Singapore's Total Diet Study (2021-2023): Study Design, Methodology, and Relevance to Ensuring Food Safety.

A total diet study is often used to evaluate a population's baseline dietary exposure to chemical hazards from across the diet. In 2021-2023, Singapore carried out a TDS, and this article presents an overview of the study design and methodological selections in Singapore's TDS, as well as its relevance to ensuring food safety. A food consumption survey was conducted on Singapore citizens and permanent residents, where food consumption patterns of the Singapore population were identified. The selection of chemical hazards and foods for inclusion in Singapore's TDS, as well as principal considerations on sampling, food preparation, and analytical testing are discussed. Commonly consumed foods by the Singapore population in food categories such as grain and grain-based products, meat and meat products, fish and seafood, vegetables, fruits, milk and dairy products were included in this study, and mean concentrations of chemicals tested in each food category were reported, with food categories possessing higher levels identified. Future work will include dietary exposure assessments for the population and analysis of the contributions by food and cooking method.

Surveillance of veterinary drug residues in food commonly consumed in Singapore and assessment of dietary exposure.

Non-judicious and indiscriminate use of veterinary drugs in animal husbandry may result in accumulation of residues in animal tissues, and consequently in food for human consumption. The abuse of veterinary drugs presents a potential risk to consumer health, especially if the residue level is higher than the health-based guidance value (HBGV) such as the acceptable daily intake (ADI). Contamination of drug residues in food also promotes the emergence of antimicrobial resistance (AMR) which poses a serious threat to public health globally. There has been limited information on the occurrence and dietary exposure to veterinary drug residues in Singapore to date. In this study, the occurrence of four classes of veterinary drugs, namely beta-agonists, coccidiostats, fluoroquinolones and macrolides, were determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in food widely consumed by Singapore residents. The magnitude of dietary exposure was assessed based on the consumption profile of Singapore population. Out of 216 food samples, 9.72 % were detected positive with veterinary drug residues, where majority of the positive samples were poultry and its derived products, followed by eggs and egg products. 7 veterinary drugs, specifically ciprofloxacin, enrofloxacin, clopidol, diclazuril, lasalocid, nicarbazin and tilmicosin, were detected in the samples, with clopidol and enrofloxacin being the most frequently detected drugs. Dietary exposure was evaluated using the estimated daily intake (EDI) of the detected drugs and benchmarked against the corresponding acceptable daily intake (ADI). All the %ADI values were far less than 100 in both the average and high consumer scenarios, indicating that the health risk associated with dietary exposure to these drugs in Singapore is low.

Occurrence and Dietary Exposure to Acrylamide from Foods Consumed within and outside Main Meals in Singapore.

This study investigated the influence of 'snackification' in Singaporean diets, leading to increased dietary acrylamide exposure. Acrylamide concentrations in commonly consumed foods within and outside the main meals were measured using liquid chromatography with tandem mass spectrometry (LC-MS/MS). High acrylamide concentrations were detected in vegetables cooked at high temperatures (ranging from 0.5 to 478.4 µg/kg) and potato-based crackers and chips (ranging from 81.8 to 2095.8 µg/kg). The estimated total dietary exposure for the Singapore population was 0.165 µg/kg bw/day for general consumers and 0.392 µg/kg bw/day for high consumers (95th percentile). The acrylamide exposure from outside main meals was nearly equivalent to that from within the main meals. The calculated margins of exposure (MOE) were below 10,000, indicating potential human health concern. These findings highlight the need for industry practices and consumer advisories to reduce acrylamide exposure from foods consumed both within and outside main meals.

Food Safety Controls during Bulk Food Preparation-An Observational Analysis.

Seeing the palpable impact of food poisoning associated with catering operations, we surveyed caterers with and without past hygiene violations to examine staffing, food safety practices and correlations with microbial counts in food and the processing environment. Past infringements did not negatively impact the current execution of food safety measures nor the microbial quality of food. In preference to added stringencies for errant operators, we discuss alternative efforts to augment food safety, as well as the policy implications thereof.

Occurrence and Dietary Exposure of 3-MCPD Esters and Glycidyl Esters in Domestically and Commercially Prepared Food in Singapore.

This study investigated the prevalence and occurrence of 3-monochloropropanediol esters (3-MCPDEs) and glycidyl esters (GEs) in domestically and commercially prepared food in Singapore and assessed the total dietary exposure for the Singaporean population. Minimal impact on the formation of 3-MCPDEs and GEs was observed from the domestic cooking methods commonly practiced in Singapore such as deep frying and stir frying. The estimated total dietary exposure to 3-MCPDEs for the Singaporean population (aged 15 to 92) was 0.982 µg/kg bw/day for general consumers and 2.212 µg/kg bw/day for high consumers (95th percentile), which accounted for 49.1% and 110.6% of the tolerable dietary intake (TDI) at 2 µg/kg bw/day by the European Food Safety Authority (EFSA). The calculated margins of exposure (MOE) for GEs based on the dietary exposure for general consumers at 0.882 µg/kg bw/day and 2.209 µg/kg bw/day for high consumers were below 10,000, indicating a potential health concern. Our study showed that the occurrence of 3-MCPDEs and GEs varied among vegetable oils, and domestic cooking methods did not significantly impact the levels of 3-MCPDEs and GEs in prepared food. The critical factor influencing the prevalence and occurrence of 3-MCPDEs and GEs was the choice of oil used for cooking, which absorbed into the cooked food. It is essential to encourage the food industry to continue its innovation on mitigation measures to control and reduce 3-MCPDEs and GEs in vegetable oil production. Consumers are advised to make informed choices on food consumption and cooking oil for food preparation to reduce their exposure to 3-MCPDEs and GEs.

Consumption Patterns of Processed Foods in Singapore-A Cross-Sectional Study.

The consumption of processed foods is increasingly widespread and could have an impact on diet quality and health. Understanding the factors influencing people's eating habits is useful for assessing such impact. There are limited data on the consumption patterns of processed foods and associated factors influencing the dietary patterns in Singapore. This cross-sectional study based on a food frequency questionnaire aimed to examine how the consumption of processed foods among 2079 Singapore residents aged 18 to 89 years varies with sociodemographic factors. The analysis of the consumption by processed food groups showed that the studied factors, i.e., age, gender, ethnicity, housing and health status, all contributed to differences in processed food consumption to varying extents, with ethnicity being the key factor driving the variation. Such differences were also confirmed to a limited degree by determining another measure of consumption, i.e., a processed food variety score. The findings in this study could inform further work in relation to dietary risks.

Application of Neuron-Selective Fluorescent Probe, NeuA, To Identify Mouse Retinal Degeneration.

The retina is part of the central nerve system (CNS) and has various interneurons and sensory neurons such as photoreceptor cells. Retinitis pigmentosa (RP) is an inherited condition that is characterized by photoreceptor degeneration. Herein, we developed a fluorescent probe-NeuA-for detecting retinal neuronal cells and applied NeuA to discriminate between healthy and RP retinas. The staining pattern of NeuA in the retinas of healthy and RP mouse models was examined in vitro, ex vivo and in vivo using confocal microscopy, the fluorescent fundus microscopy and optical coherent tomography (OCT). NeuA strongly stained the outer segment layer of photoreceptor cells and some bipolar cells in the healthy retina, but there was only weak staining in the photoreceptor degenerated retinas. Therefore, NeuA probe can be used as the detecting RP tools in the preclinical conditions.

A highly selective fluorogenic probe for the detection and in vivo imaging of Cu/Zn superoxide dismutase.

Copper/zinc superoxide dismutase (Cu/Zn SOD) is an essential enzyme that protects tissue from oxidative damage. Herein we report the first fluorogenic probe (SODO) for the detection and in vivo imaging of Cu/Zn SOD. SODO represents a unique chemical probe for translational imaging studies of Cu/Zn SOD in inflammatory disorders.

A Multisite-Binding Switchable Fluorescent Probe for Monitoring Mitochondrial ATP Level Fluctuation in Live Cells.

Adenosine triphosphate (ATP), commonly produced in mitochondria, is required by almost all the living organisms; thus fluorescent probes for monitoring mitochondrial ATP levels fluctuation are essential and highly desired. Herein, we report a multisite-binding switchable fluorescent probe, ATP-Red 1, which selectively and rapidly responds to intracellular concentrations of ATP. Live-cell imaging indicated that ATP-Red 1 mainly localized to mitochondria with good biocompatibility and membrane penetration. In particular, with the help of ATP-Red 1, we successfully observed not only the decreased mitochondrial ATP levels in the presence of KCN and starvation state, but also the increased mitochondrial ATP levels in the early stage of cell apoptosis. These results indicate that ATP-Red 1 is a useful tool for investigating ATP-relevant biological processes.

Development of fluorescent probes specific for parallel-stranded G-quadruplexes by a library approach.

A 241-membered cyanine-based library was constructed by the combinatorial chemistry strategy. Combined with high-throughput screening, we successfully discovered a novel fluorescent probe (CyC-M716) capable of identifying a subset of parallel G-quadruplexes with propeller loops stretching across three tetrad layers with high sensitivity and selectivity.

NeuO: a fluorescent chemical probe for live neuron labeling.

To address existing limitations in live neuron imaging, we have developed NeuO, a novel cell-permeable fluorescent probe with an unprecedented ability to label and image live neurons selectively over other cells in the brain. NeuO enables robust live neuron imaging and isolation in vivo and in vitro across species; its versatility and ease of use sets the basis for its development in a myriad of neuronal targeting applications.

High-efficiency in vitro and in vivo detection of Zn2+ by dye-assembled upconversion nanoparticles.

Development of highly sensitive and selective sensing systems of divalent zinc ion (Zn(2+)) in organisms has been a growing interest in the past decades owing to its pivotal role in cellular metabolism, apoptosis, and neurotransmission. Herein, we report the rational design and synthesis of a Zn(2+) fluorescent-based probe by assembling lanthanide-doped upconversion nanoparticles (UCNPs) with chromophores. Specifically, upconversion luminescence (UCL) can be effectively quenched by the chromophores on the surface of nanoparticles via a fluorescence resonant energy transfer (FRET) process and subsequently recovered upon the addition of Zn(2+), thus allowing for quantitative monitoring of Zn(2+). Importantly, the sensing system enables detection of Zn(2+) in real biological samples. We demonstrate that this chromophore-UCNP nanosystem is capable of implementing an efficient in vitro and in vivo detection of Zn(2+) in mouse brain slice with Alzheimer's disease and zebrafish, respectively.

NeuO for Neuronal Labeling in Zebrafish.

We report the application of our newly developed fluorescent probe 3-(benzylamino)-4,4-difluoro-5-(4-propyl-1H-1,2,3-triazol-1-yl)-8-(4-(2-hydroxyacetamido)phen-yl)-4-bora-3a,4a-diaza-s-indacene (NeuO) to label and image live neurons in zebrafish. Immersing zebrafish embryos in NeuO or injecting NeuO into zebrafish brain ventricles results in nontoxic in vivo neuronal labeling. We demonstrate the applicability of NeuO and envisage the potential of this compound as a rapid and simple labeling reagent for studying neuron development and degeneration.

"Orange alert": a fluorescent detector for bisphenol A in water environments.

Due to the prevalent use of polycarbonate plastics and epoxy resins in packaging materials and paints for ships, there has been a widespread global contamination of environmental water sources with bisphenol A (BPA). BPA, an endocrine disruptor, has been found to cause tremendous health problems. Therefore, there is an urgent need for detecting BPA in a convenient and sensitive manner to ensure water safety. Herein, we develop a fluorescent turn-on BPA probe, named Bisphenol Orange (BPO), which could conveniently detect BPA in a wide variety of real water samples including sea water, drain water and drinking water. BPO shows superior selectivity toward BPA and up to 70-fold increase in fluorescence emission at 580 nm when mixed with BPA in water. Mechanistic studies suggest a plausible water-dependent formation of hydrophobic BPA clusters which favorably trap and restrict the rotation of BPO and recover its inherent fluorescence.

Discovery of a structural-element specific G-quadruplex "light-up" probe.

The development of a fluorescent probe capable of detecting and distinguishing the wide diversity of G-quadruplex structures is particularly challenging. Herein, we report a novel BODIPY-based fluorescent sensor (GQR) that shows unprecedented selectivity to parallel-stranded G-quadruplexes with exposed ends and four medium grooves. Mechanistic studies suggest that GQR associates with G-quadruplex grooves close to the end of the tetrad core, which may explain the dye's specificity to only a subset of parallel structures. This specific recognition favours the disaggregation of GQR in aqueous solutions thereby recovering the inherent fluorescence of the dye. Due to its unique features, GQR represents a valuable tool for basic biological research and the rapid discovery of novel, specific ligands that target similar structural features of G-quadruplexes.

Fluorescent dye cocktail for multiplex drug-site mapping on human serum albumin.

Elucidating how molecules bind to HSA is fundamental for predicting drug incompatibilities. Through combinatorial screening, we identified a novel fluorescent dye (BD140) with turn-on fluorescence emission and specific binding at HSA drug site 2. We further combined it with dansylamide to develop a fluorescent dye cocktail for high-throughput mapping of the interaction between therapeutics at HSA drug-binding sites.

Make caffeine visible: a fluorescent caffeine "traffic light" detector.

Caffeine has attracted abundant attention due to its extensive existence in beverages and medicines. However, to detect it sensitively and conveniently remains a challenge, especially in resource-limited regions. Here we report a novel aqueous phase fluorescent caffeine sensor named Caffeine Orange which exhibits 250-fold fluorescence enhancement upon caffeine activation and high selectivity. Nuclear magnetic resonance spectroscopy and Fourier transform infrared spectroscopy indicate that π-stacking and hydrogen-bonding contribute to their interactions while dynamic light scattering and transmission electron microscopy experiments demonstrate the change of Caffeine Orange ambient environment induces its fluorescence emission. To utilize this probe in real life, we developed a non-toxic caffeine detection kit and tested it for caffeine quantification in various beverages. Naked-eye sensing of various caffeine concentrations was possible based on color changes upon irradiation with a laser pointer. Lastly, we performed the whole system on a microfluidic device to make caffeine detection quick, sensitive and automated.

Lewis basic selenium catalyzed chloroamidation of olefins using nitriles as the nucleophiles.

A Lewis base catalyzed chloroamidation of olefinic substrates was achieved using diphenyl selenide as the catalyst. The reaction conditions are mild and suitable for a wide range of substrates including those which are acid labile.

Rational exploration of N-heterocyclic carbene (NHC) palladacycle diversity: a highly active and versatile precatalyst for Suzuki-Miyaura coupling reactions of deactivated aryl and alkyl substrates.

As less attention has been focussed on the design of highly efficient palladium precatalysts to ensure the smooth formation of the active catalyst for metal-mediated cross coupling reactions, we herein demonstrate that combining the bulky N-heterocyclic carbene (NHC) 1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene (IPr) with cyclopalladated acetanilide as the optimal palladium precatalyst leads to superior catalytic activity compared with the state-of-the-art NHC-Pd catalysts. The complex was discovered through the evaluation of a small, rationally designed library of NHC-palladacycles prepared by a novel, practical and atom-economic method, the direct reaction of IPrHCl with palladacycle acetate dimers.