Isotopic Fingerprinting and NMR Analysis Detects Designer Blends Made with Animal Fats Marketed as Saw Palmetto Extracts

Reviewed: Perini M, Paolini M, Camin F, Appendino G, Vitulo F, De Combarieu E, Sardone N, Martinelli EM, Pace R. Combined use of isotopic fingerprint and metabolomics analysis for the authentication of saw palmetto (Serenoa repens) extracts. Fitoterapia. 2018;127:15-19.

Keywords: Adulteration, animal fats, GC-FID, ¹H NMR, isotopic fingerprint, metabolomics, Serenoa repens

Saw palmetto (Serenoa repens, Arecaceae) berry extracts are widely used in dietary supplement products for urinary problems related to benign prostatatic hyperplasia. The geographic area where the saw palmetto tree naturally occurs is limited to the southeastern United States, and due to a limited supply, growing demand, and increasing prices, the appearance of vegetable oils, or mixtures of vegetable oils (e.g., canola, coconut, olive, palm, sunflower oils) with saw palmetto extracts marketed as authentic saw palmetto oil has been observed.¹ The problem of saw palmetto adulteration is exacerbated in years when the harvest volume is low due to inclement weather conditions.

Over the past years, suppliers have noticed the sale of saw palmetto extracts that appeared to comply with the pharmacopeial fatty acid ratio requirements,^2,3 one of the main tests to detect saw palmetto extract adulteration. However, these ingredients sometimes failed other tests, such as color specifications, or limits on fatty alcohol and sterols.

The approach taken by the researchers from Indena, SpA (Milan, Italy), the Fondazione Edmund Mach (San Michele all’Adige, Italy), and the University of Eastern Piedmont (Novara, Italy), was to evaluate nine commercial extracts (from suppliers outside Europe and North America) and 30 authentic saw palmetto extracts by the fatty acid method of the United States Pharmacopeia (USP),² and by orthogonal methods, such as ¹H NMR and isotopic fingerprinting analysis with a subsequent statistical assessment by principal component analysis (PCA).

For the isotopic fingerprint analysis, the ¹⁴C concentration, and the ratios of ⁸⁷Sr/⁸⁶Sr, ¹³C/¹²C, ¹⁸O/¹⁶O, and ²H/¹H were determined. The amount of ¹⁴C in a sample is indicative of its being derived from plants or from fossil-fuel (i.e., made by chemical synthesis). The strontium (⁸⁷Sr/⁸⁶Sr) ratio provides information about the nature of the soil and the climatic conditions from where the sample is obtained. The carbon (¹³C/¹²C) ratio can be used to distinguish among certain plant-derived materials, since not all plants use the same photosynthetic pathways. As such, materials derived from corn or sugar cane have higher ¹³C/¹²C ratios than most of the plants used medicinally. The oxygen (¹⁸O/¹⁶O) and hydrogen (²H/¹H) ratios are based on the local water composition, mainly influenced by precipitation in form of snow and rain in the area.

No differences were observed in the ¹⁴C concentrations, and the ⁸⁷Sr/⁸⁶Sr ratio among authentic saw palmetto extracts and the commercial samples. However, the ¹³C/¹²C ratio was slightly, but significantly lower in the commercial samples, suggesting adulteration. This was confirmed by the ¹⁸O/¹⁶O, and ²H/¹H ratios, which differed substantially from those of the authentic material. Comparison with ¹⁸O/¹⁶O, and ²H/¹H ratios from various fats and oils suggested that six out of the nine commercial samples were derived from animal fats.

The GC-FID analysis of the fatty acids with subsequent PCA led to a clustering of the six animal-derived ingredients with the authentic saw palmetto, suggesting that these ingredients were blended with the fatty acid composition of authentic saw palmetto in mind. Two of the three remaining commercial saw palmetto extracts clustered with coconut (Cocos nucifera, Arecaceae) and babaçu (Attalea speciosa, Arecaceae) oil, while one sample neither clustered with authentic saw palmetto extract nor with any of the vegetable oils.

A clear separation between adulterated and genuine saw palmetto extracts based on differences in the ¹H NMR metabolomics was also obtained for seven of the ten samples analyzed (apparently one additional commercial sample was evaluated by ¹H NMR). The remaining three commercial samples clustered closely to the authentic materials. It is not clear if the ¹H NMR approach would have been sufficient to detect the adulteration in these three cases, although a refinement of the statistical model may have provided more clarity on this.

Comment: This paper provides the first evidence for the use of animal fats to adulterate saw palmetto extracts. It also shows the sophistication with which fraudulent suppliers are able to create extracts that are so similar in the composition to the authentic extracts that the adulteration most likely will go unnoticed when using standard analytical methods. For some of these materials, differences in color and smell (as saw palmetto smells very foul) may provide the first indication of a quality issue. For other commercial extracts, only additional tests, such as the isotopic fingerprinting described here, may provide sufficient evidence of adulteration. In order to ensure authentic saw palmetto extract, a tight control of the supply chain, including assurance that the berries were grown in the United States and processed by a reputable company, are necessary in addition to a sound quality control regimen. Extra caution needs to be taken when considering the purchase of saw palmetto extracts from countries in Asia.

References

1. Gafner S, Baggett S. Adulteration of Saw Palmetto (Serenoa repens) – Botanical Adulterants Bulletin. Austin, TX: ABC-AHP-NCNPR Botanical Adulterants Prevention Program. 2017;1-6. http://cms.herbalgram.org/BAP/BAB/SawPalmettoBulletin.html. Accessed June 25, 2018.
2. United States Pharmacopeial Convention. Saw Palmetto Extract. In: United States Pharmacopeia and National Formulary (USP 41-NF 36). Rockville, MD: United States Pharmacopeial Convention. 2018.
3. The European Directorate for the Quality of Medicines & HealthCare. European Pharmacopoeia (EP 9.1). Sabalis serrulatae extractum. Strasbourg, France: Council of Europe; 2016:1509-1512.