Development of Methylmercury Analysis by Ultra-High Performance Liquid

Overview

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Key numbers

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DOI: 10.22146/ijc.82512 C18 Hypersil Gold analytical column with a mobile gradient phase of 0.5% (w/v) L-
cysteine in 2% (v/v) HNO3 and 100% methanol. The MeHg was extracted from 100 mg of
shark meat using 1 mL of 0.5% (w/v) L-cysteine in 2% (v/v) HNO3 and sonicated for
acutus was 0.22–0.63 mg/kg wet weight (w.w.), Squalus hemipinis 0.68–1.14 mg/kg w.w.,
and 0.29–1.22 mg/kg w.w. for Sphyrna lewini. This study provides a quick and easy
predominant chemical form, accounting for 80–90% of
Mercury concentrations of up to 4,000 mg/kg and MeHg
performance chromatography combined with ICP-MS mobile phase used was 0.5% L-cysteine in 2% HNO3 (A)
with a fast procedure and less waste-containing MeHg. and methanol 100% (B), and the flow rate of the gradient
adjusted to 2% and gradually increased to 90% in 132 s.
Materials After that, hold at 90% for 20 s and continue at the initial
water (Thermo Fisher, USA), HNO3 65% (Merck), at 40 °C, and the volume of injection was 50 μL. The
DORM-4 as CRM for MeHg and L-cysteine (99% purity
Sigma, USA). All glassware was washed by soaking in 10%
1,000 mg/L standard solution for CH3HgCl was prepared September 11-16, 2019, from Binuangeun Fish Auction,
by weight 1 mg CH3HgCl diluted in 1 mL with 0.5% L- Lebak, Banten, Indonesia (Southern Java Sea areas).
cysteine 2% HNO3. A series of standard solutions (1, 5, 10, Sixteen sharks were identified as Rhizoprionodon acutus
25, 50, 75, 100 μg/L) were prepared by dilution of (RA), six as Squalus hemipinis (SH), and eight as
1,000 mg/L MeHg stock solution with 0.5% L-cysteine 2% Sphyrna lewini. The total weight (TW) was measured
Fig 1. MeHg concentration calibration curve (μg/L) in duplicates with R2 = 0.999, slope = 52.336.311,
Table 1. Liquid chromatography and ICPMS operating condition for mercury speciation (MeHg)
Mobile phase A: 0.5% L-cysteine in 2% HNO3
oven-dried for 24 h at 60 °C for MeHg analysis and with 0.5% L-cysteine in 2% HNO3. Two replicates of
Dried samples were ground using mortar and pestle to get mercury) in the range of 1–100 μg/L were used to form
based on de Souza et al. (9) with modifications for the were measured based on (14) with the equation; LOD
phase, and elution conditions. The use of L-cysteine in measurement of blank/pseudo blank (n = 10), Sb was the
acid conditions as the solvent and mobile phase and also slope of the calibration curve, and the LOQ was 3.33 of
of 0.5% L cysteine in 2% HNO3 and applied sonication
A CRM DORM4 (n = 10) was used as sample of MeHg in DORM4 with 1 mL of L-cysteine in acid
and Rodrigues et al. (12) used 4.75 mL solvent. Previous results can reduce 80% on the use of solvent compared
studies commonly use two compounds, such as L-cysteine with Vallant et al. (13) and reduce by almost 90%
conditions. L-cysteine was used in acid conditions as containing MeHg (Table 3). The observed limit of
method gives less solvent for extraction, less chemical The total length of milk sharks ranged from 34.5–
(9,11-13) and showed good separation with mobile phase (Table 4).
0.5% L-cysteine and 2% HNO3 and 100% methanol under The concentration of MeHg in the meat/muscle
gradient conditions of less than 300 s. Inorganic mercury tissue ranged between 0.22 and 0.63 mg/kg w.w. (mean
(InHg) was showed a peak at 196.5 s and MeHg around concentration of MeHg: 0.41 ± 0.13 mg/kg w.w.) for R.
266 s (Fig. 2). Gradient elution shows low signal noise at acutus, 0.68 to 1.14 mg/kg w.w. (0.81 ± 0.17 mg/kg w.w.)
a low concentration compared to isocratic elution (9). The Table 2. Recovery of MeHg concentration in DORM-4
MeHg analysis by comparing isocratic elution (9). L-cysteine in 2% (v/v) HNO3 using gradient elution
method shows good result measurement (Table 2) with Certified value Measurement value
values of 96.90%. Our method uses less solvent in this CRM Analyte

Methods (brief)

Chromatography Coupled with ICP-MS and Its Application on Sharks’ Meat
Accepted: March 24, 2023 shark meat samples. Effective MeHg separation was achieved in less than 300 s using a
samples in a single assay.
Keywords: UHPLC-ICP-MS; methylmercury; sonication-assisted extraction; shark meat
coupled plasma mass spectrometry (ICP-MS)).
Performance Liquid Chromatography (HPLC)
HPLC-ICPMS on biological samples. However, they all
produce large amounts of waste products containing UHPLC-ICPMS (Thermo Scientific) with PFA Cyclonic
method for determining the MeHg concentration in shark Vanquish™ UHPLC Quaternary Pump with reverse-
performance chromatography combined with ICP-MS mobile phase used was 0.5% L-cysteine in 2% HNO3 (A)
methylmercury chloride (CH3HgCl) standard (Sigma, optimal experimental conditions for UHPLC and
USA), methanol for LC grade (Merck, Germany), ICPMS are shown in Table 1.
(v/v) HNO3 for 12 h, rinsed three times with double- Sample preparation and extraction MeHg
distilled deionized water and air drying for 12 h. A Shark samples were collected by direct purchase on
Table 1. Liquid chromatography and ICPMS operating condition for mercury speciation (MeHg)
Dried samples were ground using mortar and pestle to get mercury) in the range of 1–100 μg/L were used to form
The development method of MeHg extraction was of detection (LOD) and limit of quantification (LOQ)
based on de Souza et al. (9) with modifications for the were measured based on (14) with the equation; LOD

Implications

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Update history

The five most recent substantive edits to this page, classified major (evidence or structure moved), correction (a published value or statement was wrong and has been fixed), or minor (narrative rewritten without changing the underlying evidence). Each description is derived from what the edit did to this page; the linked commit is the authoritative record, routine regeneration passes are excluded, and the full version history lives in git. When DOI minting comes online (see schema docs), each entry below will also link to a version-pinned DataCite DOI.

Commit	Date	Change	Description
b01ec52c	2026-08-04	major	2 sections added
d49e450f	2026-08-03	major	5 sections added; narrative text revised