Planta Med 2010; 76(5): 495-497
DOI: 10.1055/s-0029-1186225
Biochemistry, Molecular Biology and Biotechnology
Letters
© Georg Thieme Verlag KG Stuttgart · New York

Successful Isolation and PCR Amplification of DNA from National Institute of Standards and Technology Herbal Dietary Supplement Standard Reference Material Powders and Extracts

Matthew T. Cimino1
  • 1Stoney Forensic, Inc., Chantilly, Virginia, USA
Further Information

Publication History

received May 21, 2009 revised September 11, 2009

accepted September 23, 2009

Publication Date:
20 October 2009 (online)

Abstract

Twenty-four herbal dietary supplement powder and extract reference standards provided by the National Institute of Standards and Technology (NIST) were investigated using three different commercially available DNA extraction kits to evaluate DNA availability for downstream nucleotide-based applications. The material included samples of Camellia, Citrus, Ephedra, Ginkgo, Hypericum, Serenoa, and Vaccinium. Protocols from Qiagen, MoBio, and Phytopure were used to isolate and purify DNA from the NIST standards. The resulting DNA concentration was quantified using SYBR Green fluorometry. Each of the 24 samples yielded DNA, though the concentration of DNA from each approach was notably different. The Phytopure method consistently yielded more DNA. The average yield ratio was 22 : 3 : 1 (ng/µL; Phytopure : Qiagen : MoBio). Amplification of the internal transcribed spacer II region using PCR was ultimately successful in 22 of the 24 samples. Direct sequencing chromatograms of the amplified material suggested that most of the samples were comprised of mixtures. However, the sequencing chromatograms of 12 of the 24 samples were sufficient to confirm the identity of the target material. The successful extraction, amplification, and sequencing of DNA from these herbal dietary supplement extracts and powders supports a continued effort to explore nucleotide sequence−based tools for the authentication and identification of plants in dietary supplements.

References

  • 1 Cole M R, Fetrow C W. Adulteration of dietary supplements.  Am J Health Syst Pharm. 2003;  60 1576-1580
  • 2 Betz J M, Fisher K D, Salanha L G, Coates P M. The NIH analytical methods and reference materials program for dietary supplements.  Anal Bioanal Chem. 2007;  389 19-25
  • 3 Crockett S L, Douglas A W, Scheffler B E, Khan I A. Genetic profiling of Hypericum (St. John's Wort) species by nuclear ribosomal ITS sequence analysis.  Planta Med. 2004;  70 929-935
  • 4 Sirvent T M, Walker L, Vance N, Gibson D M. Variation in hypericins from wild populations of Hypericum perforatum L. in the Pacific Northwest of the U.S.A.  Econ Bot. 2002;  56 41-48
  • 5 Southwell I A, Bourke C A. Seasonal variation in hypericin content of Hypericum perforatum L. (St. John's Wort).  Phytochemistry. 2001;  56 437-441
  • 6 Bruni R, Sacchetti G. Factors affecting polyphenol biosynthesis in wild and field grown St. John's Wort (Hypericum perforatum L.).  Molecules. 2009;  14 682-725
  • 7 Sucher N J, Carles M C. Genome-based approaches to the authentication of medicinal plants.  Planta Med. 2008;  74 603-623
  • 8 Lum M R, Hirsch A M. Molecular methods for the authentication of botanicals and detection of potential contaminants and adulterants.  Acta Hort. 2006;  720 59-72
  • 9 White T J, Bruns T, Lee S, Taylor J. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. Innis MA, Gelfand DH, Sninky JJ, White TJ PCR protocols. New York; Academic Press 1990
  • 10 Alschul S F, Gish W, Miller W, Myers E W, Lipman D J. Basic local alignment search tool.  J Mol Biol. 1990;  215 403-410

Ph.D. Matthew T. Cimino

Stoney Forensic, Inc.

Willard Road, Suite E 14101

Chantilly, VA 20151

USA

Phone: + 1 70 38 17 96 06

Fax: + 1 70 38 17 96 08

Email: matt@stoneyforensic.com