Ann Lab Med.  2021 Jul;41(4):414-418. 10.3343/alm.2021.41.4.414.

A Stabilizing Agent, PCA/DTPA, Improves Plasma Storage Life for the Chromsystems Vitamin C Assay up to Six Months

• Collie JT
• Hudson EP
• Deane AM
• Bellomo R
• Greaves RF

• KMID: 2512722
• DOI: http://doi.org/10.3343/alm.2021.41.4.414

Abstract

The commonly used Chromsystems vitamin C (ascorbate) assay (Munich, Germany) has a sample storage life of five days at –20°C. Stabilizing agents have been successfully used to increase longevity; however, their suitability with this commercial assay is unclear. We investigated the compatibility of a stabilizing agent, perchloric acid/diethylenetriaminepentaacetic acid (PCA/DTPA), with the Chromsystems assay. Plasma was stored at –80°C, with or without PCA/DTPA. Storage up to six months was assessed through baseline and repeat analyses, stability was assessed by comparing paired non-stabilized and PCA/ DTPA-stabilized plasma, and performance was assessed using allowable performance specifications of an external quality assurance program. Ascorbate concentration was significantly lower in non-stabilized plasma than in paired PCA/DTPA-stabilized plasma, with a proportional difference of 11% (P = 0.01). All storage analysis results were within the allowable performance specifications. Storage at –80°C prevented plasma ascorbate oxidation; however, substantial oxidation occurred during sample processing. In conclusion, PCA/DTPA significantly reduces ascorbate oxidation, and PCA/DTPA-stabilized ascorbate plasma is compatible with the Chromsystems assay and stable for up to six months, when stored at –80°C.

Keyword

Ascorbate; Vitamin C; Storage; Stability; Chromsystems; PCA/DTPA

Figure

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  Fig. 1 Schematic representation of the plasma sample handling and processing procedures utilized in this study. (A) Immediate storage of non-stabilized plasma at –80°C, followed by HPLC-UV analysis. (B) Addition of the PCA/DTPA stabilizing agent to the plasma aliquot (1:1, v:v) and storage at –80°C, followed by HPLC-UV analysis. Abbreviations: PCA, perchloric acid; DTPA, diethylenetriaminepentaacetic acid; HPLC-UV, high performance liquid chromatography with ultra-violet detection.

• 

  Fig. 2 Difference plots and Passing–Bablok regression of storage and stability experiments. The data displayed in panels A, B, and C are results from the T1 analyses for each plasma sample and are expressed as a percentage change in concentration from the baseline, T0, on the Y-axis. The X-axis represents weeks in storage before the T1 analysis. (A) Storage of non-stabilized plasma. Mean percentage concentration difference of +1.2% (median+0.5%). (B) Storage of PCA/DTPA-stabilized plasma. Mean percentage concentration difference of –0.2% (median+1.5%). (C) Comparison of PCA/DTPA-stabilized plasma with paired non-stabilized plasma. Numbers of samples per time point analyzed: 2 weeks (N=4), 7 weeks (N=7), 15 weeks (N=14), and 26 weeks (N=14). Mean percentage concentration difference of +9.9% (median+3.6%). (D) Passing–Bablok regression of non-stabilized plasma against PCA/DTPA-stabilized plasma. Concentrations determined through all experiments ranged from 2–812 μmol/L. The dashed lines in panels A, B, and C represent the ±25% APS for results ≥36.0 μmol/L. Abbreviations: PCA, perchloric acid; DTPA, diethylenetriaminepentaacetic acid; APS, allowable performance specifications; CI, confidence interval.

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