A single colony of bacteria was inoculated overnight (16 hours) in 50 ml of LB medium. Equal OD of cells from OP50 and OP50PQE was centrifuged at 4400 rpm for 15 min. The harvested pellets were the washed once with will Milli-Q water and then three times with 1 mM EDTA, followed by one wash with Milli-Q water (62 (link)). Total cellular iron content was then measured as described previously (63 (link)). Briefly, bacterial pellets were treated with total wet weight nitric acid (2 ml/g; trace metal grade, Thermo Fisher Scientific) for 24 hours and then digested with total wet weight hydrogen peroxide (1 ml/g; trace metal grade, Thermo Fisher Scientific) for 24 hours at room temperature. The samples were preserved at 4°C until quantification of metals. Ultrapure water (VWR Chemicals, ARISTAR ULTRA) was used for final sample dilution. Samples were then analyzed using ICP-MS (PerkinElmer, Nexion 2000) using 50–part per billion Bismuth as internal standard.
Trace metal grade
Manufactured by Thermo Fisher Scientific
Sourced in United States, United Kingdom, India
Trace Metal Grade is a high-purity chemical reagent designed for trace metal analysis. It is formulated to have extremely low levels of impurities, making it suitable for applications that require the detection and quantification of trace metals in various matrices.
Automatically generated - may contain errors
Lab products found in correlation
Nexion 2000, by PerkinElmer (4 mentions)
Aristar ultra, by Avantor (3 mentions)
7900 icp ms instrument, by Agilent Technologies (2 mentions)
X series 2, by Thermo Fisher Scientific (2 mentions)
Nitric acid, by Merck Group (2 mentions)
Hydrochloric acid (hcl), by Thermo Fisher Scientific (2 mentions)
Icpe 9000, by Shimadzu (2 mentions)
Titan mps, by PerkinElmer (2 mentions)
Urea, by Merck Group (2 mentions)
Potassium chloride (kcl), by Merck Group (2 mentions)
Icpms 2030, by Shimadzu (1 mentions)
Suprapur, by Merck Group (1 mentions)
Agilent 7700, by Agilent Technologies (1 mentions)
Emx spectrometer, by Bruker (1 mentions)
Certiprep, by SPEX (1 mentions)
Mars 6, by CEM Corporation (1 mentions)
Nexion 300 icp ms system, by PerkinElmer (1 mentions)
Aanalyst 600, by PerkinElmer (1 mentions)
Nafion 1100 w, by Merck Group (1 mentions)
Ampicillin sodium salt, by Merck Group (1 mentions)
37 protocols using trace metal grade
1
Quantification of Bacterial Iron Content
2
Quantifying Metal Oxide Nanoparticle Stability
3
Trace Element Quantification by ICP-MS
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High purity HNO3 (≥70% w/w, TRACEMETAL™ Grade, Fisher Scientific, Waltham, MA, USA) and HCl (≥38% w/w TRACEMETAL™ Grade, Fisher Scientific, Waltham, MA, USA) were used as received. The internal standard solution was prepared by appropriate dilution of the Internal Standard Mix solution (Agilent Technologies, Santa Clara, CA, USA). Calibration standards were prepared from 10 µg/mL and 100 µg/mL custom mix multi-element ICP-MS standard (HPS, North Charleston, SC, USA), and from the 1000 µg/mL mineral element ICP-MS standard solution (ICP-AM-17 Mineral Calibration Standard, HPS, North Charleston, SC, USA). All solutions were prepared by using ultrapure water (>18.2 MΩ·cm at 25 °C) from a Mili-Q IQ-7000 (Merck KGaA, Darmstadt, Germany) water purification system. All the glassware and plastic materials were decontaminated with a 10% (v/v) HNO3 solution for 48 h and rinsed with ultrapure water prior to use.
4
Analytical Standards for Food Composition
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Authentic standards of phenolic acids (GA, 4--BA, CA, TFA, p-CA, VA, and SA), and flavonoids (API7G, HES, kaempferol (KAM), luteolin (LUT), myricetin (MYR), naringenin (NA), quercetin (QE), and RUT) were procured from Sigma-Aldrich, India. The rice flour reference material (NIST1568b) and ammonium formate were also purchased from Sigma-Aldrich, India. Disodium methyl arsonate hexahydrate and dimethylarsinic acid were obtained from Chem Service inc, West Chester, PA, USA. Phenolic acids, CGA and PA, were purchased from HWI Analytik, GmbH, Germany. Apigenin (API) was obtained from Fluka (India). Ultrapure water (resistivity 18.2 M Ω-cm) was obtained from an ultra-pure Lab Q water purification system (Indion Lab Q, India) and used throughout the experiment. Nitric acid (65%–69%, TraceMetal™ Grade) and hydrochloric acid (35%–37%, TraceMetal™ Grade) were purchased from Fisher, India. Standards of arsenic species, As (III) and As (V) were purchased from Supelco, India. Sodium arsenate, potassium dihydrogen phosphate, urea, and potassium chloride were procured from Merck, India. All other chemicals used were of analytical grade.
5
Serum Metal Quantification by ICP-MS
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Whole blood from indicated mice were obtained via submandibular vein puncture or from the orbital sinus. Whole blood was collected in untreated sterile 1.5 mL Eppendorf tubes and allowed to coagulate for 1–2 hrs at RT. Coagulated samples were spun at 13,000×g for 15 min and serum was collected. Collected serum was further clarified with a second spin at 13,000×g for 15min. 10 μL of clarified serum was treated with 2 mL/g total wet weight nitric acid (20 μL) (Trace metal grade; Fisher) for 24 hr, and then digested with 1 mL/g total wet weight hydrogen peroxide (10 μL) (Trace metal grade; Fisher) for 24 h at room temperature. The samples were preserved at 4 °C until quantification of metals. Ultrapure water (VWR Chemicals ARISTAR®ULTRA) was used for final sample dilution to 3 mL. Samples were then analyzed using inductively coupled plasma mass spectrometry (ICP-MS) (Perkin Elmer) utilizing Bismuth as an internal standard.
6
Quantifying Cellular Metal Uptake
7
Inductively Coupled Plasma Mass Spectrometry for Metal Analysis
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Tissue samples were analyzed for metals by inductively coupled plasma mass spectrometry (Perkin Elmer Nexion 2000). The internal standard was 50 ppb Bismuth. Briefly, the tissues were digested with 2 mL/g total wet weight nitric acid (Trace metal grade; Fisher) for 24 h, and then digested with 1 mL/g total wet weight hydrogen peroxide (Trace metal grade; Fisher) for 24 h at room temperature. The samples were preserved at 4 °C until quantification of metals. Ultrapure water (VWR Chemicals ARISTAR®ULTRA) was used for final sample dilution.
8
Quantitative Iron Analysis by AAS
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The iron content of the stock solutions was verified by AAS. Briefly, each stock solution (1 mL) was mixed with ultrapure water (3–4 mL), concentrated nitric acid (0.75 mL; trace metal grade; Fisher Scientific, Loughborrough, U.K.), and concentrated hydrochloric acid (0.15 mL; trace metal grade; Fisher Scientific, Loughborrough, U.K.) in Teflon microwave digestion vessels. The microwave digestion was then carried out in an Ethos plus microwave lab station (Milestone; Sorisole, Italy) at 180 °C for 15 min. Iron concentration in the microwave-digested samples was determined using an Agilent 240/280 Series AA spectrometer (Santa Clara, CA, USA).
9
Trace Metal Analysis of Spirits
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To prepare samples for ICP-MS analysis, 20 mL of each spirit was mixed with 0.4 mL of 50% nitric acid and 0.2 mL of 50% hydrochloric acid (TraceMetal Grade, Thermo Fisher Scientific, Waltham, MA, USA) in a digestion tube and refluxed on a heating block (Environmental Express, Charleston, SC, USA) at 93 °C for 2 h. The digested samples were cooled, filtered (No. 40 Whatman filter paper, Thermo Fisher Scientific, Waltham, MA, USA), and adjusted with distilled/deionized water back to 20 mL for analysis using a Thermo Scientific X-Series 2 ICP-MS (Thermo Electron North America LLC).
To ensure that Falcon tubes into which samples were transferred were not themselves introducing extractable metals into samples, two fresh tubes were filled with a 50/50 (v/v) mixture of 100% ethanol/distilled water and incubated for three months, the contents of which were analyzed as regular samples. A well characterized Scotch whisky (The Glenlivet (TGL); Adam et al., 2002 ) purchased in San Diego, California (USA) was also included as a ‘certified reference material.’ To ensure control over the reference conditions under which trace amounts of metals could be detected with 99% confidence in ethanol, a 50/50 (v/v) mixture of 100% ethanol/distilled water was used as a blank sample in each batched analysis, and as a platform for laboratory control spiked duplicate samples.
To ensure that Falcon tubes into which samples were transferred were not themselves introducing extractable metals into samples, two fresh tubes were filled with a 50/50 (v/v) mixture of 100% ethanol/distilled water and incubated for three months, the contents of which were analyzed as regular samples. A well characterized Scotch whisky (The Glenlivet (TGL); Adam et al., 2002 ) purchased in San Diego, California (USA) was also included as a ‘certified reference material.’ To ensure control over the reference conditions under which trace amounts of metals could be detected with 99% confidence in ethanol, a 50/50 (v/v) mixture of 100% ethanol/distilled water was used as a blank sample in each batched analysis, and as a platform for laboratory control spiked duplicate samples.
10
Preparation and Analysis of Serum Samples for ICP-MS
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Three days before the blood collection, all study participant maintained normal diet and exercise levels. At least 2 ml of venous blood was collected on an empty stomach, and the serum was isolated within 3 h (3500 rpm, 35 min), and frozen samples were then used for inspection. Samples with visible hemolysis, lipids, or jaundice were excluded from the analysis. One milliliter 5% HNO3 (Trace Metal Grade, Thermo Fisher, USA) solution was added to 0.5 ml serum sample to obtain a mixed solution, which was centrifuged at 12000 rpm for 5 min to obtain the supernatant. After that, 3.5 ml 1% HNO3 solution was added to 0.5 ml supernatant, and then the mixture was centrifuged for 2 min at 2500 rpm, and incubated at room temperature for 1 min to obtain the samples to be analyzed by ICP-MS (ICPMS-2030, Shimadzu, Japan). During the determination, accuracy and precision were checked by certified commercial element standard solution (1000 mg/L, the National Center of Analysis and Testing for Nonferrous Metals and Electronic Materials, Beijing, China).
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