subjected to in vitro release studies. This was quantified through HPLC analysis using two major bioactive diterpenoids as reference. The prototype formulation was tested for efficacy in a TPA-induced acute murine skin inflammation model as well as a 3D human skin model for irritancy/toxicity (Epiderm™). RESULTS: The prototype resin cream was a chartreuse-coloured homogenous semisolid preparation that was readily spreadable upon contact with skin with no sensation of tackiness, residual greasiness, or irritation. The optimized cream showed no phase separation after 30 min centrifugation at 825 g. In the TPA-induced inflammation model, the resin formulation significantly reduced ear thickness and interleukin-1 beta levels in mouse ear tissue. The 5% w/w resin cream formulation showed no irritancy in a 3D human skin model. CONCLUSIONS: Our results demonstrate that bioactive resin from D. polyandra can be formulated into a stable and non-irritant semi-solid dosage form and reduce parameters of acute skin inflammation in vivo. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Polyandric acid A, a clerodane diterpenoid from the Australian medicinal plant Dodonaea polyandra, attenuates pro-inflammatory cytokine secretion in vitro and in vivo.

Dodonaea polyandra is a medicinal plant used traditionally by the Kuuku I'yu (Northern Kaanju) indigenous people of Cape York Peninsula, Australia. The most potent of the diterpenoids previously identified from this plant, polyandric acid A (1), has been examined for inhibition of pro-inflammatory cytokine production and other inflammatory mediators using well-established acute and chronic mouse ear edema models and in vitro cellular models. Topical application of 1 significantly inhibited interleukin-1ß production in mouse ear tissue in an acute model. In a chronic skin inflammation model, a marked reduction in ear thickness, associated with significant reduction in myeloperoxidase accumulation, was observed. Treatment of primary neonatal human keratinocytes with 1 followed by activation with phorbol ester/ionomycin showed a significant reduction in IL-6 secretion. The present study provides evidence that the anti-inflammatory properties of 1 are due to inhibition of pro-inflammatory cytokines associated with skin inflammation and may be useful in applications for skin inflammatory conditions including psoriasis and dermatitis.

A LC-MS/MS method for the simultaneous quantification of 17 opioids in biosolids.

The solid product of wastewater treatment plants is commonly used as a fertiliser to increase sustainability and waste reuse. It has undergone extensive treatment to remove high nutrient loads, pathogens and heavy metals but the extensive matrix of household chemicals, pesticides and pharmaceuticals remains, untargeted by most treatment technologies. These compounds, particularly pharmaceuticals, have been detected in biosolids with there being evidence of uptake by plants. With the current opioid pandemic in North America and overprescription, a simple method is required for the extraction of opioids from a solid medium as to ascertain the concentrations the environment is exposed to. A sonication-liquid-liquid extracted method was developed where biosolids were suspended in water and extracted using ethyl acetate before analysis on LC MS/MS. Sodium and potassium chloride were compared along with acidic and alkaline conditions. The optimised method utilised NaCl at a pH of 12 and was validated for 17 opioids, achieving linearity >0.987, 86-113% matrix effect and 0.1-10 µg/kg limits of detection. Upon analysis of biosolids destined for agriculture, 14 opioids were detected across all samples in a concentration range of 1-289 µg/kg.

Amphetamine-type drug-related fatalities in the context of 8 years of methamphetamine use measured by wastewater analysis in South Australia.

Methamphetamine is the illicit stimulant of choice in Australia. Countless initiatives have been employed to reduce methamphetamine use and drug-related harm. Wastewater analysis (WWA) has been used effectively as an objective measure of drug use at a population level and can be compared to indicators of harm, such as the number of drug-related fatalities. This paper attempts to describe methamphetamine use in the context of changes in levels measured in wastewater in South Australia whilst recognising considerable interventions over an 8-year period. Validated liquid chromatography-mass spectrometry methods were used to determine methamphetamine (and MDMA) levels in wastewater over an 8-year interval. The number of drug-induced deaths and driver fatalities involving methamphetamine (and MDMA) was presented and described in the context of changes in use measured by WWA. A rise in methamphetamine use according to WWA was evident up to 2017, followed by a gradual decrease to 2020 back to 2015 levels. Both driver fatalities and drug-induced deaths correlated well with use measured by WWA over the 8-year period. Multiple initiatives to curb supply, distribution and harm within the state and nationally have been implemented. The decrease in methamphetamine use after 2017 suggests that timely interventions have successfully reduced overall drug use and has led to fewer fatalities. This study shows that the response to increasing methamphetamine use in South Australia has resulted in a reversal of the upward trend in consumption and fewer drug-related fatalities.

Monitoring the use of novel psychoactive substances in Australia by wastewater-based epidemiology.

Users of novel psychoactive substances (NPS) are at risk, due to limited information about the toxicity and unpredictable effects of these compounds. Wastewater-based epidemiology (WBE) has been used as a tool to provide insight into NPS use at the population level. To understand the preferences and trends of NPS use in Australia, this study involved liquid chromatography mass spectrometry analysis of wastewater collected from Australian states and territories from February 2022 to February 2023. In total, 59 different NPS were included across two complementary analytical methods and covered up to 57 wastewater catchments over the study. The NPS detected in wastewater were 25-B-NBOMe, buphedrone, 1-benzylpiperazine (BZP), 3-chloromethcathinone, N,N-dimethylpentylone (N,N-DMP), N-ethylheptedrone, N-ethylpentylone, eutylone, 4F-phenibut, 2-fluoro deschloroketamine, hydroxetamine, mephedrone, methoxetamine, methylone, mitragynine, pentylone, phenibut, para-methoxyamphetamine (PMA), alpha-pyrrolidinovalerophenone (α-PVP) and valeryl fentanyl. The detection frequency for these NPS ranged from 3 % to 100 % of the sites analysed. A noticeable decreasing trend in eutylone detection frequency and mass loads was observed whilst simultaneously N,N-DMP and pentylone increased over the study period. The emergence of some NPS in wastewater pre-dates other sources of monitoring and provides further evidence that WBE can be used as an additional early warning system for alerting potential NPS use.

Seven-years of alcohol consumption in Australia by wastewater analysis: Exploring patterns by remoteness and socioeconomic factors.

BACKGROUND: Wastewater analysis provides a complementary measure of alcohol use in whole communities. We assessed absolute differences and temporal trends in alcohol consumption by degree of remoteness and socioeconomics indicators in Australia from 2016 to 2023. METHODS: Alcohol consumption estimates from 50 wastewater treatment plants (WWTP) in the Australian National Wastewater Drug Monitoring Program were used. Trends were analysed based on 1) site remoteness: Major Cities, Inner Regional and a combined remoteness category of Outer Regional and Remote, and 2) using two socioeconomic indexes from the Australian Bureau of Statistics (ABS) relating to advantage and disadvantage for Income, education, occupation, and housing. RESULTS: Consumption estimates were similar for Major Cities and Inner Regional areas (14.3 and 14.4L/day/1000 people), but significantly higher in Outer Regional and Remote sites (18.6L/day/1000 people). Consumption was decreasing in Major cities by 4.5% annually, Inner Regional by 2.4%, and 3.5% in the combined Outer Regional and Remote category. Consumption estimates were higher in socioeconomically advantaged quartiles than those of lower advantage (0%-25% mean = 13.0, 75%-100% mean = 17.4). Consumption in all quartiles decreased significantly over the 7 year period with annual rates of decrease of 0.9%, 3.7%, 3.6%, and 3.0% for the lowest to highest quartile, respectively. CONCLUSIONS: Declines in Australian alcohol consumption have been steeper in large urban areas than regional and remote areas. There were smaller annual decreases in the most socioeconomically disadvantaged areas. If continued, these trends may increase Australian health inequalities. Policy and prevention work should be appropriately targeted to produce more equitable long-term outcomes.

Learning from both sides: Experiences and opportunities in the investigation of Australian aboriginal medicinal plants.

With one of the oldest surviving cultures in the world, Australian Aboriginal people have developed immense knowledge about the diverse Australian flora. Western scientific investigation of some Australian Aboriginal medicinal plants has demonstrated interesting pharmacological activities and chemistry, however the majority of these species have not yet been extensively examined. We argue that research that is locally initiated and driven by Indigenous traditional owners in collaboration with Western scientists has significant potential to develop new plant-based products. Locally driven medicinal plants research in which traditional owners work as researchers in collaboration with University-based colleagues in the investigation of medicines rather than "stakeholders" or "informants" is one model that may be used in characterising plants with the potential to be developed into sustainable plant-based medicinal products with commercial value. Our team has taken this approach in research located both on traditional homelands and in the laboratory. Research being conducted by the University of South Australia and Chuulangun Aboriginal Corporation has led to patent filing for protection of intellectual property associated with novel compounds and extracts with the potential for development through cosmetic, complementary medicine and pharmaceutical routes. Ongoing research is examining the commercial developmental pathways and requirements for product development in these spaces. This review will address the opportunities that might exist for working in partnership with Australian Indigenous communities, some of the scientific knowledge which has been generated so far from our work together and the lessons learnt since the inception of the collaboration between the Chuulangun Aboriginal Corporation and scientists from the University of South Australia.

Dioecy in plants--is it an important factor for phytochemists to consider?

Plants with separate male and female flowers are termed dioecious. Dioecy is not rare, yet is a characteristic that could potentially impact the phytochemical and subsequent pharmacological properties of a species. This is a brief insight which highlights why the sex of the plant might be an important factor to consider for researchers within phytochemistry related fields.

Using wastewater-based epidemiology to evaluate the relative scale of use of opioids.

Wastewater analysis (WWA) has been used as a tool to monitor population drug use, both pharmaceutical and illicit, for over 15 years. Policymakers, law enforcement and treatment services may use WWA-derived data to seek an objective understanding of the extent of drug use in specific areas. Therefore, wastewater data should best be reported in a meaningful form to allow those that are not experts in the field to compare the scale within and between drug classes. Excreted drug loads quantified in wastewater describe the mass of drug present in the sewer. Normalisation for wastewater flow and population is standard practice and critical for comparing drug loads between different catchments and indicates a transition to an epidemiological approach (wastewater-based epidemiology (WBE)). A further consideration is necessary to accurately compare the measured level of one drug to another. The standard dose of a drug taken to elicit a therapeutic effect will vary, with some compounds requiring microgram amounts, while others are administered in the gram range. When WBE data is expressed with units representing excreted or consumed loads without considering dose amounts, the scale of drug use when comparing multiple compounds becomes distorted. To demonstrate the utility and significance of including known excretion rates, potency and typical dose amounts into back-calculations of the measured drug load, this paper compares the levels of 5 prescribed (codeine, morphine, oxycodone, fentanyl and methadone) and 1 illicit (heroin) opioid from South Australian wastewater. The data is presented at each stage of the back-calculation starting with the total mass load measured, to consumed amounts factoring in excretion rates and finally the number of doses the load equates to. This is the first paper to describe the levels of 6 opioids measured in wastewater over a 4-year period in South Australia that demonstrate the relative scale of use.

Partitioning of phytocannabinoids between faeces and water - Implications for wastewater-based epidemiology.

Evaluating consumption estimates for lipophilic drugs in wastewater has proven to be a challenge. A common feature for these compounds is that they are excreted in faeces and in conjugated form in urine. Limited research with no obvious experimental evidence has been conducted to investigate the degree to which faecal-bound chemical markers contribute towards mass loads in wastewater. Cannabis chemical markers, known as phytocannabinoids, have been suggested in literature to fall into this category. In this study, cannabis users (n = 9) and non-cannabis users (n = 5) were recruited and provided faecal and urine samples after using the substance. The common chemical markers of cannabis consumption, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THC-COOH), 11-hydroxy-Δ9-tetrahydrocannabinol (THC-OH), Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD), were investigated. An extraction method was developed for the cannabis chemical markers in faecal matter and urine and analysis was performed by liquid chromatography-mass spectrometry. Participant samples were used to establish adsorption and desorption dissolution kinetics models and to assess the equilibrium between faeces and water for these compounds. Equilibration between phases were found to be fast (<5 min). THC-COOH, which is the primary metabolite used in wastewater studies, partitioned ~40% in water while the less polar metabolite and CBD remained largely associated with the particulate fraction. Faecal loads of both cannabis users and non-users affected the total measured amounts of cannabinoids in the aqueous phase. The implications for wastewater monitoring of lipophilic substances are discussed.