One bottle, two numbers

Why NMR and HPLC give different polyphenol figures for the same olive oil. Scroll, and the drawing follows the words.

One drawing of the same olive oil measured two ways Left, a bottle of olive oil with a magnified view of the phenolic compounds inside it. Top right, the NMR bench: a glass tube lowered into a superconducting magnet, a radio pulse, and a spectrum with one signal per compound, tallied in mg per kg. Bottom right, the HPLC bench: the phenols extracted into methanol and water, pumped through a packed column where some change form, measured by a detector against tyrosol. Bottom left, the two numbers side by side, with no conversion between them, and the rule: compare NMR with NMR and HPLC with HPLC. EXTRA VIRGIN OLIVE OIL polyphenols: a number inside the oil, magnified enormously oleocanthal oleacein oleuropein aglycone ligstroside aglycone hydroxytyrosol tyrosol Method one: NMR Method two: HPLC Method one: NMR Method two: HPLC a thin glass tube superconducting magnet chilled with liquid helium a shortradio pulse oleocanthal oleacein oleuropein aglycone ligstroside aglycone hydroxytyrosol tyrosol one signal per compound, each in its own place each counted as itself, mg per kg (example) oleocanthal4 oleacein3 oleuropein aglycone1 ligstroside aglycone1 hydroxytyrosol2 tyrosol1 total12 oil phenols pulled out of the oil into methanol and water solvent pump injector the column, packed with tiny particles detector two of the big compounds changed into other forms on the way through shown by the Athens group, 2014 what came out, counted against tyrosol tyrosol, the reference 9 tyrosol units example the same bottle, measured twice NMR 12 mg per kg each compound counted as itself HPLC 9 tyrosol units what came through the column example counts. the papers give the direction, not the size no conversion Compare NMR with NMR. HPLC with HPLC.
What is in the bottle

This is a bottle of extra virgin olive oil. When a label says high phenolic, it is talking about natural compounds inside the oil.

What is in the bottle

Those compounds are called phenols. They are far too small to see, but magnified enormously they would look something like this.

What is in the bottle

Four big ones matter most: oleocanthal, oleacein, oleuropein aglycone and ligstroside aglycone. These are the ones the phenol number is mostly about.

What is in the bottle

Two smaller ones matter too: hydroxytyrosol and tyrosol. Keep an eye on tyrosol. It comes back later.

What is in the bottle

Nobody can count these by eye, so a laboratory measures them. There are two main ways to do it. Here, the same oil goes to both.

Method one: NMR

Method one is NMR, short for nuclear magnetic resonance. A few drops of the oil, gently prepared, go into a thin glass tube.

Method one: NMR

The tube is lowered into a very strong magnet, and a short radio pulse is sent through the sample.

Method one: NMR

Every compound answers with its own signal, in its own place, a bit like a fingerprint. Oleocanthal here, oleacein there. Nothing is separated out and nothing is changed.

Method one: NMR

The lab reads each signal and writes down how much of each compound there is, in mg per kg: milligrams in every kilogram of oil. Add them up and that is the NMR number.

Method two: HPLC

Method two is HPLC, high performance liquid chromatography. First the phenols are washed out of the oil into a mix of methanol (a type of alcohol) and water.

Method two: HPLC

That liquid is injected into a flowing stream and pumped through a narrow tube called a column, packed with tiny particles.

Method two: HPLC

Different compounds cling to the particles for different lengths of time, so they spread out and leave the column one after another.

Method two: HPLC

On the way through, some of the big compounds change into other forms. A team in Athens showed this in 2014 by testing the liquid before and after the column.

Method two: HPLC

A detector at the end measures what comes out. Then everything is counted as if it were one compound, tyrosol. The result is a total in tyrosol units.

What it means

Same bottle. Two methods. Two different numbers. NMR counted each compound as itself. HPLC counted what came through the column, in another compound's units.

What it means

You cannot convert one into the other. The gap depends on how much of the big compounds the oil had to start with, and that changes from oil to oil.

What it means

So compare NMR with NMR, and HPLC with HPLC. If a bottle just says a number, find out how it was measured. If you can't, the number tells you nothing.

The counts on screen are illustrative. What the papers show is the direction: NMR reads each compound directly, HPLC's column changes some of them on the way through. Karkoula, Skantzari, Melliou, Magiatis, J. Agric. Food Chem. 2012 (10.1021/jf3032765) and 2014 (10.1021/jf404421p). Starec et al., University of Trieste, Molecules 2021 (10.3390/molecules26010242), both methods on the same oils. International Olive Council COI/T.20/Doc. No 29 Rev. 2, 2022. I'm not a doctor, so nothing here is medical advice. These are measurement methods, not health claims. Deon Bryan, official Atsas distributor for the UK and Ireland.