Immune Modulating Activities of Echinacea

Published Immune Stimulation Activity:
Joseph A. Rininger¹, Kerry Ringer² and Mark Savarese²
Columbia PhytoTechnology LLC , NE Hickman Court, Suite 4 Pullman, WA 99163

Summary

EchImmune® (CPT-121) has been optimized to immunostimulatory activity. We have identified polysaccharides as the compounds responsible for this activity. EchImmune is unique because it is specially processed (including use of our patented Radiant Zone Drying technology) to preserve the bioactivity of the polysaccharides. Spray drying and other processes that utilize high heat or non-ambient pressure change destroy the size, shape and therefore activity of polysaccahrides. Spray dried products include short chain or damaged polysacchrides (also called oligosaccharides) in standardization. A special step in analysis assures we are only measuring the larger, proven bioactive polysaccharides in EchImmune standardization.

Polysaccharides Are Not Found In Echinacea Extracts

There are four types of constituents purported as pharmacologically active molecules found in Echinacea species: phenolic caffeic acid derivatives, alkylamides/isobutylamides, polysaccharides and glycoproteins. In commercially prepared Echinacea extracts, the quantities of some of these constituents are measured to ensure that these presumed active ingredients are present. The development of "standardized" Echinacea extracts has occurred from the need to produce a more consistent product and the desired benefits on a continual basis. However, techniques to enrich for one class of constituents typically reduce or exclude the others.

Products standardized to phenolics, such as cichoric acid, and isobutylamides do not contain polysaccharides due to differences in solubility. Also, current industry practices which include atomization, boiling under vacuum, and excessive heat deactivate active polysaccharides by reducing the size and shape of these complex carbohydrates.

Polysaccharides Are Proven To Be Immunostimulatory

The most common constituents found in standardized extracts include polyunsaturated alkylamides or caffeic acid derivatives such as cichoric, chlorogenic and caftaric acids. These compounds have been shown to inhibit cyclooxygenase and 5-lipoxygenase, key enzymes associated with inflammation via the production of prostaglandins and leukotrienes¹^,². Overall, these activities are not direct immunostimulatory activities. This is further supported by in vitro and in vivo studies assessing immune parameters in laboratory animals that have shown no immune stimulating activity of chlorogenic and cichoric acid tested as a single agents.³^,⁴^,⁵

In contrast to the limited in vitro experimental evidence of immunostimulatory activity of caffeic acid derivatives and alkylamides, there is consistent and convincing evidence for the role of Echinacea polysaccharides to directly stimulate immune cells. Wagner et al (1988)6and Steinmüller et al (1993)7 have worked extensively to investigate the immunostimulatory effects of polysaccharides from Echinacea. ⁶^,⁷^,^8,⁹^,¹⁰^,¹¹ These researchers were successful in isolating several polysaccharide structures, including a variety of arabinogalactans. The complex and high molecular weight (10 to 75 kDa) polysaccharides were found to directly activate non-specific immune cell types such as monocytes, macrophages, and natural killer (NK) cells. Echinacea polysaccharide-induced stimulation of these cell types initiated cytokine production (TNF-a), elevated phagocytic activity and oxidative burst, resulting in enhanced in vitro and in vivo killing of Leishmania, Listeria and Candida pathogens⁶^,⁷^,^8,⁹^,¹⁰^,¹¹. Importantly, the in vitro characterization of the polysaccharide activity was dose-dependent and with potent stimulation occurring at concentrations <10 mg/mL. In addition, there is a likely mechanism of action for polysaccharide-induced stimulation of immune cell types through the binding and activation of cell surface receptors present on target immune cells. The Echinacea polysaccharides were subsequently shown to activate non-specific immune cells when evaluated in animal models as well as human subjects ⁷^,^8,⁹^,¹⁰. This characterization of Echinacea polysaccharides is the best demonstration of in vitro bioassay activity yielding reproducible in vivo pharmacological effects.

Immune Cell Stimulation Assay

The macrophage stimulation assay published by Rininger et. al. 20005 measures the stimulation of immune cells (murine macrophages) by quantifying cytokine production (including TNF-α), important markers for immune cell stimulation. A simulated digestion protocol (CACO II) was also used to test the bioavailability of the immunostimulatory polysaccharides in EchImmune®.

The first step in the development of CPT-121 was to perform an assessment of macrophage immunostimulatory activity of different Echinacea extracts and fractionations. This included various Echinacea standard extracts, a raw herb powder, and preliminary fractionation of E. purpurea aerial parts as starting material for the development of CPT-121. The results showed that Echinacea extracts standardized to phenolic or isobutylamide constituents and fresh pressed juice tested negative for macrophage activation (see the comparison graph).

Testing of subsequent production batches of CPT-121 extract consistently demonstrated a similar level of the immunostimulatory potency. Dose-response experiments showed that the stimulatory activity was enhanced approximately 10-fold in comparison to the initial starting herb material (see the comparison graph).

Our Unique Process Extracts And Preserves The Activity of Polysaccharides

The immunomodulatory active polysaccharides in Echinacea, 4-0-methyl-glucuronoarabinoxylan and acidic arabinorhamnogalactan are found in the hemicellulose. ¹²^,¹³ A less active pectin polysaccharide, xyloglucan, has been isolated from the expressed juice of E. purpurea, but possesses only weak immunostimulatory activity. ¹³
Our unique process is capable of extracting the hemicellulosic polysacchrides without degrading their activity. Once they are extracted we use a cold, no-vacuum concentration method, and our patented drying technology to preserve the size, shape and, activity of the polysaccharides.

Conclusion And Continuing Research

The design and implementation of in vitro experimental approaches are of central importance to define pharmacological profiles of herbal medicines and provide credible evidence for efficacy to be assessed in clinical studies. Extracts from other popular herbal medicines including Hypericum perforatum (St. John’s Wort) and Serenoa repens (Saw Palmetto) that possess a defined in vitro pharmacology have subsequently been proven to have efficacy in human clinical trials as an anti-depressant and for a treatment of benign prostate hyperplasia, respectively⁶^,¹⁴^,¹⁵^,¹⁶^,¹⁷^,¹⁸. Unlike these two examples, other Echinacea extracts currently do not have a truly defined pharmacology. Therefore, it is not surprising that the clinical effectiveness of Echinacea preparations for the treatment (to provide faster resolution), prevention, or alleviation of symptoms of colds and flu are inconclusive ¹⁹^,²⁰^,²¹.

CPT-121 (EchImmune®) is an Echinacea preparation with a quantified dose of polysaccharides that shows in-vitro immunostimulatory properties. Polysaccharide standardized Echinacea products need to be included in effective formulations, particularly those purported as immunostimulatory where the prevention and reduction in length of colds and flues is an important pharmacological response. Recently, polysaccharide content in CPT-121 has been correlated directly to macrophage activation of both murine and human cell lines. Our current research with the product is addressing important questions for future clinical work.

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