Determination of Residual Peroxide in Food Packaging Materials using RQflex^® 20 with Reflectoquant^® Peroxide Test Strips

May May Lee, Chemistry R&D¹

Merck Pte Ltd

Introduction

Hydrogen peroxide (H₂O₂) is a mild antiseptic for cleaning and bleaching. It is approved by the US Food and Drug Administration (FDA) for packaging and surface sterilization in the food industry. The permissible limit is <0.5 ppm hydrogen peroxide in distilled water packaged under production conditions (assay to be performed immediately after packaging).¹

There is a need to determine residual peroxide in a rapid and sensitive manner in the fast production setting as conventional peroxide determination is mainly by titrimetric or spectrophotometric methods.

Experimental Procedure

Sample and Standard Preparation

Sample/Standard	Preparation
Samples are containers and packaging with direct food contact e.g., cups, tubs, boxes etc.	Rinse the hydrogen peroxide-treated container/packaging with an appropriate volume of pure water (minimum distilled or Type 2 from a suitable  Milli-Q® system, 1 – 10 mL depending on container size). Use the least volume practical to rinse the container. If the container is sealed, pierce it with a syringe filled with pure water and discharge the water fully into the container. Swirl the container thoroughly to wash the sides and use this solution immediately for the peroxide test.
Standards are varying concentrations of hydrogen peroxide from 0.25 to 15 mg/L	Dilute PerhydrolTM 30% H2O2 with pure water as follows: 10 mL PerhydrolTM 30% H2O2 to 100 mL (A) 6 mL (A) to 200 mL (B), resulting H2O2 concentration = 1000 mg/L 1.5 mL (B) to 100 mL (C), 15 mg/L H2O2 1 mL (B) to 100 mL (D), 10 mg/L H2O2 20 mL (D) to 25 mL (E), 8 mg/L H2O2 10 mL (E) to 20 mL (F), 4 mg/L H2O2 10 mL (F) to 20 mL (G), 2 mg/L H2O2 10 mL (G) to 20 mL (H), 1 mg/L H2O2 10 mL (H) to 20 mL (I), 0.5 mg/L H2O2 10 mL (I) to 20 mL (J), 0.25 mg/L H2O2 Use standards on the day prepared.
Assay of 1000 mg/L H2O2 standard	Pipette 50 mL of 1000 mg/L H2O2 solution into a 500 mL conical flask. Add 200 mL of pure water and 30 mL of 25% sulfuric acid. Mix well and titrate against 0.02 M potassium permanganate solution until a faint pink color is observed. Record titer (mL) and repeat for triplicates. Concentration of peroxide solution in mg/L = titer x (mL potassium permanganate solution) x 34.02. (See ref. 2)

RQflex^® 20 with Reflectoquant^® Peroxide Test Kit (0.2 to 20 mg/L)

Test Procedure
RQflex® 20 instrument calibration	Turn on the instrument and perform the calibration using the Recalibration Set (1.16954).
Negative control – pure water	Insert the barcode strip of the Reflectoquant® Peroxide Test into the RQflex® 20. Press the START key and simultaneously dip the test strip into the pure water (15 - 30 °C) for ~2 seconds ensuring that both reaction zones are immersed. Allow excess liquid to run off via the long edge of the strip onto an absorbent paper towel. Insert the strip into instrument and wait for the readout at 15 seconds.
Positive control check - H2O2 standards	Measure 0.5 mg/L H2O2 standard as above along with other concentrations as required.
Sample rinse solutions	Measure sample solutions as described above using the sample solution instead of pure water.

Results and Discussion

Results determined for a blank water solution, a positive control standard with 0.5 mg/L H₂O₂ and an example sample are shown in Table 1.

Table 1. Hydrogen peroxide values in pure water, sample, and standard.

Identity	1st reading (mg/L peroxide)	2nd reading (mg/L peroxide)	3rd reading (mg/L peroxide)	Ave Reading (mg/L peroxide)
Water Blank	< 0.2	< 0.2	< 0.2	< 0.2
0.5 mg/L H2O2 standard	0.6	0.6	0.6	0.6
Sample solution	< 0.2	< 0.2	< 0.2	< 0.2

If the peroxide-treated containers were fully dried before the water rinsing step, no peroxide would be detectable. This residual peroxide test can be employed to optimize the disinfection spray-dry cycles so that the residual peroxide is below the safe limit before the filling process.

Notes on the measurement:

1. For rinse solutions, the peroxide concentration measured in the sample solution is used.
2. If the measurement value exceeds the measuring range (HI is shown on the display), repeat the measurement using fresh, diluted samples until a value less than 20.0 mg/L H₂O₂ is obtained.
3. For the peroxide concentration at the full container capacity, the result of the analysis is:
   Measurement value x (rinse volume/container volume)

Conclusion

The RQflex^® 20 reflectometer along with the Reflectoquant^® peroxide test (0.2-20.0 mg/L) are suitable for the determination of residual peroxide in food packaging and containers. The RQflex^® 20 is a rugged and portable hand-held device, ideal for use in production lines. The system is sensitive to peroxide levels from 0.2 mg/L and does not require additional reagents or extensive sample preparation. This is an easy yet rapid quantitative test with a total measurement time of ~20 seconds for each sample compared to conventional titrimetric or spectrophotometric methods.

References

1. 2023. Food and Drugs, Chapter I, Subchapter B – Food for Human Consumption. [Internet]. CFR Title 21: Available from: https://www.ecfr.gov/current/title-21/chapter-I/subchapter-B/part-178/subpart-B/section-178.1005

2. 2023. Analytical Method: Analytical Quality Assurance, Standard for Hydrogen peroxide. [Internet]. Available from: https://www.sigmaaldrich.com/technical-documents/protocol/analytical-chemistry/photometry-and-reflectometry/analytical-quality-assurance-standard-for-hydrogen-peroxide