Study on Echinacea Enhance Pig Immune System

Echinacea, also known as “coneflower”, is a perennial herb in the Asteraceae family that is native to North America. It is one of the oldest herbal medicines in Western history, with a medicinal history that can be traced back to the time of the North American Indians. There are nine species and several varieties of Echinacea, three of which are of medicinal value: Echinacea purpurea, Echinacea angustifolia and Echinacea pallida. Echinacea is widely distributed in the United States, Canada and Europe (especially Germany). In the 18th century, North American immigrants began to use echinacea, which quickly spread. It has since been used throughout the history of American medicine. In the 19th century, Dr. Merle learned herbal medicine from Native Americans and introduced echinacea to Europe. In recent years, China has successfully introduced echinacea, which is cultivated in many regions such as Shaanxi, Henan and Hunan.

Currently, most research on echinacea at home and abroad focuses on immune regulation. The whole herb is an important raw material for preparations. In China, echinacea-related preparations are also approved for use in enhancing the immune function of livestock and poultry vaccines. On November 13, 2014, the former Ministry of Agriculture approved the use of echinacea root and Echinacea root powder for enhancing the immune effect of the inactivated vaccine against porcine blue ear disease. On December 28, 2016, the former Ministry of Agriculture approved the use of echinacea granules to enhance the immune response of pigs to swine fever vaccine. Echinacea and its related preparations have a very good enhancing effect on the immune function of pigs and the immunogenicity of vaccines. Therefore, this article describes the active ingredients, pharmacological activity, safety pharmacology, toxicology and clinical application of echinacea to guide the rational use of echinacea in pigs.

1 Active ingredients

The main active ingredients of Echinacea are caffeic acid and its derivatives, alkylamide compounds, polysaccharides and glycoproteins.

Caffeic acid is one of the common phenolic acids in Echinacea. Caffeic acid derivatives (CADs) are a class of metabolites containing the hydroxyl cinnamate ester of the basic structural unit of caffeic acid. They are the main polar active ingredients of Echinacea. Currently, 15 CADs have been isolated and identified from Echinacea purpurea, including caffeic acid, chicoric acid, chlorogenic acid, echinacoside, and articholic acid. CADs have a variety of biological activities, including the inhibition of the key enzyme in bacterial infection, hyaluronidase, as well as significant immunostimulatory and anti-inflammatory effects, and they also have strong antioxidant properties [1].

Alkylamides are a large group of non-polar active ingredients in echinacea, mainly found in the roots. Currently,  more than 20 echinacoside alkylamides have been identified. These substances mainly have isobutene amides of straight-chain fatty acids with alkene and/or acetylene bonds, and the alkyl group has 4 to 5 carbon atoms. The content of alkylamides can be used as an important quality control indicator for echinacea preparations [2].

Echinacea contains a variety of macromolecules such as polysaccharides and glycoproteins. Two immunopolysaccharides, 4-methoxy-glucurono-arabino-xylan with a relative molecular weight of 35,000 and arabino-rhamno-galactan with a relative molecular weight of 50,000, were found in the aqueous extract of the aerial parts of Echinacea. Three glycoproteins with relative molecular masses of 17 000, 21 000 and 30 000 can be isolated from the roots of Echinacea purpurea. Echinacea polysaccharides and glycoproteins have strong immunological activity and anti-inflammatory, antioxidant and immune-enhancing functions [3].

In addition, Echinacea also contains essential oils, polyacetylene, flavonoids and other ingredients.

2 Pharmacological effects

Echinacea and its active ingredients have a wide range of pharmacological effects, including immunomodulatory activity, anti-inflammatory activity, antibacterial, antiviral, antitumor activity and antioxidant capacity.

2.1 Immunomodulatory activity

Echinacea has significant immunomodulatory properties and can enhance the body's immune function (both nonspecific and specific immunity). Its immunomodulatory effect is closely related to the enhancement of macrophage and lymphocyte function and stimulation of the production of cytokines and antibodies. Immunomodulatory activity is the most important biological activity of echinacea.

Echinacea can promote the growth and development of immune organs. Echinacea can effectively increase the weight of the spleen in mice, increase the level of immunoglobulins in mouse serum and reduce the level of total cholesterol, enhance resistance to pathogens, improve the immune function of mice and lower blood lipids [4].

Echinacea can enhance cellular immunity. Echinacea can significantly enhance the killing ability of natural killer (NK) cells in broiler chickens, significantly stimulate the proliferation of T lymphocytes, and promote the proliferation of peripheral blood mononuclear cells [5].

Echinacea can enhance humoral immunity. Echinacea can significantly increase antibody levels against swine fever, porcine reproductive and respiratory syndrome vaccines, and swine erysipelas [6,7], and can significantly increase antibody titers in the peripheral blood of broilers against chicken infectious bursal virus vaccine and Newcastle disease vaccine, and can significantly enhance the antibody titer against Newcastle disease virus [8,9]. In addition, Echinacea can also improve the immune response of dogs to canine distemper vaccine and canine parvovirus vaccine, and enhance antibody titers [10,11].

Echinacea can improve the immune function and biochemical indicators of blood. Echinacea can not only increase the level of serum immunoglobulin and reduce the total cholesterol level in mice, but also increase the red blood cell RBC-C3bR rosette rate and interleukin-2 and tumor necrosis factor-α content in broiler chickens, and can also improve the blood routine indicators [12,13].

2.2 Anti-inflammatory activity

The roots and above-ground parts of Echinacea purpurea contain polyunsaturated isobutylamide, which has immunomodulatory properties and can prevent pathogen-induced inflammation. It also has strong anti-inflammatory activity. In addition, echinacoside and polysaccharides in Echinacea purpurea also have immunomodulatory and anti-inflammatory activities. Echinacoside can inhibit hyaluronidase and protect collagen III from the effects of free radicals that can lead to degradation [14].

2.3 Antibacterial, antiviral and antitumor activity

Echinacea's main active ingredients, chicoric acid and caffeic acid, have significant antibacterial effects on both gram-positive and gram-negative bacteria [15]. The polysaccharides in echinacea can exert bacteriostatic and bactericidal effects by mobilizing the body's immune defense function [16]. In addition, echinacea can prevent infections caused by Candida albicans [2].

Echinacea stems and leaves and extracts exhibit antiviral activity, and the mechanism of action is related to immune stimulation and anti-hyaluronic acid. Echinacea has a certain inhibitory effect on influenza and herpes viruses, and can also inhibit the activity of hyaluronidase, thereby blocking the binding of viral receptors on the cell surface [17].

In addition, Echinacea can kill tumor cells by stimulating macrophages to produce tumor necrosis factor, interleukin, interferon, etc. [18].

2.4 Antioxidant capacity

Echinacea root and its derivatives are a good natural source of antioxidants and can be used to prevent damage caused by free radical induction. Echinacoside and polyphenols in echinacea have the ability to scavenge free radicals and purify free radicals in the body, protecting the collagen in the skin from free radical damage [19].

3 Safety pharmacology

Echinacea has no potential for undesirable effects on the cardiovascular, respiratory or central nervous systems in animals such as dogs, cats and mice at therapeutic or supratherapeutic doses.

Pang Yunlu et al. [20] studied the effects of echinacea on heart rate, electrocardiogram, blood pressure and respiration in conscious Beagle dogs. The results showed that oral administration of 0.125–1.25 g/kg Echinacea had a significant effect on heart rate in conscious beagles, but had no significant effect (P > 0.05) on the P, T, and R waves of the electrocardiogram, Q-T interval, QRS interval, P-R interval, blood pressure, or respiration. This indicates that Echinacea has no significant effect on the respiratory and cardiovascular systems of dogs, and is safe and reliable when applied at an effective dose.

Zhang Yingtao et al. [21] reported that after echinacea extract equivalent to 5, 10, or 20 g/kg of raw herb was administered to anaesthetised cats via the duodenum, there was no significant effect on blood pressure, electrocardiogram, heart rate, heart rhythm, respiratory rate and depth were not significantly affected. When echinacea herb was given to mice at a dose of 5, 10 or 20 g/kg by gavage once a day for 7 consecutive days, there was no significant effect on the mice's spontaneous activity or nervous system.

Zhang Wei et al. [22] gave healthy Kunming mice a continuous 1-week course of Echinacea root powder in different concentrations to observe the effects on the central nervous system, respiratory system and heart rate of healthy Kunming mice, in order to examine the potential adverse effects of Echinacea root powder. The results showed that Echinacea root powder had no significant effect on the central nervous system, respiratory system and heart rate of healthy Kunming mice.

4 Toxicological studies

Echinacea and its extracts are relatively non-toxic and have a large safety margin. They are a safe Chinese herbal medicine. Echinacea has no carcinogenic, teratogenic or mutagenic effects and no reproductive or genetic toxicity.

The European Medicines Agency (EMA) reported [23] that Echinacea has relatively low acute toxicity. The LD50 of fresh pressed echinacea juice is higher than the maximum recommended dose, for example, higher than 15 g/kg in rats administered orally, higher than 5 g/kg in rats administered intravenously, higher than 30 g/kg in mice administered orally, and higher than 10 g/kg in mice administered intravenously.

The EMA reported the repeated dose toxicity of freshly pressed Echinacea purpurea juice, which was administered at doses of 800, 2400 or 8000 mg per kg of body weight per day for 4 weeks. Compared with the control group, there were no changes in clinical chemistry indicators or autopsy indicators. The EMA reported that Echinacea is not mutagenic. Regarding the carcinogenicity of Echinacea, the EMA stated that considering that Echinacea is not mutagenic, it is therefore considered unnecessary to conduct carcinogenicity studies.

Zhao Dan et al. [24] studied the teratogenic effect of Echinacea powder on rats to evaluate the reproductive toxicity of Echinacea. The experiment involved the mating of sexually mature male and female SD rats in pairs. Pregnant rats were randomly divided into three dose groups: high (1 g/kg), medium (0.5 g/kg), and low (0.05 g/kg) Echinacea. A negative control group and a positive control group were also included. The test animals were given the drugs orally from gestation day 7 to 16, and the growth and development of the mother rats and fetuses were observed. The results showed that the weight gain during pregnancy, live birth weight, body length and tail length of the test animals in each dose group were significantly higher than those of the positive control group (P < 0.01), while the number of absorbed fetuses, the number of stillbirths and the malformation rate were significantly lower than those of the positive control group. There were no significant differences in all indicators between the test animal groups at each dose and the negative control group. The results showed that Echinacea purpurea powder had no maternal toxicity, embryotoxicity or teratogenic effects on rats.

Xu Peiyu et al. [25] studied the toxicity and teratogenicity of Echinacea purpurea extract on pregnant rats and rat fetuses. The study set up three dose groups based on the recommended human dose, which were 833.3 mg/kg, 1 666.6 mg/kg and 3 333.3 mg/kg, equivalent to 25, 50 and 100 times the recommended human dose, respectively, as well as a negative control group and a positive control group. The test animals were orally administered with the test substance on the 7th to 16th day of pregnancy, and the relevant indicators of pregnant rats and fetuses were examined. The results showed that the weight gain of pregnant rats in each dose group was normal, and there was no significant effect on the average weight, length and tail length of live fetuses in each litter. No external or visceral abnormalities were found, and there was no significant reduction in weight gain of pregnant rats in each dose group. Studies have shown that Echinacea has no maternal toxicity or embryotoxicity and is not teratogenic.

Zhao Dan et al. [26] evaluated the potential mutagenicity of Echinacea extract by conducting the reverse mutation test with Salmonella typhimurium in Kunming mice. The test used an echinacea extract suspension for the Ames test, and the plate infiltration method was used to count the number of revertant colonies of the TA97, TA98, TA100, and TA102 standard test strains after incubation at 37 °C for 48 h at four different concentrations. The results showed that, in both the +S9 and -S9 conditions, none of the dose groups caused a significant increase in the number of revertant colonies in the test strain, and the Ames test result was negative. This indicates that the Echinacea purpurea extract does not cause an increase in the number of revertant mutations in Salmonella typhimurium and has no genotoxicity.

Yu Meng et al. [27] studied the genotoxicity of Echinacea purpurea extract. The conventional methods of the bone marrow cell micronucleus test, mouse sperm deformity test and Ames test were used in the experiment. The results showed that there was no statistically significant difference between the Echinacea extract groups and the negative control group (P > 0.05), and there was a statistically significant difference compared with the negative control group (P < 0.01). The study concluded that Echinacea purpurea extract has no genotoxicity on mice and is not mutagenic to Salmonella typhimurium.

5 Clinical application

Ma Shaopeng et al. [7] studied the effect of Echinacea compound (Echinacea purpurea and Astragalus membranaceus) on the immune function of piglets and the vaccine against porcine reproductive and respiratory syndrome. Different doses of Echinacea compound were added to the piglets' diet to test the conversion rate of peripheral blood lymphocytes in piglets and the antibody level of the porcine reproductive and respiratory syndrome vaccine, and to study the effect of Echinacea compound preparations on the immune effect of the porcine reproductive and respiratory syndrome vaccine. The results showed that a certain amount of Echinacea compound preparation can significantly increase the conversion rate of peripheral blood lymphocytes in piglets and extremely significantly increase the antibody level of the porcine reproductive and respiratory syndrome vaccine, indicating that Echinacea has a certain promoting effect on the immune function of piglets and the effect of vaccines.

Fu Yanfang et al. [28] studied the effect of a Echinacea complex (Echinacea and Astragalus) on the immune function of piglets and the effect on the immune effect of vaccines. The experiment randomly divided three-way crossbred piglets into four groups, with the control group being the no-drug group, and test groups I, II, and III groups (Echinacea compound 1.5%, 1.0%, 0.5%), and blood was taken at 20, 35, 50, 60, 70, and 80 days of age to test the piglets' immune indicators such as the percentage of peripheral blood neutrophils, lymphocyte transformation rate, antibody levels against swine fever and porcine reproductive and respiratory syndrome. The results showed that the application of a certain amount of Echinacea compound preparation can significantly increase the percentage of peripheral blood neutrophils and lymphocyte transformation rate in piglets, and extremely significantly increase the antibody levels of swine fever and porcine reproductive and respiratory syndrome vaccines. The study concluded that Echinacea can enhance the immune function of pigs and enhance the immune effect of vaccines.

Yao Weiping et al. [29] studied the effect of adding a compound preparation of Echinacea purpurea (Echinacea and Astragalus) to feed on the growth performance and immune function of weaned piglets. The results showed that the daily weight gain, positive lymphocyte rate in serum, antibody titer against swine fever, the antibody titer of porcine reproductive and respiratory syndrome, etc., were significantly higher than those of the other groups (P < 0.05), and the diarrhea rate was significantly lower than that of the other groups (P < 0.05). The study showed that Echinacea compound can improve the production performance of piglets to a certain extent and enhance the body's immunity.

Wen Suhua et al. [30] studied the effect of Echinacea on pig performance. The drugs selected for the experiment were Zibingfen (Echinacea purpurea powder) and Kangtishu (microecological preparation). The animals selected for the experiment were healthy 3-month-old piglets. The effects of Zibingfen and Kangtishu on the detection of routine blood tests in pigs, the detection of cytokines in pigs, the antibody levels of sows against swine fever, and the reproductive performance of sows were studied. The results showed that Zi Ping Fen and Kang Ti Shu can maintain normal blood routine and serum biochemistry indicators in pigs, ensure normal basal metabolism and physiological functions, and ensure the functional state of normal pig organs. At the same time, Zibingfen and Kangsishu can improve the non-specific immunity of pigs, increase the antibody level of swine fever vaccine, and improve the reproductive performance of sows, such as shortening the farrowing time of sows, reducing the dystocia rate, significantly increasing the estrus rate 1 week after weaning, and significantly increasing the weight of weaned piglets.

Yu Xiaobin et al. [31] studied a compound biological feed additive containing Echinacea polyphenols, which was added to adult pig feed at 0.3%, and no antibiotics were added during the 32-day feeding period. The results showed that the compound biological additive containing Echinacea polyphenols reduced the feed-to-weight ratio by 12.52% and increased daily weight gain by 9.89%.

Maass et al. [6] studied the growth performance, blood picture, plasma enzymes including lymphocyte proliferation, antibody status, and immunoglobulin content in colostrum of sows, piglets, and finishing pigs after adding Echinacea as a feed additive. Alfalfa powder was used as the control in the experiment. 36 sows were given Echinacea at 0%, 1.2% or 3.6% during the 85th to 110th day of pregnancy, and at 0%, 0.5% or 1.5% from the beginning of lactation to 28 days. The results showed that Echinacea had no significant effect on the growth performance, weight loss, blood picture, plasma enzymes and colostrum composition of pigs. The health status of pigs in all treatment groups was similar. In the second trial of the study, which lasted 6 weeks and used 36 piglets, Echinacea was fed at 1.8% or 20 mg/kg, and there were no differences in the parameters evaluated.

The feed conversion rate of the Echinacea-fed group of pigs was slightly higher (about 4% higher). In the third study, 48 finishing pigs were selected for the experiment, and the test cycle was 9 weeks, divided into two stages, 1-3 weeks and 7-9 weeks. The test group was administered with 0% or 1.5% Echinacea powder or 4-6 mL of pressed juice. In the first and fifth weeks, the pigs were vaccinated against swine erysipelas to induce a specific immune response. The results showed that the feed conversion rate of the echinacea-fed group was significantly higher than that of the non-fed group, and the antibody level against swine erysipelas was significantly higher. The study concluded that echinacea can be used as a feed additive to enhance the immune effect of pig vaccines and improve feed conversion.

Zhao Xianmin et al. [32] used a lymphocyte transformation test to determine the effects of different concentrations of Echinacea polysaccharide (EPPS), Isatis indigotica polysaccharide (IRPS), Astragalus polysaccharide (APS), Chinese yam polysaccharide (CYPS) and Achyranthes bidentata polysaccharide (ABPS) were used to determine the effects of five traditional Chinese medicine polysaccharides of different concentrations (EPPS, IRPS, APS.

Xu Shouzhen et al. [33] studied the effects of various drugs, including Echinacea, on confirmed cases of infectious gastroenteritis in pigs. The study randomly selected 57 diseased pigs and divided them into 4 groups according to pen. They were treated with Astragalus polysaccharide-Echinacea injection + compound sulfadiazine injection at acupoints, with psoralen injection + florfenicol injection, with Astragalus polysaccharide injection + norfloxacin sodium solution, and with ribavirin injection + gentamicin sulfate injection. The test results showed that the treatment effect of the formula of the acupoint injection of astragalus polysaccharide-echinacea injection + compound sulfated amphotericin injection was the best. It can be seen that in clinical use, echinacea also has a certain therapeutic effect on porcine infectious gastroenteritis.

6 Conclusion

Echinacea's immunostimulatory effect on pigs is gradually being recognized. In China, the root powder of Echinacea purpurea has been approved for use in enhancing the immunogenicity of inactivated vaccines against porcine blue ear disease, and Echinacea granules have been approved for use in enhancing the immune response to swine fever vaccines. This article provides a detailed account of the active ingredients, pharmacological and toxicological effects, and clinical applications of Echinacea, with the aim of providing a reference for the rational use of Echinacea in pigs.

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