Nexion 350x

Urine cadmium measurement by inductively coupled plasma mass spectrometry. Urine specimens were collected and stored in 10 mL metal-free plastic collection tubes. To avoid hydration bias during urine sample collection, urine samples that were overdiluted or overconcentrated (urine creatinine level <10 or >300 mg/dL) were excluded from analysis. Urine specimens were stored at 4 °C. Cadmium was quantified by means of inductively coupled plasma mass spectrometry on a PerkinElmer NexION 350X instrument (MA, USA) and analyzed using a no-gas mode. Urine specimens (500 μL) were diluted (1 + 9) with a 1.5% nitric acid (JT Baker, NJ, USA) solution containing yttrium as an internal standard. The cadmium and yttrium standards were purchased from AccuStandard (CT, USA). The standard range was 1.25 to 40 μg/L. The calibration curve had an R ≥ 0.995. Lypocheck quality controls and urine metal levels 1 and 2 (Bio-Rad Laboratories, Hemel Hempstead, UK) were used and analyzed at the start and end of each analytical run and again after every 10 samples. The lower limit of quantitation (LOQ) for cadmium was 0.3 μg/L. Values below the LOQ were assigned to LOQ for analysis.

Quantification of platinum (Pt) in blood (pharmacokinetic analysis) as well as in tumor samples and organ tissues (biodistribution analysis) was performed by Excite Pharma Services LLC (Lee’s Summit, MO, USA) in their FDA inspected facility which adheres to Good Manufacturing Practices (GMP) and Good Laboratory Practices (GLP). The analysis was performed using Perkin Elmer Nexion 350X inductively coupled plasma mass spectrometry (ICP-MS) instrument (Waltham, MA, USA) and a validated (ICP-MS) method^{34 (link)–37 (link)}. Lower limit of quantitation [LLOQ] = 1.06 ng/mL.

Root tissues (0.3 g) of four-week-old plants were used to isolate intact protoplasts and vacuoles according to Robert et al. (2007) (link). The purified protoplasts were divided into two fractions, one of which was used for the releases of vacuoles according to the method described in Dürr et al. (1975) (link). The purified protoplasts and vacuoles were used for the determination of NO₃^- and Cd²⁺ concentrations (Vögeli-Lange and Wagner, 1990 (link); Han et al., 2016 (link)). NO₃^- concentrations in protoplasts and vacuoles were measured by a continuous-flow auto-analyzer (Auto Analyzer 3, Bran and Luebbe) as described previously by Han et al. (2016) (link). Cd²⁺ concentrations in the protoplasts and vacuole were determined with an ICP Mass Spectrometer (NexION 350X, PerkinElmer) as described in Huang et al. (2012) (link).

After all experiments were complete, blood, urine, and several types of tissue (kidney, blood, liver, bone, fat, and muscle) were collected. Concentrations of chromium in samples were measured according to a prior report [40 (link)]. In brief, each sample (0.1 g of tissue and 25 µL of blood and urine) was digested in 65% nitric acid, and then each sample was subjected to digestion with nitric acid overnight (temperature: 100 °C). Chromium concentrations were determined using an ICP Mass Spectrometer (NexION 350X, PerkinElmer, MA, USA). Next, distilled water was used to dilute the digested solution to a 5 mL solution prior to measurements. The relative chromium recovery rate was calculated at 10 ppb and 100 ppb of the quantification levels by 5% (n = 5) and 8% (n = 5), respectively. The absorption data were plotted onto a 1–500 ppb standard curve, and regression analysis was performed to identify the total chromium level in samples (R² > 0.996).

Human sera (0.2 g) were added to 5 µL of IS solution (10 µg/mL Rh) and 1 mL of nitric acid in 15 mL centrifuge tubes (polypropylene conical tube, Falcon, Tamaulipas, Mexico). The samples were transferred to a microwave digestion system (MARS 6, CEM, Matthews, NC, USA), which was equipped with a fiber optic temperature sensor and a high-throughput accessory (CEM High Throughput Vessel set, DV-120 turntable, Matthews, NC, USA). The temperature was ramped up for 30 min and held at 100 °C for 20 min with the oven power set at 750 W. After this process, the samples were cooled to room temperature, and the final volume was turned up to 5 mL with ultrapure water. The samples were centrifuged at 4000 rpm for 5 min, and the supernatant was introduced directly into an inductively coupled plasma mass spectrometry (ICP-MS, PerkinElmer NexIon 350X, Waltham, MA, USA) system to analyze the elements present. In addition, 0.2 g of human sera was added to the IS solution, and the 5-element solution was added to the spiked sample. The final concentrations were 15 ppb for copper and Zn, and 3 ppb for Cr, Mn, and Se in the instrumental analysis. Spike samples were used to calculate the recovery and confirm the accuracy of this method.