RAPD-Based Method for the Quality Control of Mediterranean Oregano and Its Contribution to Pharmacognostic Techniques

Abstract

A pharmacognostic survey of 84 commercial samples of Mediterranean oregano, obtained from wholesale traders between 2001 and 2007, pinpointed the presence of extraneous plant material in 90.5% of the samples. In 59% of them extraneous material of plant origin was above 20%. Two major groups of botanical foreign matter were identified: oregano-like flavored plants ( Satureja montana L., Origanum majorana L.) and plants lacking a clearly detectable essential oil profile ( Rubus sp., Cistus incanus L., Rhus coriaria L.), added as bulk extraneous material. A random amplified polymorphic DNA (RAPD) method was developed to make the detection of the second group of adulterants easier and speed pharmacognostic analysis of large batches of samples. Thirteen primers discriminating between Origanum spp. and Rubus caesius , R.coriaria, and C. incanus were individuated, allowing their detection in oregano samples with a limit of detection of 1%. The utilization of RAPD as a reliable test to probe the authenticity of Mediterranean oregano or previously screen the presence of specific contaminants is proposed as a complementary approach to pharmacognostic and phytochemical screening.

Development of a SCAR marker for the identification of Olea europaea L.: A newly detected adulterant in commercial Mediterranean oregano

“…In order to ascertain whether this new adulterant was correctly identified a specific PCR system was designed based on previous knowledge of Mediterranean oregano RAPD and SCAR markers and developing specific SCAR markers for O. europaea leaves. Two genotypes of O. europaea coming from different geographical zones of Mediterranean basin (Italy and Montenegro) were subjected to RAPD analysis and compared with ten genotypes belonging to seven (Marieschi et al, 2009) allowed the identification of a RAPD band monomorphic (OPB12Oe 392 ) for the two considered olive genotypes and absent in the profiles generated by the amplification of the Origanum samples. This discriminating RAPD amplicon was selected for the conversion into a SCAR marker (Fig.…”

“…Genomic plant DNA was isolated from fresh and dried material as previously described (Marieschi et al, 2009) with some modifications proposed by Tibbits, McManus, Spokevicius, and Bossinger (2006), to increase the yield and the purity of the DNA extracted and reduce the inhibitory effects of carbohydrates and polyphenols in the subsequent PCR reactions. Briefly, 100-200 mg of plant material were placed in 2 ml tubes with 900 ll of CTAB buffer in the presence of activated charcoal and PVP.…”

“…This hindrance was avoided by enhancing the PCR amplification through the addition of BSA 0.4% and a non ionic detergent like Triton X-100 0.5% or Tween 20 0.5% (Demeke & Adams, 1992;Monteiro et al, 1997;Rådström, Knutsson, Wolffs, Lövenklev, & Löfström, 2004;Tibbits et al, 2006). The integrity and availability of DNA from dried commercial samples for SCAR analysis was checked by performing an amplification with a couple of primers, constructed on a monomorphic RAPD amplicon (OPA07Ospp 670 ) obtainable from the amplification with the primer OPA07 on different Origanum species (Marieschi et al, 2009). The bands, nearly 670 bp long, obtained from O. onites, O. vulgare, O. vulgare subsp.…”

“…In particular, the more experienced the perpetrator gets in avoiding routine and often non specie-specific controls, the higher the risk of adulteration, to the detriment of both consumers and honest traders. A recent survey of the European market from 2000 to 2007 revealed that, more often than not, products sold and purchased as true Mediterranean oregano were not corresponding to what declared on the label (Marieschi, Torelli, Poli, Sacchetti, & Bruni, 2009). A continuation of this survey pinpointed a previously unreported adulteration of commercial Mediterranean oregano with dried crushed leaves having a similar silvery grey colour, but devoid of glandular hairs and with characteristic xerophytic adaptations (thick lamina, peltate trichomes, abundant cuticular coating).…”

“…Fresh plant material was collected, immediately freeze-dried in liquid nitrogen and stored at À80°C until molecular analysis. Mediterranean oregano dried samples were purchased from international wholesale traders as previously reported (Marieschi et al, 2009). Once acquired, in order to simulate real commercial conditions, samples have been stored in dry and dark conditions at 5°C until performing molecular analysis.…”