Browsing by Author "Dheer Singh"

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Advances in analytical methods for BPA detection in commercial milk: Current techniques and future prospects
(Academic Press Inc., 2025) Hagera Dilnashin; Dheer Singh; Partha Pratim Roy; Rakesh Kumar Tyagi; Surya Pratap Singh
Bisphenol A (BPA) is a widely produced chemical, primarily used in the manufacture of polycarbonate plastics and epoxy resins. It is commonly found in food and beverage packaging, such as milk, water, and infant bottles, as well as in dental sealants and food can coatings. The primary source of BPA exposure in humans is dietary intake, with milk and dairy products contributing significantly, particularly among younger populations. Other exposure pathways include air, water, and dust. This review provides an overview of the analytical methods developed in recent years for detecting BPA. It examines the advantages and limitations of various sample preparation techniques, including microextraction, molecularly imprinted polymer solid-phase extraction, and solid-phase extraction. Furthermore, we discuss the advancements in analytical techniques, such as immunoassays, liquid and gas chromatography–mass spectrometry, and emerging biosensors. The review highlights recent progress in BPA detection methodologies and outlines potential future developments in this field. © 2024 Elsevier Inc.
Detection of endocrine and metabolism disrupting xenobiotics in milk-derived fat samples by fluorescent protein-tagged nuclear receptors and live cell imaging
(Taylor and Francis Ltd., 2023) Keshav Thakur; Emmagouni Sharath Kumar Goud; Yashika Jawa; Chetan Keswani; Suneel Onteru; Dheer Singh; Surya P. Singh; Partha Roy; Rakesh K. Tyagi
Nuclear receptors (NRs) are ligand-modulated transcription factors that regulate multiple physiological functions in our body. Many NRs in their unliganded state are localized in the cytoplasm. The ligand-inducible nuclear translocation of NRs provides a valuable tool for studying the NR-ligand interactions and their downstream effects. The translocation response of NRs can be studied irrespective of the nature of the interacting ligand (agonist, antagonist, or a small molecule modulator). These nuclear translocation studies offer an advantage over promoter-reporter-based transcription assays where transcription response is observed only with the activating hormones or agonistic ligands. Globally, milk serves as a major dietary source. However, suspected presence of endocrine/metabolism-disrupting chemicals like bisphenols, parabens, organochlorine pesticides, carbamates, non-steroidal anti-inflammatory drugs, chloramphenicol, brominated flame retardants, etc. has been reported. Considering that these chemicals may impart serious developmental and metabolism-related health concerns, it is essential to develop assays suitable for the detection of xenobiotics present at differing levels in milk. Since milk samples cannot be used directly on cultured cells or for microscopy, a combination of screening strategies has been developed herein based on the revelation that i) lipophilic NR ligands can be successfully retrieved in milk-fat; ii) milk-fat treatment of cells is compatible with live-cell imaging studies; and finally, iii) treatment of cells with xenobiotics-spiked and normal milk derived fat provides a visual and quantifiable response of NR translocation in living cells. Utilizing a milk-fat extraction method and Green Fluorescent Protein (GFP) tagged NRs expressed in cultured mammalian cells, followed by an assessment of NR response proved to be an effective approach for screening xenobiotics present in milk samples.Highlights Diverse endocrine and metabolism-disrupting chemicals are suspected to contaminate milk. Nuclear receptors serve as ‘xenosensors’ for assessing the presence of xenobiotics in milk. Nuclear import of steroid receptors with (ant)agonist can be examined in live cells. Lipophilic xenobiotics are extracted and observed enriched in milk-fat fraction. A comprehensive cell-based protocol aids in the detection of xenobiotics in milk. © 2022 Informa UK Limited, trading as Taylor & Francis Group.
Evaluation of (Anti)androgenic Activities of Environmental Xenobiotics in Milk Using a Human Liver Cell Line and Androgen Receptor-Based Promoter-Reporter Assay
(American Chemical Society, 2022) Himanshu Agrawal; Keshav Thakur; Shreyasi Mitra; Debarghya Mitra; Chetan Keswani; Debabrata Sircar; Suneel Onteru; Dheer Singh; Surya P. Singh; Rakesh K. Tyagi; Partha Roy
The recent reports on milk consumption and its associated risk with hormone related disorders necessitates the evaluation of dairy products for the presence of endocrine disrupting chemicals (EDCs) and ensure the safety of consumers. In view of this, we investigated the possible presence of (anti)androgenic contaminants in raw and commercialized milk samples. For this purpose, a novel HepARE-Luc cell line that stably expresses human androgen receptor (AR) and the androgen responsive luciferase reporter gene was generated and used in the present study. Treatment of this cell line with androgens and corresponding antiandrogen (flutamide) stimulated or inhibited expression of reporter luciferase, respectively. Real time polymerase chain reaction and immunostaining results exhibited transcription response and translocation of AR from the cytoplasm to the nucleus in response to androgen. Observations implied that a cell-based xenobiotic screening assay via AR response can be conducted for assessing the (anti)androgenic ligands present in food chain including milk. Therefore, the cell line was further used to screen the (anti)androgenic activity of a total of 40 milk fat samples procured as raw or commercial milk. Some of the raw and commercial milk fat samples distinctly showed antiandrogenic activities. Subsequently, some commonly used environmental chemicals were also evaluated for their (anti)androgenic activities. Initial observations with molecular docking studies of experimental compounds were performed to assess their interaction with AR ligand binding domain. Furthermore, (anti)androgenic activities of these compounds were confirmed by performing luciferase assay using the HepARE-Luc cell line. None of the test compounds showed androgenic activities rather some of them like Bisphenol A (BPA) and rifamycin showed antiandrogenic activities. In conclusion, our results provide a valuable information about the assessment of (anti)androgenic activities present in milk samples. Overall, it is proposed that a robust cell-based CALUX assay can be used to assess the (anti)androgenic activities present in milk which can be attributed to different environmental chemicals present therein. © 2022 American Chemical Society. All rights reserved.
Global footprints of organochlorine pesticides: a pan-global survey
(Springer Science and Business Media B.V., 2022) Chetan Keswani; Hagera Dilnashin; Hareram Birla; Partha Roy; Rakesh K. Tyagi; Dheer Singh; Vishnu D. Rajput; Tatiana Minkina; Surya P. Singh
Organochlorine pesticides (OCPs) are ubiquitous environmental contaminants widely used all over the world. These chlorinated hydrocarbons are toxic and often cause detrimental health effects because of their long shelf life and bioaccumulation in the adipose tissues of primates. OCP exposure to humans occurs through skin, inhalation and contaminated foods including milk and dairy products, whereas developing fetus and neonates are exposed through placental transfer and lactation, respectively. In 1960s, OCPs were banned in most developed countries, but because they are cheap and easily available, they are still widely used in most third world countries. The overuse or misuse of OCPs has been rising continuously which pose threats to environmental and human health. This review reports the comparative occurrence of OCPs in human and bovine milk samples around the globe and portrays the negative impacts encountered through the long history of OCP use. © 2021, The Author(s), under exclusive licence to Springer Nature B.V.
Organochlorine pesticide dieldrin upregulate proximal promoter (PII) driven CYP19A1 gene expression and increases estrogen production in granulosa cells
(Elsevier Inc., 2021) Deeksha Sharma; Suman Kumari; Payal Rani; Suneel Kumar Onteru; Partha Roy; Rakesh Kumar Tyagi; Surya Pratap Singh; Dheer Singh
Organochlorine pesticides are highly persistent environmental pollutants, generally shown to act through estrogen receptor alpha and alter estrogen biosynthesis. However, the molecular mechanism of regulation of estrogen biosynthesis by these pesticides is not clear. Estrogen is main female fertility hormone regulated by rate-limiting enzyme aromatase. It is encoded by the CYP19A1 gene, which is expressed using specific promoters. In the present study, the attempt has been made to elucidate the effect of dieldrin on the promoter-specific CYP19A1 gene expression and estrogen hormone production in buffalo granulosa cells. The buffalo granulosa cells were cultured and treated with dieldrin in a dose (100,150 and 200 ng/mL) and time (6, 12, and 24 h) dependent manner, followed by analysis of CYP19A1, promoter-specific CYP19A1 transcript expression, and estrogen production. Results showed that dieldrin significantly increased the expression of the CYP19A1 gene after 6 and 12 h while its expression was decreased after 24 h. To understand the upregulation of CYP19A1 gene, promoters’ specific CYP19A1 transcript analysis was done. The finding showed that dieldrin significantly increased the proximal promoter specific CYP19A1 transcript while there was no effect on distal promoter specific CYP19A1 transcripts. This specific-promoter activity was quantified by chromatin immunoprecipitation assay (ChIP). Results confirmed the involvement of the proximal promoter in the overexpression of CYP19A1 gene. Furthermore, a significant increase in estradiol-17β level was also observed. Overall, the present study demonstrated the stimulatory effect of dieldrin on the CYP19A1 gene and will help to understand the toxicological role of dieldrin on the reproductive system. © 2021
Process parameter optimization for textural properties of ready-to-eat extruded snack food from millet and legume pieces blends
(2009) Subir Kumar Chakraborty; Daya S. Singh; B.K. Kumbhar; Dheer Singh
Five blends of millets and legume pieces (viz. 12, 16, 20, 24, 28% legume pieces) at varying moisture contents (12, 15, 18, 21, 24%, wet basis [w.b.]) were subjected to extrusion cooking in a single-screw laboratory extruder Brabender DA47055. The extruder was operated at different die head temperatures (160-200C), barrel temperatures (100-140C) and screw speeds (100-140 rpm). Designed experiments were conducted based on central composite rotatable design to determine the optimum processing condition for extruded products. The processing parameters were optimized with the help of response surface methodology to obtain product with maximum crispness and with minimum hardness and cutting strength. The optimum processing conditions obtained were moisture content 24%, w.b.; blend ratio 18.7% legume pieces; die head temperature 171.2C; barrel temperature 140C; and screw speed 103.8 rpm. The corresponding textural properties of the extrudates in terms of crispness, hardness and cutting strength were 50, 48.2 N and 79.7 N, respectively. © 2009, Wiley Periodicals, Inc.