Marker Compounds, Active Components and Reference Standards - Their Meaning and Use in Quality Determination of Botanical Products
By Ted Waszkuc, Ph.D., R&D Methods Department, NOW Foods
Botanical and herbal products are preparations containing one or more plant materials. Unlike conventional pharmaceutical products that are typically prepared from synthetic chemical compounds – usually present as single chemical - herbal products contain plant preparations that are mixtures of multiple components, in most cases representing multiple classes of natural compounds. Different preparations or batches from the same type of plant may vary in their chemical profile, depending on plant growing area, method of harvest, extraction procedures and storage conditions. For these reasons, assessing the quality of herbal products presents a far greater analytical challenge than a quantitative analysis of pharmaceutical products. The inherent complexity and variability of botanicals pose many problems for those charged with maintaining consistent product quality.
The concept of standardization is widely accepted to establish standard criteria for consistency and quality control of specific herbal preparations. As botanical preparations are mixtures of multiple constituents, the determination of a consistent chemical composition for a given plant preparation is one way of assuring product quality. Chemical standardization generally concerns the choice of one or more measurable compounds as being characteristic components of that herbal preparation. Specific levels of these characteristic compounds must be defined and then consistently maintained for each batch.
Despite the enormous scientific efforts to explain the mechanism of action for herbal preparations, there are still very few herbs in which the observed therapeutic effects can be assigned to the activity of one, or even a few, well-defined constituents. So called “active components” are substances which are both chemically identified and known to contribute to the therapeutic activity of a plant material or preparation.
If we don’t know which compounds are specifically responsible for the biological activity, one or more natural constituents present in the plant material, referred to as “marker compounds”, are selected and used to test for quality. A good marker should be characteristic of (or unique to) the given herbal preparation, be a substance with an established chemical structure, and not be present in other herbs contained in the finished product (when it is necessary to selectively quantify the content of one herb in a multi-herb product). For proper quality control a selected marker should be present in sufficient quantities to develop a scientifically valid test method, be accessible to quantify with common analytical equipment, be sufficiently stable and commercially available. Some compounds with a proven therapeutic activity can also be used as markers for quality assurance if they meet the criteria for analytical markers (e.g. kavalactones in Kava Kava or silybins/silydinins in Milk Thistle).
The most common chromatographic techniques used in quality control laboratories include HPLC, TLC and GC. In order to produce accurate and reliable results, these tests require the use of pure substances known to professionals as ‘reference standards’. A reference standard is a high purity (usually 99+%) chemical compound used as a comparison when detecting and determining the quantity of the same compound within a blended formula. Reference standards are used to calibrate the test instruments. This calibration allows us to compare the unknown amount of the active ingredient or marker contained within the test material to a known amount of the same substance in a highly purified form. Analytical reference standards for many botanical preparations are difficult to obtain or very expensive to purchase. Those that are not commercially available may be isolated from corresponding plant material or synthesized using appropriate methods.