A UV assay counts everything that absorbs at one wavelength. HPLC separates the compounds first and counts the ones it names. The two figures answer different questions.
Two offers arrive for what looks like the same extract. One states the active content by UV, the other by HPLC, and the UV figure is clearly the higher. Neither has to be wrong, and the material may even be the same. The two methods count different things, and each states its result in terms of a named substance.
Here are six checks to make before you compare two assay figures, or compare a certificate with your own laboratory’s result. They draw on European guidance for herbal medicines, the international guideline on validating analytical methods and published studies; the example comes from our own specifications. Our earlier articles covered the specification, the extract ratio, the Certificate of Analysis, the solvent and carrier, and the contaminant rows; this one stays with the assay method.
01 / THE METHOD
Does the assay row name the method?
An assay figure belongs to the method that produced it. European guidance for herbal medicines builds this into its definition of a specification, “a list of tests, references to analytical and biological procedures, and appropriate acceptance criteria”, and conformance means that the material, “when tested according to the listed analytical procedures, will meet the listed acceptance criteria”.1
On a specification the method is often one short word after the figure: UV, HPLC or another technique. A row without it is incomplete. A 2023 review by authors from the American Botanical Council and partner organisations notes that results from spectrophotometric assays “are sometimes the sole data on the chemical composition of a botanical ingredient provided on a Certificate of Analysis”, and it adds that declaring the methodology behind a quantitative result “should become standard industry practice”.2
Check: that the assay row names the method, and that the specification, the quotation and the certificate all name the same one.
02 / UV
What does a UV figure count?
On a specification, UV stands for UV-visible spectrophotometry. The extract is dissolved, in many assays a reagent turns the compounds of interest into a coloured product, and the light the solution absorbs at one wavelength is measured. The compounds in the solution are not separated from one another, so everything that absorbs at that wavelength adds to the result.2
The review states the consequence: concentrations obtained by spectrophotometry “are often higher than the results from other quantitative methods due to interference from nontarget analytes that absorb at the wavelength of interest”. Its example is milk thistle extract, where silymarin concentrations “are generally between 30% and 65% by HPLC-UV and 65–80% using spectrophotometric tests”.2
The guideline on validating analytical methods has a name for this property. Specificity and selectivity describe “the extent to which other substances interfere with the determination of an analyte”.3 European guidance for herbal medicines takes a UV-visible assay as its example of a non-specific assay.1 A UV figure is therefore a figure for a group of compounds, and it should be read as one.
Check: for a UV figure, which group of compounds it is meant to count, and the reagent and wavelength the method uses.
03 / HPLC
What does an HPLC figure count?
HPLC, high-performance liquid chromatography, separates the dissolved extract into its compounds before anything is measured. The compounds then show as separate peaks, and the method counts the peaks it names. The detector behind the column is usually a light measurement too: the review calls the UV/vis detector the “most widely used detection system”.2 The main difference between the two methods is the separation.
So an HPLC figure stands for the compounds the method names, one or several added together, and the specification should say which. The European Medicines Agency’s report on bilberry shows the pattern. It quotes the European Pharmacopoeia’s definition of the refined and standardised dry extract of fresh bilberry fruit: 32.4% to 39.6% of anthocyanins, “expressed as cyanidin 3-O-glucoside chloride”. The chromatogram for that assay, as the report lists it, shows twenty named peaks: fifteen anthocyanins and five anthocyanidins, their sugar-free forms.4
A compound the method does not name is not counted, however much of it the extract holds. And some groups can be separated only in part: for proanthocyanidins, the review notes, separation of the larger molecules “by chromatographic means is difficult; hence, suppliers generally use spectrophotometric methods for quantitative analysis”.2
Check: for an HPLC figure, the compounds that are counted, and whether the figure is their sum or one named compound.
04 / THE STANDARD
Against which standard, and expressed as what?
In a content assay, neither method reads a percentage off the sample. The signal is converted into an amount by reference to a named substance, which is either measured alongside as a standard or built into the calculation; in European guidance, reference standards are used “for identity and purity testing and for content assignment”.1 The result is then stated in terms of a named substance or group. The guidance on declaring herbal substances and preparations shows the wording: for senna leaf, hydroxyanthracene glycosides “calculated as sennoside B”; for a ginkgo leaf extract, flavonoids “expressed as flavone glycosides”.5 What is named is not always what was measured: pharmacopoeia methods for ginkgo usually determine the flavonoids after acid hydrolysis, which splits off their sugars.2
Change the reference substance and the figure changes, although the method and the material stay the same. For cranberry proanthocyanidins measured by a colour reaction, the DMAC assay, one study reports that a standard prepared from cranberry itself “resulted in values that were 2.2 times higher than those determined by procyanidin A2”.6
The form of the molecule counts as well. The bilberry report describes one extract by two figures, “36% anthocyanosides, corresponding to 25% anthocyanidins”: the second figure counts the same molecules without their sugar part.4
Our cranberry extract is an example of an assay statement that names its method and its standard. It is specified at 30% or more total soluble proanthocyanidins (PACs) by BL-DMAC / UV-Vis, expressed as procyanidin A2 equivalents. The assay uses procyanidin A2 as its reference standard and measures absorbance at 640 nm; it does not establish isolated A-type PAC purity. Results obtained using different analytical methods or calibration standards are not automatically comparable.
Check: the reference substance, and what the result is expressed as: one named compound, a group, or the molecules with or without their sugar part.
05 / THE CHOICE
Which method suits the figure you need?
HPLC is not the better method in every case. The validation guideline states what validating any method has to show: “that the analytical procedure is fit for the intended purpose”.3 Where the marker is one compound, or a few, a separating method can count them directly. Where it is a whole group that chromatography cannot resolve, a group assay is the usual tool.2
A multi-laboratory study of the DMAC assay for cranberry called it “a simple, robust and relatively specific spectrophotometric assay for total PACs in cranberry samples”, and added that it “may not be appropriate for comparing concentrations across different food types”.7
A group assay cannot show where the compounds came from: of the colour reactions used for proanthocyanidins, the review says that none “can distinguish among PACs from different sources”.2 European guidance for herbal medicines prefers a specific procedure wherever one is possible; where a non-specific assay is justified, it points to a second test: “a combination of the assay and a suitable test for identification (e.g. fingerprint chromatography) can be used”.1 The validation guideline puts it generally: a lack of specificity “should be compensated by one or more other supporting analytical procedures, unless appropriately justified”.3
Check: that the method suits the marker and, where a group assay is used, which identification test stands beside it.
06 / YOUR LABORATORY
Will your own test use the same method?
A figure can be confirmed only by the method that produced it. If your laboratory tests a delivery by HPLC against a specification written by UV, its result is not a result against that specification, whether it comes out lower or higher. There is no general factor for converting one figure into the other: the gap depends on what else the extract contains and on how each result is expressed.
Even with one method, two laboratories do not return the same number. In the multi-laboratory study, five laboratories analysed the same eleven cranberry samples by the same DMAC method. Results varied by 4.1% on average within a laboratory and by 16.9% on average between laboratories (range 8% to 32%), measured as relative standard deviation.7
So settle three things before the order: the method, by name and by reference to a pharmacopoeia or a written procedure; the reference substance; and the difference between two laboratories that both sides will accept. For medicines with a pharmacopoeia monograph, European guidance says which method decides when a different one was used: “In case of doubt or dispute, the methods of analysis of the Pharmacopoeia are authoritative.”1 Where no pharmacopoeia method applies, the agreed specification has to do that work. Our article on the Certificate of Analysis covers the checks to make when the certificate arrives.
Check: that your laboratory and the laboratory behind the certificate use the same method and the same reference substance, and what difference between their results both sides accept.
WHAT TO SEND
State the method with your enquiry
When an assay figure matters to your product, tell us:
- the ingredient, and the plant part if more than one is offered;
- your intended application and destination market;
- the compound or group you need, and the target level;
- the assay method your quality team uses, and the reference standard where it forms part of the assay;
- whether you will test the delivery yourself, and by which method;
- which technical documents you need, such as a specification sheet or Certificate of Analysis;
- your name, company and contact details.
An active-content percentage is more useful when it is paired with the analytical method and reference standard. Tell us these requirements with your target level so that the specification and quotation describe the material your project needs. Specification ranges in our catalogue are references, and feasibility, availability and price are confirmed with your quotation.
The product example is taken from YOHOLIFE® specifications and technical documents. This article gives no other assay values for our ingredients; the method and specification for a selected grade are confirmed when you enquire. Sources 1, 3, 4 and 5 were written for medicines and are cited here for their definitions and examples; source 2 is a review of published work; sources 6 and 7 are studies of one assay for cranberry and were read in their published abstracts. Sources were checked on 12 November 2026.
