HPLC-FD Determination of Catechin, Epicatechin, and Procyanidins A2 and B2 for the Classification of Cranberry Products Sold in Europe

Reviewed: Bakhytkyzy I, Nuñez O, Saurina J. Determination of flavanols by liquid chromatography with fluorescence detection. Application to the characterization of cranberry-based pharmaceuticals through profiling and fingerprinting approaches. J Pharm Biomed Anal. 2018;156:206-213.

Keywords: Adulteration, cranberry, fingerprint, HPLC-FD, metabolomics, Vaccinium macrocarpon

Dietary supplements containing processed cranberry (Vaccinium macrocarpon or V. oxycoccos, Ericaceae) ingredients are among the most popular in the United States.¹ However, due to differences in the manufacturing process to make cranberry dietary ingredients, the composition of cranberry dietary supplements is highly variable. In addition, the inclusion of other extracts to cranberry, e.g., grape (Vitis vinifera, Vitaceae) seed extracts or heather (Erica spp. Ericaceae) extracts, make the evaluation of quality a challenging task. The quality assessment is further complicated by cases of substitution of cranberry powders or extracts with undisclosed ingredients mainly derived from plants that have high concentrations of proanthocyanidins (PACs), which has been reported in the literature.²

For this project, two cranberry extracts (provided by Deiters, S.L., Barcelona, Spain) and 17 commercial cranberry dietary supplements (2 tablets, 14 capsules, 2 sachets, and 1 syrup) purchased in pharmacies and specialty shops in Poland and Spain were analyzed by high-performance liquid chromatography combined with fluorescence detection (HPLC-FD). The results, either using quantitative data on catechin, epicatechin, and procyanidin dimers A2 and B2 (procyanidin C1 was measured as well, but was not detected in any of the commercial products), or a fingerprint measuring the areas of 14 most abundant peaks, were subjected to chemometric analysis using principal component analysis (PCA), or partial least squares (PLS) regression.

The commercial cranberry products were separated into four clusters, regardless of whether the data for flavan-3-ols or the fingerprinting of the 14 major peaks were used for the chemometric analysis. With the data from the flavan-3-ol quantification, one cluster with three commercial products aligned with the cranberry reference extract, characterized by the absence of catechin, and the presence of approximately equal amounts of procyanidins A2 and B2. Another cluster, which included an extract reportedly made from grapes, included two products that were predominantly composed of catechin and epicatechin, and contained only small concentrations of the procyanidin dimers. Two products having mainly procyanidin B2, smaller amounts of catechin and epicatechin, but no procyanidin A2, were also found in a separate cluster. Finally, the majority of the products (n = 10) clustered in the center between the three clusters described above.

A loadings plot (a graph representing the correlation between the peaks in a chromatogram and their contribution to the separation into principal components) provided information about the molecules contributing to the clustering of the various commercial products. Besides the flavan-3-ols, a number of phenolic acids (gallic acid, 3,4-dihydroxybenzoic acid, 4-hydroxybenzoic acid, vanillic acid, and syringic acid) were found to impact the discrimination among cranberry dietary supplements. The phenolic acids, and a number of stilbenes were identified in the fingerprint using chemical reference standards.

Comment: While not very common in industrial laboratories, the use of HPLC with a fluorescence detector provides a cleaner chromatogram of the main cranberry flavan-3-ol monomers and dimers than an ultraviolet/visible (UV/Vis) detector. The data suggest that cranberry dietary supplements can be categorized using catechin, epicatechin, and procyanidins A2 and B2. However, since some of the commercial products contained other PAC-rich materials, e.g., grape seed or heather extracts, it is unclear how much the additional ingredients impacted the results. As such, it would have been helpful to indicate which commercial products were made solely from cranberry, and which ones were made with a combination of ingredients. However, even the addition of grape or heather extracts cannot explain why some of the commercial products consisted mainly of catechin and epicatechin, and why procyanidin A2, which is known to be one of the main procyanidin dimers in authentic cranberry, was absent in two products.

References

1. Smith T, Kawa K, Eckl V, Morton C, Stredney R. herbal supplement sales in US increase 8.5% in 2017, topping $8 billion. HerbalGram. 2018;119:62-71.
2. Brendler T, Gafner S. Adulteration of cranberry (Vaccinium macrocarpon). Botanical Adulterants Prevention Bulletin. Austin, TX: ABC-AHP-NCNPR Botanical Adulterants Prevention Program; 2017:1-8.