Displaying publications 61 - 77 of 77 in total

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  1. Molecular basis of the anti-diabetic properties of camel milk through profiling of its bioactive peptides on dipeptidyl peptidase IV (DPP-IV) and insulin receptor activity
    Ashraf A, Mudgil P, Palakkott A, Iratni R, Gan CY, Maqsood S, et al.
    J Dairy Sci, 2021 Jan;104(1):61-77.
    PMID: 33162074 DOI: 10.3168/jds.2020-18627
    The molecular basis of the anti-diabetic properties of camel milk reported in many studies and the exact active agent are still elusive. Recent studies have reported effects of camel whey proteins (CWP) and their hydrolysates (CWPH) on the activities of dipeptidyl peptidase IV (DPP-IV) and the human insulin receptor (hIR). In this study, CWPH were generated, screened for DPP-IV binding in silico and inhibitory activity in vitro, and processed for peptide identification. Furthermore, pharmacological action of intact CWP and their selected hydrolysates on hIR activity and signaling and on glucose uptake were investigated in cell lines. Results showed inhibition of DPP-IV by CWP and CWPH and their positive action on hIR activation and glucose uptake. Interestingly, the combination of CWP or CWPH with insulin revealed a positive allosteric modulation of hIR that was drastically reduced by the competitive hIR antagonist. Our data reveal for the first time the profiling and pharmacological actions of CWP and their derived peptides fractions on hIR and their pathways involved in glucose homeostasis. This sheds more light on the anti-diabetic properties of camel milk by providing the molecular basis for the potential use of camel milk in the management of diabetes.
  2. New insights into the cholesterol esterase- and lipase-inhibiting potential of bioactive peptides from camel whey hydrolysates: Identification, characterization, and molecular interaction
    Baba WN, Mudgil P, Baby B, Vijayan R, Gan CY, Maqsood S
    J Dairy Sci, 2021 Jul;104(7):7393-7405.
    PMID: 33934858 DOI: 10.3168/jds.2020-19868
    Novel antihypercholesterolemic bioactive peptides (BAP) from peptic camel whey protein hydrolysates (CWPH) were generated at different time, temperature, and enzyme concentration (%). Hydrolysates showed higher pancreatic lipase- (PL; except 3 CWPH) and cholesterol esterase (CE)-inhibiting potential, as depicted by lower half-maximal inhibitory concentration values (IC50 values) compared with nonhydrolyzed camel whey proteins (CWP). Peptide sequencing and in silico data depicted that most BAP from CWPH could bind active site of PL, whereas as only 3 peptides could bind the active site of CE. Based on higher number of reactive residues in the BAP and greater number of substrate binding sites, FCCLGPVPP was identified as a potential CE-inhibitory peptide, and PAGNFLPPVAAAPVM, MLPLMLPFTMGY, and LRFPL were identified as PL inhibitors. Molecular docking of selected peptides showed hydrophilic and hydrophobic interactions between peptides and target enzymes. Thus, peptides derived from CWPH warrant further investigation as potential candidates for adjunct therapy for hypercholesterolemia.
  3. Simulated gastrointestinal digestion of camel and bovine casein hydrolysates: Identification and characterization of novel anti-diabetic bioactive peptides
    Mudgil P, Kamal H, Priya Kilari B, Mohd Salim MAS, Gan CY, Maqsood S
    Food Chem, 2021 Aug 15;353:129374.
    PMID: 33740505 DOI: 10.1016/j.foodchem.2021.129374
    Camel milk proteins are an important substrate for bioactive peptides generation. This study investigates in-vitro antidiabetic effect (via inhibition of α-amylase (AA), α-glucosidase (AG) and dipeptidyl peptidase IV (DPP-IV)) of bovine (BC) and camel casein (CC) hydrolysates. Further, effect of simulated gastrointestinal digestion (SGID) on inhibitory potential of generated hydrolysates was also explored. Both BC and CC hydrolysates displayed potent inhibitory properties against AA (IC50 value- 0.58 & 0.59 mg/mL), AG (IC50 value- 1.04 & 0.59 mg/mL) and DPP-IV (IC50 value- 0.62 & 0.66 mg/mL), respectively. Among different peptides identified in BC and CC hydrolysates, it was observed that FLWPEYGAL was predicted to be most potent inhibitory peptide against AA. While LPTGWLM, MFE and GPAHCLL as most active inhibitor of AG and HLPGRG, QNVLPLH and PLMLP were predicted to be active against DPP-IV. Overall, BC and CC hydrolysates can be proposed to be used in different food formulations as functional antidiabetic agents.
  4. ACE Inhibitory and Antioxidant Activities of Collagen Hydrolysates from the Ribbon Jellyfish (Chrysaora sp.)
    Barzideh Z, Latiff AA, Gan CY, Abedin MZ, Alias AK
    Food Technol Biotechnol, 2014 Dec;52(4):495-504.
    PMID: 27904323
    Collagen isolated from the ribbon jellyfish (Chrysaora sp.) was hydrolysed using three different proteases (i.e. trypsin, alcalase and Protamex) to obtain bioactive peptides. Angiotensin-I-converting enzyme (ACE) inhibitory activity and antioxidant activities (i.e. ferric reducing antioxidant power (FRAP) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity) of the peptides were measured and compared, and the effect of the duration of hydrolysis on the bioactivity (ACE inhibitory and antioxidant activities) of peptides was also evaluated. FRAP activity was the highest in Protamex-induced (25-27 mM) and trypsin-induced hydrolysates (24-26 mM) at 7 and 9 h, respectively. Conversely, hydrolysates produced by trypsin for 1 and 3 h showed the highest DPPH radical scavenging activities (94 and 92%, respectively). Trypsin-induced hydrolysates (at 3 h) also showed the highest ACE inhibitory activity (89%). The peptide sequences with the highest activities were identified using tandem mass spectrometry, and the results show that the hydrolysates had a high content of hydrophobic amino acids as well as unique amino acid sequences, which likely contribute to their biological activities.
  5. Fulltext Camel whey protein hydrolysates induced G2/M cellcycle arrest in human colorectal carcinoma
    Murali C, Mudgil P, Gan CY, Tarazi H, El-Awady R, Abdalla Y, et al.
    Sci Rep, 2021 03 29;11(1):7062.
    PMID: 33782460 DOI: 10.1038/s41598-021-86391-z
    Camel milk has been gaining immmense importance due to high nutritious value and medicinal properties. Peptides from milk proteins is gaining popularity in various therapeutics including human cancer. The study was aimed to investigate the anti-cancerous and anti-inflammatory properties of camel whey protein hydrolysates (CWPHs). CWPHs were generated at three temperatures (30 ℃, 37 ℃, and 45 ℃), two hydrolysis timepoints (120 and 360 min) and with three different enzyme concentrations (0.5, 1 and 2 %). CWPHs demonstrated an increase in anti-inflammatory effect between 732.50 (P-6.1) and 3779.16 (P-2.1) µg Dicolfenac Sodium Equivalent (DSE)/mg protein. CWPHs (P-4.3 & 5.2) inhibited growth of human colon carcinoma cells (HCT116) with an IC50 value of 231 and 221 μg/ml, respectively. P-4.3 induced G2/M cell cycle arrest and modulated the expression of Cdk1, p-Cdk1, Cyclin B1, p-histone H3, p21 and p53. Docking of two peptides (AHLEQVLLR and ALPNIDPPTVER) from CWPHs (P-4.3) identified Polo like kinase 1 as a potential target, which strongly supports our in vitro data and provides an encouraging insight into developing a novel peptide-based anticancer formulation. These results suggest that the active component, CWPHs (P-4.3), can be further studied and modeled to form a small molecule anti-cancerous therapy.
  6. Novel antihypertensive peptides from lupin protein hydrolysate: An in-silico identification and molecular docking studies
    Fadimu GJ, Gan CY, Olalere OA, Farahnaky A, Gill H, Truong T
    Food Chem, 2023 May 01;407:135082.
    PMID: 36493485 DOI: 10.1016/j.foodchem.2022.135082
    Application of non-thermal treatment to proteins prior to enzymatic hydrolysis can facilitate the release of novel bioactive peptides (BPs) with unique biological activities. In this study, lupin protein isolate was pre-treated with ultrasound and hydrolysed using alcalase and flavourzyme to produce alcalase hydrolysate (ACT) and flavourzyme hydrolysate(FCT). These hydrolysates were fractionated into 1, 5, and 10 kDa molecular weight fractions using a membrane ultrafiltration technique. The in vitro angiotensin-converting enzyme (ACE) studies revealed that unfractionated ACT (IC50 = 3.21 mg mL-1) and FCT (IC50 = 3.32 mg mL-1) were more active inhibitors of ACE in comparison to their ultrafiltrated fractions with IC50 values ranging from 6.09 to 7.45 mg mL-1. Molecular docking analysis predicted three unique peptides from ACT (AIPPGIPY, SVPGCT, and QGAGG) and FCT (AIPINNPGKL, SGNQGP, and PPGIP) as potential ACE inhibitors. Thus, unique BPs with ACE inhibitory effects might be generated from ultrasonicated lupin protein.
  7. Fulltext In-Silico Analysis and Antidiabetic Effect of α-Amylase and α-Glucosidase Inhibitory Peptides from Lupin Protein Hydrolysate: Enzyme-Peptide Interaction Study Using Molecular Docking Approach
    Fadimu GJ, Farahnaky A, Gill H, Olalere OA, Gan CY, Truong T
    Foods, 2022 Oct 26;11(21).
    PMID: 36359988 DOI: 10.3390/foods11213375
    The use of natural ingredients for managing diabetes is becoming more popular in recent times due to the several adverse effects associated with synthetic antidiabetic medications. In this study, we investigated the in vitro antidiabetic potential (through inhibition of α-glucosidase (AG) and α-amylase (AA)) of hydrolysates from lupin proteins pretreated with ultrasound and hydrolyzed using alcalase (ACT) and flavourzyme (FCT). We further fractionated ACT and FCT into three molecular weight fractions. Unfractionated ACT and FCT showed significantly (p < 0.05) higher AG (IC50 value = 1.65 mg/mL and 1.91 mg/mL) and AA (IC50 value = 1.66 mg/mL and 1.98 mg/mL) inhibitory activities than their ultrafiltrated fractions, where lower IC50 values indicate higher inhibitory activities. Then, ACT and FCT were subjected to peptide sequencing using LC-MS-QTOF to identify the potential AG and AA inhibitors. Molecular docking was performed on peptides with the highest number of hotspots and PeptideRanker score to study their interactions with AG and AA enzymes. Among the peptides identified, SPRRF, FE, and RR were predicted to be the most active peptides against AG, while AA inhibitors were predicted to be RPR, PPGIP, and LRP. Overall, hydrolysates prepared from lupin proteins using alcalase and flavourzyme may be useful in formulating functional food for managing diabetics.
  8. Camel Milk-Derived Bioactive Peptides and Diabetes: Molecular View and Perspectives
    Ayoub MA, Yap PG, Mudgil P, Khan FB, Anwar I, Muhammad K, et al.
    J Dairy Sci, 2023 Sep 12.
    PMID: 37709024 DOI: 10.3168/jds.2023-23733
    In dairy science, camel milk (CM) constitutes a center of interest for scientists due to its known beneficial impact on diabetes as demonstrated in many in vitro, in vivo, and clinical studies and trials. Overall, CM had positive effects on various parameters related to glucose transport and metabolism as well as the structural and functional properties of the pancreatic β-cells and insulin secretion. Thus, CM consumption may help manage diabetes, however, such a recommendation will become rationale and clinically conceivable only if the exact molecular mechanisms and pathways involved at the cellular levels are well understood. Moreover, the application of CM as an alternative antidiabetic tool may first require the identification of the exact bioactive molecule(s) behind such antidiabetic properties. In this review, we describe the advances in our knowledge of the molecular mechanisms reported to be involved in the beneficial effects of CM in managing diabetes using different in vitro and in vivo models. This mainly includes the effects of CM on the different molecular pathways controlling (i) insulin receptor signaling and glucose uptake, (ii) the pancreatic β-cell structure and function, and (iii) the activity of key metabolic enzymes in glucose metabolism. Moreover, we described the current status of the identification of CM-derived bioactive peptides and their structure-activity relationship study and characterization in the context of molecular markers related to diabetes. Such an overview will not only enrich our scientific knowledge of the plausible mode of action of CM in diabetes but should ultimately rationalize the claim of the potential application of CM against diabetes. This will pave the way toward new directions and ideas for developing a new generation of antidiabetic products taking benefits from the chemical composition of CM.
  9. Fulltext Hydrogen Sulphide Treatment Prevents Renal Ischemia-Reperfusion Injury by Inhibiting the Expression of ICAM-1 and NF-kB Concentration in Normotensive and Hypertensive Rats
    Hashmi SF, Rathore HA, Sattar MA, Johns EJ, Gan CY, Chia TY, et al.
    Biomolecules, 2021 Oct 19;11(10).
    PMID: 34680182 DOI: 10.3390/biom11101549
    Our main objective was to investigate the effect of chronic administration of hydrogen sulphide donor (sodium hydrosulphide) on the expression of intercellular adhesion molecule-1 (ICAM-1) and concentration of nuclear factor-kappa B (NF-kB) in a renal ischemia-reperfusion injury (IRI) model of WKY and L-nitro-arginine-methyl-ester (L-NAME)-induced hypertensive rats. Sodium hydrosulphide (NaHS) was administered intraperitoneally (i.p.) for 35 days while cystathionine gamma lyase (CSE) inhibitor dL-propargylglycine (PAG) was administered at a single dose of 50 mg/kg. Animals were anesthetised using sodium pentobarbitone (60 mg/kg) and then prepared to induce renal ischemia by clamping the left renal artery for 30 min followed by 3 h of reperfusion. Pre-treatment with NaHS improved the renal functional parameters in both WKY and L-NAME-induced hypertensive rats along with reduction of blood pressure in hypertensive groups. Oxidative stress markers like malondialdehyde (MDA), total superoxide dismutase (T-SOD) and glutathione (GSH) were also improved by NaHS treatment following renal IRI. Levels of ICAM-1 and NF-kB concentration were reduced by chronic treatment with NaHS and increased by PAG administration after renal IRI in plasma and kidney. Treatment with NaHS improved tubular morphology and glomerulus hypertrophy. Pre-treatment with NaHS reduced the degree of renal IRI by potentiating its antioxidant and anti-inflammatory mechanism, as evidenced by decreased NF-kB concentration and downregulation of ICAM-1 expression.
  10. Grandilodines A-C, biologically active indole alkaloids from Kopsia grandifolia
    Yap WS, Gan CY, Low YY, Choo YM, Etoh T, Hayashi M, et al.
    J Nat Prod, 2011 May 27;74(5):1309-12.
    PMID: 21428274 DOI: 10.1021/np200008g
    Three new indole alkaloids (1-3), named grandilodines A-C, and five known ones were obtained from the Malayan Kopsia grandifolia. The structures were established using NMR and MS analyses and, in the case of 1 and 2, were confirmed by X-ray diffraction analyses. Alkaloids 1, 3, and lapidilectine B (8) were found to reverse multidrug resistance in vincristine-resistant KB cells.
  11. Changes in blood-brain barrier permeability and ultrastructure, and protein expression in a rat model of cerebral hypoperfusion
    Sekaran H, Gan CY, A Latiff A, Harvey TM, Mohd Nazri L, Hanapi NA, et al.
    Brain Res Bull, 2019 10;152:63-73.
    PMID: 31301381 DOI: 10.1016/j.brainresbull.2019.07.010
    Cerebral hypoperfusion involved a reduction in cerebral blood flow, leading to neuronal dysfunction, microglial activation and white matter degeneration. The effects on the blood-brain barrier (BBB) however, have not been well-documented. Here, two-vessel occlusion model was adopted to mimic the condition of cerebral hypoperfusion in Sprague-Dawley rats. The BBB permeability to high and low molecular weight exogenous tracers i.e. Evans blue dye and sodium fluorescein respectively, showed marked extravasation of the Evans blue dye in the frontal cortex, posterior cortex and thalamus-midbrain at day 1 following induction of cerebral hypoperfusion. Transmission electron microscopy revealed brain endothelial cell and astrocyte damages including increased pinocytotic vesicles and formation of membrane invaginations in the endothelial cells, and swelling of the astrocytes' end-feet. Investigation on brain microvessel protein expressions using two-dimensional (2D) gel electrophoresis coupled with LC-MS/MS showed that proteins involved in mitochondrial energy metabolism, transcription regulation, cytoskeleton maintenance and signaling pathways were differently expressed. The expression of aconitate hydratase, heterogeneous nuclear ribonucleoprotein, enoyl Co-A hydratase and beta-synuclein were downregulated, while the opposite observed for calreticulin and enhancer of rudimentary homolog. These findings provide insights into the BBB molecular responses to cerebral hypoperfusion, which may assist development of future therapeutic strategies.
  12. Smartphone-controlled patch electro-acupuncture versus conventional pain relief during colonoscopy: a randomized controlled trial
    Joan Gan CY, Chan KK, Tan JH, Tan Chor Lip H, Louis Ling LL, Mohd Azman ZA
    ANZ J Surg, 2021 06;91(6):E375-E381.
    PMID: 33876547 DOI: 10.1111/ans.16870
    BACKGROUND: Smartphone-controlled patch electro-acupuncture (SCEA) is a novel device which gives the same analgesic effect as with conventional acupuncture. There are no published articles in the English literature on the use of this device as a primary mode of pain relief during colonoscopy. Primary aims of this study were to investigate the efficacy of SCEA as a substitute for pain relief during colonoscopy.

    METHODS: Thirty-seven patients were randomized to receive SCEA (n = 19) or placebo (n = 18) during colonoscopy. Additional rescue sedation was administered to patients if they had pain or discomfort during the procedure. Visual analogue scale was used to quantify the intensity of pain from the beginning to end of the procedure. Other variables analysed were the amount of sedation used, duration from start to caecal intubation, length of time for completion of colonoscopy and recovery time to home discharge.

    RESULTS: Patients who received SCEA had a lower median pain score of 4.6 (interquartile range 5.7) compared to the placebo group of 6.0 (interquartile range 3.2). Statistical analysis comparing the groups revealed a non-significant P-value of 0.12, although more than 90% of the patients indicated willingness for SCEA as the primary analgesia if they were to repeat the procedure. Throughout the study, there were no adverse complications that occurred during the use of SCEA.

    CONCLUSIONS: Even though this study did not demonstrate, a significance in pain reduction, SCEA remains a safe modality which, more than 90% of patients favoured as a substitute for pain relief during colonoscopy.

  13. A comparative investigation into novel cholesterol esterase and pancreatic lipase inhibitory peptides from cow and camel casein hydrolysates generated upon enzymatic hydrolysis and in-vitro digestion
    Mudgil P, Baba WN, Kamal H, FitzGerald RJ, Hassan HM, Ayoub MA, et al.
    Food Chem, 2022 Jan 15;367:130661.
    PMID: 34348197 DOI: 10.1016/j.foodchem.2021.130661
    Cow (CwC) and camel casein (CaC) hydrolysates were generated using Alcalase™ (CwCA and CaCA) and Pronase-E (CwCP and CaCP) each for 3 and 6 h, and investigated for their potential to inhibit key lipid digesting enzymes i.e., pancreatic lipase (PL) and cholesteryl esterase (CE). Results revealed stronger PL and CE inhibition by CaC hydrolysates compared to CwC. Potent hydrolysates (CwCP-3 h and CaCA-6 h) upon simulated gastrointestinal digestion (SGID) showed significant improvement in inhibition of both PL and CE. However, both the SGID hydrolysates showed similar extent of PL and CE inhibition and were further sequenced for peptide identification. Peptides MMML, FDML, HLPGRG from CwC and AAGF, MSNYF, FLWPEYGAL from CaC hydrolysates were predicted to be most active PL inhibitory peptides. Peptide LP found in both CwC and CaC hydrolysates was predicted as active CE inhibitor. Thus, CwC and CaC could be potential source of peptides with promising CE and PL inhibitory properties.
  14. Microwave-assisted extraction of polysaccharide from Cinnamomum cassia with anti-hyperpigmentation properties: Optimization and characterization studies
    Al-Ajalein AAS, Shafie MH, Yap PG, Kassim MA, Naharudin I, Wong TW, et al.
    Int J Biol Macromol, 2023 Jan 31;226:321-335.
    PMID: 36502951 DOI: 10.1016/j.ijbiomac.2022.12.023
    The anti-hyperpigmentation effect and tyrosinase inhibitory mechanism of cinnamon polysaccharides have not been reported. The current study focused on the extraction of polysaccharides from Cinnamomum cassia bark using microwave-assisted approach and optimization of the extraction process (i.e., microwave power, irradiation time and buffer-to-sample ratio) by Box-Behnken design to obtain a high yield of polysaccharides with high sun protection factor (SPF), anti-hyperpigmentation and antioxidant activities. The extracted pectic-polysaccharides had low molecular weight and degree of esterification. The optimal extraction process had polysaccharides characterized by (a) monophenolase inhibitory activity = 97.5 %; (b) diphenolase inhibitory activity = 99.4 %; (c) ferric reducing antioxidant power = 4.4 mM; (d) SPF = 6.1; (e) yield = 13.7 %. The SPF, tyrosinase inhibitory and antioxidant activities were primarily contributed by the polysaccharides. In conclusion, the polysaccharides from C. cassia could be an alternative therapeutic source for skin hyperpigmentation treatment.
  15. Fulltext Molecular docking studies on α-amylase inhibitory peptides from milk of different farm animals
    Mudgil P, Al Dhaheri MKO, Alsubousi MSM, Khan H, Redha AA, Yap PG, et al.
    J Dairy Sci, 2024 May;107(5):2633-2652.
    PMID: 38101739 DOI: 10.3168/jds.2023-24118
    Milk-derived peptides have emerged as a popular mean to manage various lifestyle disorders such as diabetes. Fermentation is being explored as one of the faster and efficient way of producing peptides with antidiabetic potential. Therefore, in this study, an attempt was made to comparatively investigate the pancreatic α-amylase (PAA) inhibitory properties of peptides derived from milk of different farm animals through probiotic fermentation. Peptide's identification was carried out using liquid chromatography-quadrupole time-of-flight mass spectrometry and inhibition mechanisms were characterized by molecular docking. Results obtained showed a PAA-IC50 value (the amount of protein equivalent needed to inhibit 50% of enzymes) between 2.39 and 36.1 µg protein equivalent for different fermented samples. Overall, Pediococcus pentosaceus MF000957-derived fermented milk from all animals indicated higher PAA inhibition than other probiotic derived fermented milk (PAA-IC50 values of 6.01, 3.53, 15.6, and 10.8 µg protein equivalent for bovine, camel, goat, and sheep fermented milk). Further, molecular docking analysis indicated that camel milk-derived peptide IMEQQQTEDEQQDK and goat milk-derived peptide DQHQKAMKPWTQPK were the most potent PAA inhibitory peptides. Overall, the study concluded that fermentation derived peptides may prove useful in for managing diabetes via inhibition of carbohydrate digesting enzyme PAA.
  16. Biochemical and radical-scavenging properties of sea cucumber (Stichopus vastus) collagen hydrolysates
    Abedin MZ, Karim AA, Latiff AA, Gan CY, Ghazali FC, Barzideh Z, et al.
    Nat Prod Res, 2014;28(16):1302-5.
    PMID: 24670209 DOI: 10.1080/14786419.2014.900617
    The molecular mass distribution, amino acid composition and radical-scavenging activity of collagen hydrolysates prepared from collagen isolated from the sea cucumber Stichopus vastus were investigated. β and α1 chains of the collagen were successfully hydrolysed by trypsin. The molecular mass distribution of the hydrolysates ranged from 5 to 25 kDa, and they were rich in glycine, alanine, glutamate, proline and hydroxyproline residues. The hydrolysates exhibited excellent radical-scavenging activity. These results indicate that collagen hydrolysates from S. vastus can be used as a functional ingredient in food and nutraceutical products.
  17. Profiling of morphine and codeine in urine after the ingestion of curry containing poppy seed as an evidence for opiates defence in Malaysia
    Gan CY, Zainuddin Z, Muhamad Noh H, Rahmat R, Mohd Akir F, Mahad NH, et al.
    Forensic Sci Int, 2019 Sep;302:109865.
    PMID: 31279278 DOI: 10.1016/j.forsciint.2019.06.023
    Consumption of curry containing poppy seeds has raised an issue concerning the opiate content in the urine that might exceed the cut-off value (300ng/mL). The main objective of this study was to examine the morphine and codeine contents in the urine of the consumers after partaking poppy seed-enriched curry in. The volunteers were asked to partake: (a) a single meal and their urines were collected within 24h, or (b) Two meals a day for three consecutive days and their urines were collected within 72h. Two different dosages were also tested in this study: (a) low dosage: 1g/100ml curry (containing 138μg of morphine and 66μg of codeine) and (b) high dosage: 5g/100ml curry (containing 690μg of morphine and 330μg of codeine). The subjects were randomised into the groups using the method of stratified randomization with age and gender groups as covariates. A total of 6 subjects was allocated for each group and placebos were used as control. Results showed that all subjects who consumed low dosage of poppy seeds either in single meal or multiple meals experiment were found negative. However, 1 out of 6high dosage subjects was confirmed positive at a period of 3-6h after the consumption of curry in the single meal study. This outlier maybe due to the lack of water consumption after consuming the curry, thus the low volume of urine was collected and the opiate was concentrated in the urine. On the other hand, 5 out of 6high dosage subjects in the multiple meals experiment were found positive. Majority of these subjects were found positive on the second and third day of the experiment after the second curry meal was consumed. The outlier (negative) in this group might be due to the high consumption of water throughout the experiment and the subject's urine volumes and frequency of urine collection were much higher compared to other subjects. From the result of this study, it can be concluded that partaking high dosages of poppy seed in curry could give a positive response (>300ng/ml+uncertainty of measurement) in the urine, and the water consumption after partaking curry has significant influence for the opiate contents in the urine.
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