K₂₆₈ Olive Oil: According to Commission Regulation (EEC) No 2568/91 Annex IX

UV-spectrophotometric determination of the specific absorbance at 268 nm; method No. 2526

INTRODUCTION

The K₂₆₈ test is a significant analytical method used for assessing the quality and oxidative stability of olive oils. Specifically, it measures the absorbance of olive oil samples at a wavelength of 268 nm. This test is crucial in detecting the presence of conjugated dienes, which are indicators of oxidation in the oil. By evaluating the K₂₆₈ values, it becomes possible to determine the extent of oxidation and monitor the freshness and overall quality of olive oil.

In this application note, the determination of K₂₆₈ values of olive oils is described using UV-spectrophotometric analysis, following the guidelines outlined in Commission Regulation (EEC) No 2568/91 Annex IX.¹

MATERIALS AND REAGENTS:

• Isooctane for spectroscopy Uvasol^® (1.04718)
• Rectangular cells quartz 10 mm (1.00784)
• Spectroquant^® UV/VIS Spectrophotometer Prove 300 (173017) or
• Spectroquant^® UV/VIS Spectrophotometer Prove 600 (173018)
• Volumetric flasks, class A, volume range 25 mL to 100 mL
• Analytical balance 0.001 g
• Paper filters

METHOD AND MEASURING Range:

• Method No. 2526
• 0 – 4.00 K₂₆₈

SAMPLE PREPARATION:

• Homogenize liquid samples at 30 °C.
• Heat solid samples to a temperature of max. 10 °C above the melting point, melt and homogenize. If contaminations are present, filter the sample over a fast-filtering paper.
• Weigh the sample into a volumetric flask, accurately weighed to 1 mg.
• The quantity of the sample and volume of the volumetric flask must be selected in such a way that the measurement solution exhibits an absorbance between 0.1 and 0.8 A at 268 nm. The following table can be used for orientation:

K268 value	Weight	Volumetric flask
<0.10	1.00 g	25 mL
0.10 - 0.80	0.25 g	25 mL
0.80 - 1.60	0.25 g	50 mL
1.60 - 4.00	0.20 g	100 mL

• Add a few milliliters of isooctane and dissolve the sample at room temperature. Subsequently, make up the contents of the volumetric flask to the mark with isooctane and mix. Filter turbid solutions over a paper filter.

ZERO ADJUSTMENT

For each measurement series, a zero adjustment is required. It is recommended to use the same cell for zero adjustment and for sample measurement. The zeroing procedure for the measurement series is automatically prompted by the instrument.

• For zero adjustment fill 10-mm rectangular quartz cell with isooctane.
• After prompting, place the rectangular cell filled with isooctane in the cell compartment, the zero adjustment is executed automatically.
• Confirm the implementation of zero adjustment with "OK".
• The zero adjustment is valid for the entire measurement series.

MEASUREMENT PROCEDURE

• Select method 2526 "K₂₆₈ (olive oil)" from the method list.
• Enter the weight in grams, and confirm with "OK".
• To enter the volume of the sample solution, tap the input field in the “Sample Solution” line, enter the volume in milliliters, and confirm with "OK".
• Press "START" to reach the measuring mode.
• Before measuring the first sample, the system automatically prompts a zero adjustment (procedure see "Zero adjustment").
• Transfer the sample solution into a 10-mm rectangular quartz cell and place the rectangular cell into the cell compartment, the measurement is performed automatically.
• Confirm measurement with "OK".
• The K₂₆₈ value is shown in the display.
• Press "START" to start the measurement procedure for the next sample. The system does not prompt a repeat of the zero adjustment.

Note: According to Commission Regulation (EEC) No 2568/91 Annex IX the method checks automatically if the measured absorption is in a range of 0.100 – 0.800 A. If the absorption is not in this range, a pop-up message appears: “Condition not met – Absorbance < 0.1” resp. “Condition not met – Absorbance > 0.8”. In these cases, the sample preparation needs to be adjusted. Use an appropriate sample weight in a range of 0.100 – 1.000 g and a volumetric flask in a range of 10 – 100 mL to achieve these absorption values.

CONCLUSION

This method is also suited for other oils and fats and in addition to Commission Regulation No 2568/91 Annex IX is based on the:

• DGF Standard Methods C-IV 6 (13)
• ISO 3656:2011
• COI/T.20/Doc. No. 19/Rev. 3 (2015)
• CODEX STAN 33-1981 (2013)
• AOCS Ch 5-91(2009)

Low K values are characteristic of high-quality, extra native olive oils; refined or aged olive oils exhibit higher K values at the 232, 268, and 270 nm wavelengths. The limits for olive oils can be referenced from Commission Regulation No. 2568/91 Annex IX.

For more applications with photometry & reflectometry see SigmaAldrich.com/wfa-applications

References

1. Commission Regulation (EEC) No 2568/91 of 11 July 1991 on the characteristics of olive oil and olive-residue oil and on the relevant methods of analysis. . [Internet]. Available from: http://data.europa.eu/eli/reg/1991/2568/oj