A New Method for the Determination of Total Content of Vitamin C, Ascorbic and Dehydroascorbic Acid, in Food Products with the Voltammetric Technique with the Use of Tris(2-carboxyethyl)phosphine as a Reducing Reagent

At a Glance

Metadata	Details
Publication Date	2023-01-13
Journal	Molecules
Authors	Artur Mazurek, Marzena Włodarczyk-Stasiak
Institutions	University of Life Sciences in Lublin
Citations	13
Analysis	Full AI Review Included

Technical Documentation & Analysis: High-Sensitivity Voltammetry Using 6CCVD Boron-Doped Diamond (BDD)

Executive Summary

This research successfully developed and validated a novel differential pulse voltammetry (DPV) method for the accurate determination of total Vitamin C (Ascorbic Acid and Dehydroascorbic Acid) in complex food matrices. The core value proposition for 6CCVD clients lies in the reliance on high-performance Boron-Doped Diamond (BDD) electrodes for superior electrochemical sensing.

Core Achievement: Developed a highly sensitive DPV method for total Vitamin C determination, demonstrating equivalence to the established HPLC-DAD reference method.
Material Requirement: The method critically depends on a Boron-Doped Diamond Electrode (BDE) modified with an electrochemically deposited mercury film.
Performance Metrics: Achieved a high sensitivity with a Limit of Detection (LOD) of 0.89 µg/mL and a Limit of Quantification (LOQ) of 2.67 µg/mL.
Matrix Compatibility: BDE modification minimized interference from complex food matrix components, a common challenge in food analysis.
Chemical Methodology: Utilized Tris(2-carboxyethyl)phosphine (TCEP) as a highly effective reducing agent for Dehydroascorbic Acid (DHAA) in acidic environments (pH < 2).
Interference Mitigation: Successfully eliminated TCEP interference using N-ethylmaleimide (NEM) prior to voltammetric measurement, ensuring accurate AA quantification.
6CCVD Relevance: This work validates the use of 6CCVD’s custom BDD substrates for developing robust, high-precision electrochemical sensors in demanding analytical applications.

Technical Specifications

The following hard data points detail the critical parameters and performance metrics of the developed voltammetric method, focusing on the electrode material and measurement conditions.

Parameter	Value	Unit	Context
Electrode Material	Boron-Doped Diamond (BDD)	N/A	Working electrode
Electrode Diameter	3	mm	Standard size used in the three-electrode system
Surface Polishing	0.3	µm	Alumina slurry used for BDE preparation
Electrode Modification	Mercury (Hg) Film	N/A	Electrodeposited from 20 mg/L Hg(II) solution
Modification Potential	-1.30	V	Applied for 180 s during Hg film formation
Voltammetry Technique	Differential Pulse Voltammetry (DPV)	N/A	Measurement method for AA/Tc
Potential Range (Initial)	-50	mV	vs Ag/AgCl reference electrode
Potential Range (Final)	200	mV	vs Ag/AgCl reference electrode
Pulse Amplitude	50	mV	DPV parameter
Potential Change Rate	10	mV/s	DPV parameter
Limit of Detection (LOD)	0.89	µg/mL	Mean detection limit for AA
Limit of Quantification (LOQ)	2.67	µg/mL	Calculated as 3 * LOD
DHAA Reduction Time	30	min	Required time using 10 mM TCEP solution
TCEP Interference Elimination Time	30	min	Required time using 40 mM N-ethylmaleimide (NEM)

Key Methodologies

The experimental procedure relies on precise BDD surface preparation, electrochemical modification, and controlled chemical reaction steps.

BDD Surface Preparation: The 3 mm diameter BDD electrode was mechanically polished using 0.3 µm alumina slurry, followed by rinsing and drying with N₂ gas.
Mercury Film Electrodeposition: A fresh mercury film was applied before each analysis by transferring the BDE to a plating solution containing 20 mg/L Hg(II) and holding the working electrode potential at -1.30 V for 180 seconds.
Sample Extraction: Food samples were homogenized in 2% metaphosphoric acid and filtered (0.45 µm) to obtain the extract.
Ascorbic Acid (AA) Determination: The initial AA content was measured directly using DPV in an acetate buffer (pH 4.6).
DHAA Reduction: A separate portion of the extract was treated with 100 mM Tris(2-carboxyethyl)phosphine (TCEP) for 30 minutes to quantitatively reduce Dehydroascorbic Acid (DHAA) to AA.
Interference Elimination: 80 mM N-ethylmaleimide (NEM) solution was added to the reduced sample and shaken for 30 minutes to eliminate the voltammetric interference caused by TCEP.
Total Vitamin C (Tc) Determination: DPV was performed on the final treated solution. Total Vitamin C content was calculated based on the concentration of AA after the reduction step.

6CCVD Solutions & Capabilities

This research highlights the critical role of high-quality Boron-Doped Diamond (BDD) substrates in developing advanced electrochemical sensors. 6CCVD is uniquely positioned to supply the materials and customization services required to replicate and advance this work.

Applicable Materials

The foundation of this high-sensitivity method is the BDD electrode.

Material Recommendation: Heavy Boron-Doped Diamond (BDD) Substrates. 6CCVD supplies MPCVD BDD wafers and plates optimized for electrochemical applications, offering the wide potential window and low background current essential for DPV analysis in complex matrices.

Customization Potential

The success of the BDE relies on precise geometry and surface quality, areas where 6CCVD excels.

Research Requirement	6CCVD Capability	Technical Advantage
Electrode Dimensions (3 mm diameter disc)	Custom Dimensions & Laser Cutting. 6CCVD provides BDD wafers up to 125 mm (PCD equivalent) and offers precision laser cutting to achieve specific geometries (e.g., 3 mm discs) with high accuracy.	Ensures reproducible electrode surface area and integration into standard electrochemical cells.
Surface Quality (Required for stable Hg film)	Ultra-High Polishing. We guarantee surface roughness (Ra) < 5 nm for BDD plates, ensuring the smooth, defect-free surface necessary for stable, uniform electrodeposition of the mercury film.	Minimizes background current and maximizes the stability and lifetime of the modified electrode.
Integrated Contacts (Three-electrode system)	In-House Metalization Services. While the paper used external contacts, 6CCVD can deposit custom metal layers (Au, Pt, Ti, W, Cu) directly onto the BDD substrate.	Facilitates the development of robust, integrated microelectrode arrays or stable electrical contacts for long-term sensor deployment.
Substrate Thickness	Custom Thickness Control. BDD layers can be grown from 0.1 µm up to 500 µm, with substrates available up to 10 mm thick.	Allows engineers to select the optimal thickness for mechanical stability or specific device integration requirements.

Engineering Support

This study demonstrates a sophisticated application of BDD in high-sensitivity electrochemistry.

Expert Consultation: 6CCVD’s in-house PhD team offers specialized engineering support for projects involving voltammetric determination of biomolecules and sensor development in complex matrices. We assist clients in optimizing BDD material selection (doping concentration, thickness) and surface preparation protocols to meet stringent analytical requirements (LOD, stability).
Global Supply Chain: We offer reliable global shipping (DDU default, DDP available) to ensure researchers worldwide receive their custom BDD materials promptly and securely.

For custom specifications or material consultation, visit 6ccvd.com or contact our engineering team directly.

View Original Abstract

The objective of the study was to develop a new method for the determination of the total content of vitamin C and dehydroascorbic acid in food, based on the technique of differential pulse voltammetry with the use of a boron-doped diamond electrode modified with mercury film. A comparison was made between the results obtained with the developed method and a proposed reference method based on high-performance liquid chromatography with spectrophotometric detection. The reduction of dehydroascorbic acid was performed with the use of tris(2-carboxyethyl)phosphine. The interference caused by the presence of tris(2-carboxyethyl)phosphine during the voltammetric determination of ascorbic acid was effectively eliminated through a reaction with N-ethylmaleimide. The conducted validation of the voltammetric method indicated that correct results of analysis of the total content of vitamin C and ascorbic acid were obtained. Analysis of the content of dehydroascorbic acid was imprecise due to the application of the differential method. The results of the analyses and the determined validation parameters of the developed method are characterised by a high degree of conformance with the results obtained with the chromatographic reference method, which indicates the equivalence of the two methods.

Tech Support

Original Source

DOI: https://doi.org/10.3390/molecules28020812

References

2005 - Biological Role of Vitamin C in Keratinocytes [Crossref]
2011 - The Genetics of Vitamin C Loss in Vertebrates [Crossref]
1994 - Healthfulness and Nutritional Quality of Fresh versus Processed Fruits and Vegetables: A Review [Crossref]
2012 - Changes of Dehydroascorbic Acid Content in Relation to Total Content of Vitamin C in Selected Fruits and Vegetables
1984 - Dehydroascorbic Acid Levels in Fresh Fruit and Vegetables in Relation to Total Vitamin C Activity [Crossref]
2000 - Non-Spectrophotometric Methods for the Determination of Vitamin C [Crossref]
2008 - HPLC Methods for Simultaneous Determination of Ascorbic and Dehydroascorbic Acids [Crossref]
2014 - Determination of Vitamin C in Foods: Current State of Method Validation [Crossref]