This is a short-term bacterial reverse mutation assay specifically designed to detect a wide range of chemical substances producing genetic damage leading to gene mutations.¹⁰ The test was developed by Ames and colleagues in the mid-1970s and became the most used test because of its initial promise of high qualitative (yes/no) predictivity for cancer in rodents and, by extension, in humans. The relationship between mutagenic potency prediction and quantitative carcinogenicity, however, is now known to be weak,¹¹ despite the fact that early studies with this assay indicated that greater than 90% of the known carcinogens tested were mutagenic and that 90% of the non-carcinogens tested were non-mutagenic. The power of this assay was derived from the use of a liver microsome fraction (S9 mix) containing the mixed function oxidase (cytochrome P450) enzymes required to activate the test substance into precarcinogens (as might occur in the body after phase I metabolism by the liver). As the basis of the selection of chemicals for mutagenicity testing shifted to relative environmental importance, the sensitivity of the Salmonella assay for detecting carcinogens decreased. A negative result does not imply that the chemical will be non-carcinogenic. There are a large number of false-negatives produced (i.e. non-genotoxic carcinogens).¹² Some plant components such as flavonoids give a false positive on this test.

This is the induction and quantitative measurement of chromosomal damage leading to the formation of micronuclei in cells that have been exposed to genotoxic agents or ionising radiation.

The SOS chromotest is a simple bacterial colorimetric assay for genotoxicity that may be used as a primary screening tool or as part of a battery of short-term tests for carcinogens.

Although Andrographis is widely used in infections and infestations, the most likely opinion is that its value here is mainly as an immune-enhancing treatment. Early reports in China attributed an antibacterial activity to the plant that was not supported in a later review.⁵ No direct antibacterial activity could be demonstrated for an aqueous extract of Andrographis against Salmonella, Shigella, Escherichia coli, group A streptococci and Staphylococcus aureus in vitro. Animal studies using orally administered Andrographis (0.12 to 0.24 g/kg) for 6 months failed to demonstrate bactericidal activity.¹³ Serum taken from 10 healthy volunteers after a single oral dose of Andrographis (ranging from 1 g to 6 g) showed no bactericidal activity against a number of organisms.¹³

However, an alcoholic extract of Andrographis did show significant activity against an E. coli enterotoxin-induced secretory response (that causes diarrhoea) in vivo,¹⁴ and in another study effected in vitro inhibition of adherence of Streptococcus mutans.¹⁵ Andrographolide potentiated the sensitivity of two strains of Pseudomonas aeruginosa to several antibiotic drugs in vitro.¹⁶ Significant growth inhibition (compared to five other herbs) was recently demonstrated for an Andrographis aqueous extract in vitro against Streptococcus agalactiae.¹⁷ Feeding Nile tilapia fish (Oreochromis niloticus) with Andrographis reduced mortality following infection with this species of Streptococcus (although such an outcome could also reflect enhanced immunity).

Liquid extract of Andrographis root demonstrated strong in vitro anthelmintic activity against human Ascaris lumbricoides.¹⁸ Subcutaneous administration of a decoction of Andrographis leaves to infected dogs reduced nematode larvae in the blood by 85%.¹⁹

Early uses as a substitute for quinine in malaria have been supported by recent research. An in vitro study revealed that xanthones from Andrographis root bearing a hydroxyl group at position 2 demonstrated the most potent antimalarial activity, while xanthones with a hydroxyl group at position 1, 4 or 8 possessed very low activity. Further, in vivo antimalarial testing of the most active xanthone on Swiss Albino mice with Plasmodium berghei infection demonstrated a substantial reduction (62%) in parasitaemia after treatment with 30 mg/kg.¹⁰ However, more relevant to current herbal use are studies on Andrographis leaf or whole plant. An earlier study showed that a chloroform extract of the whole plant demonstrated complete parasite growth inhibition within a 24 hour incubation period in vitro at concentrations as low as 0.05 mg/mL. There was significant reduction in mortality rates in mice administered Andrographis whole plant extract (5 mg/kg/day, ip for 4 days) just after malarial infection.²⁰ Methanolic extracts of Andrographis leaves²¹ and aerial parts²² were active in vitro against both chloroquine resistant and sensitive strains of Plasmodium falciparum.

Dehydroandrographolide succinic acid monoester (DASM), a drug derived from andrographolide, has been found to inhibit HIV in vitro. This effect was observed on several HIV strains and DASM was non-toxic to other cells in the concentration range. However, the diterpenoid lactones of Andrographis (dehydroandrographolide and andrographolide) were devoid of anti-HIV activity.²³ Moreover, in vitro studies with aqueous extracts of Andrographis showed little or no inhibition of HIV-1. Modes of inhibition studied comprised inhibition of HIV-1 protease,²⁴ inhibition of the interaction between HIV-1 gp 120 and immobilised CD4 receptor, inhibition of HIV-1 reverse transcriptase and inhibition of glycohydrolase enzymes.²⁵

Andrographolides showed in vitro activity against herpes simplex virus 1 in vitro²⁶ and both Andrographis ethanolic extract (25 μg/mL) and andrographolide (5 μg/mL) inhibited the expression of Epstein-Barr virus lytic proteins in vitro, thereby inhibiting viral maturation.²⁷ Andrographolide showed significant activity against influenza A viruses in vitro, including the H5N1 strain.²⁸ Administration of andrographolide (100 to 200 mg/kg/day, oral) to mice infected with avian influenza A strains H9N2 and H5N1 and the human strain H1N1 significantly reduced death rate, prolonged life, inhibited lung consolidation and reduced viral titres in the lung.²⁸ However, such in vivo activity might also be the consequence of immune effects.

Early research suggested an immunostimulant action for Andrographis. Enhanced phagocytosis was demonstrated in vitro for a decoction of the herb and in vivo after injection of the soluble derivatives.⁵ Isolated andrographolide (4 mg/kg/day, ip for 2 days) and an ethanolic Andrographis extract (25 mg/kg/day, oral for 7 days) significantly stimulated both antigen-specific and innate immune responses in mice. The whole extract produced stronger immunostimulation.²⁹ Prolonged survival in animals after snakebite was observed after pretreatment with extracts of Andrographis.³⁰

Four later studies (three from the same research group) have demonstrated a downregulation of immune response by andrographolide using both in vitro and in vivo models. In vitro, andrographolide reduced T cell activation in splenocytes,³¹ reduced IL-2 production in stimulated Jurkat T cells by interfering with nuclear factor of activated T cells (NFAT) and mitogen-activated protein kinases (MAPK),³² reduced IL-2 and interferon-gamma in stimulated murine T cells,³³ and downregulated macrophage immune responses and cytokine expression.³⁴ In vivo, andrographolide (4 mg/kg/day, ip) reduced T cell function and significantly reduced the severity of experimental autoimmune encephalomyelitis in mice, including antimyelin T cell and antibody responses.³¹ Andrographolide (1 mg/kg/day for 7 days, ip) also reduced antibody production and the number of IL-4-producing splenocytes in mice after antigen challenge.³⁴

However, several other later publications have observed an enhanced immune response from andrographolide or Andrographis extract, most notably the in vivo studies summarised in the Antitumour activity section that follows. In addition, andrographolide and a combination of Andrographis and Siberian ginseng extracts (Kan Jang) demonstrated lymphocyte proliferation and stimulation of some cytokines in vitro in a whole blood cell culture.³⁵ Andrographis extract (25 or 50 mg/kg/day, oral for 14 or 28 days) and andrographolide (1 or 4 mg/kg/day, oral for 14 or 28 days) enhanced specific antibody and cell-mediated immune responses in mice inoculated with an inactivated Salmonella vaccine.³⁶ A mixture of andrographolides (1.0, 1.5 and 2.5 mg/kg, oral) potentiated delayed-type hypersensitivity (DTH) in mice inoculated with sheep red blood cells, but also countered the increase in DTH after cyclophosphamide treatment.³⁷ This suggests an immunomodulatory activity. Further experimentation at the same doses revealed that the andrographolide mixture significantly stimulated phagocytic activity, white blood cell counts and spleen and thymus weights in mice.

A methanol extract of Andrographis showed potent cell differentiation-inducing activity on mouse leukaemia cells in vitro. Some of the isolated diterpenes also demonstrated this activity.^7,38 Andrographolide was shown to inhibit the in vitro proliferation of more than 30 tumour cell lines representing various types of cancers, specifically breast, CNS, colon, lung, melanoma, ovarian, prostate, renal and leukaemia. The compound was found to exert direct anticancer activity on cancer cells by cell-cycle arrest via induction of p27 and decreased expression of cyclin-dependent kinase 4 (CDK4). It also enhanced tumour necrosis factor-alpha (TNF-alpha) in lymphocytes, and possibly via this mechanism increased their cytotoxic activity against cancer cells in vitro.³⁹ Andrographolide (100 and 200 mg/kg/day for 10 days, oral) significantly inhibited the growth of B16 melanoma and HT-29 colon tumours in mice.³⁹

Since these publications there has been a considerable number of in vitro investigations of the antineoplastic activity of andrographolide against a wide variety of cancer cell lines. An extensive review of material up to 2008 by Varma and co-workers identified the key mechanisms involved.⁴⁰ These included induction of cell-cycle arrest (possibly due to increased levels of p21) and apoptosis (via a variety of mechanisms involving caspase-3, caspase-8, BcL-2 and TNF-alpha-related apoptosis inducing ligand). Other mechanisms have been proposed from recent research, including a downregulation of epidermal growth factor receptors,⁴¹ a decrease of cell-cycle related proteins,⁴² changes in the intracellular redox system⁴³ and a novel cell differentiating activity.⁴⁴ Other andrographolides have also been shown to induce cell-cycle arrest in vitro.⁴⁵

Neoandrographolide sensitised the cytotoxic action of etopiside against a leukaemia cell line⁴⁶ and andrographolide was found to sensitise cancer cells (such as colorectal, cervical and hepatic) to doxorubicin.⁴⁷

Other researchers have investigated inhibition of angiogenesis as an anticancer prospect for Andrographis extract and andrographolide. Intraperitoneal administration of both in angiogenesis-induced mice led to substantial reductions in elevated proinflammatory cytokines such as IL-1beta, IL-6, TNF-alpha and granulocyte-macrophage colony-stimulating factor (GM-CSF) and the most potent angiogenic factor, vascular endothelial growth factor (VEGF). Antiangiogenic factors such as tissue inhibitor of metalloproteinase 1 (TIMP-1) and IL-2 levels were elevated after treatment.⁴⁸ The inhibitory effect on VEGF production was supported by a later in vitro study.⁴⁹

Andrographolide inhibited the adhesion of gastric cancer cells to endothelial cells in vitro by blocking E-selectin expression in the latter.⁵⁰ Taiwanese scientists have found inhibition of migration and invasion of cancer cell lines in vitro via the downregulation of matrix metalloproteinase-7 (MMP-7)^51,52 and MMP-2.⁵³

There have also been additional in vivo anticancer studies on andrographolide and Andrographis extract (both administered intraperitoneally). A 70% ethanolic extract of Andrographis (10 mg/animal for 10 days, ip) substantially reduced tumour growth, helped maintain total white cell count, improved IL-2 and GM-CSF levels and reduced TNF-alpha in mice inoculated with Dalton’s lymphoma ascites cells.⁵⁴ These effects were maintained for 11 to 20 days after the final herb dose and were also observed for animals concurrently treated with a combination of cyclophosphamide, radiation and whole body hyperthermia. The same authors also observed that both Andrographis extract (10 mg/animal for 10 days, ip) and andrographolide (0.5 mg/animal for 10 days, ip) substantially prolonged the survival times of mice inoculated with EL4 thymoma cells.⁵⁵ IL-2 and interferon-gamma levels were increased and the authors concluded, based on this and a series of complex experiments, that the two treatments increased cytotoxic T lymphocyte activity. In general, the Andrographis extract was more active than andrographolide. The same research group using similar models and treatments has also demonstrated enhanced natural killer cell activity and antibody-dependent cytotoxicity in normal and tumour-bearing (Ehrlich ascites carcinoma) mice.⁵⁶

Andrographolide showed protective activity against chemically induced toxicity in rat hepatocytes in vitro. The observed hepatoprotective effect was greater than silymarin.⁵⁷ Intraperitoneal administration of andrographolide, andrographiside and neoandrographolide (100 mg/kg) to mice protected against hepatotoxic damage caused by carbon tetrachloride and tert-butylhydroperoxide. Andrographiside and neoandrographolide had the greatest effect on reducing lipid peroxidation and were comparable to silymarin.⁵⁸ Similar studies suggest that andrographolide is the major active antihepatotoxic principle in Andrographis.⁵⁹ Intraperitoneal administration of three diterpene constituents of Andrographis showed protective effects on hepatotoxicity induced in mice by various chemicals. The protective effect of andrographiside and neoandrographolide was as strong as silymarin, and could be due to the glucoside groups acting as strong antioxidants.⁶⁰ Andrographolide exhibited hepatoprotective activity after oral or intraperitoneal administration to rats with chemically induced acute hepatitis. Treatment with the herb led to complete normalisation of five biochemical parameters and improved histopathological changes in the liver.⁶¹

Intraperitoneal pretreatment of mice with different doses of andrographolide or arabinogalactan proteins from Andrographis for 7 days was followed by intraperitoneal injection of ethanol (7.5 g/kg of body weight). At 500 mg/kg and 125 mg/kg, respectively, the protective activity of these two preparations against hepatic and renal alcohol toxicity was comparable to silymarin.⁶²

Oral administration of Andrographis extract and andrographolide to rats demonstrated a protective action against carbon tetrachloride-induced hepatotoxicity. The leaf extract showed stronger activity than andrographolide.⁶³ Pre- and post-treatment with oral doses of Andrographis (0.5 g/kg/day) normalised alcohol-induced increases in serum transaminase activity in rats. The researchers concluded that Andrographis has a protective as well as a curative effect on alcohol-induced toxic liver damage.⁶⁴

Andrographolide (5, 7 and 10 mg/kg, oral) improved levels of antioxidant parameters such as glutathione, superoxide dismutase and catalase in mice treated with the liver carcinogen hexachlorocyclohexane (BHC).⁶⁵ It also reduced parameters of liver damage and the development of liver tumours in BHC-treated mice at the same doses.⁶⁶

Significant hepatoprotective activity was demonstrated for an alcohol extract of Andrographis and two of its diterpenes – andrographolide and neoandrographolide – against the hepatotoxicity caused by Plasmodium berghei infection in animals. The protective effect of Andrographis was thought to be partially due to reactivation of superoxide dismutase, which in turn counteracted peroxidative damage caused by the infection. Andrographis may also induce drug metabolising systems that detoxify hepatotoxins.⁶⁷ Administration of Andrographis (0.5 g/kg/day) or andrographolide (5.0 mg/kg/day) to rats for 7 to 30 consecutive days induced the liver microsomal drug-metabolising enzymes aniline hydroxylase, N-demethylase and O-demethylase.⁶⁸

Andrographolide produced a dose-dependent choleretic effect (increased bile flow, bile salt and bile acids) in rats and guinea pigs after oral administration⁶⁹ and by intraperitoneal injection in rats.⁷⁰ The effect was stronger than silymarin.⁶⁹ Aqueous extract of Andrographis orally administered to rats at 3.75 mL/kg increased bile flow and liver weight. A maximal increase in flow and weight was reached after 2 days.⁷¹

An early study investigating an aqueous extract by intravenous administration suggested that Andrographis may limit the expansion of the ischaemic focus, may exert a marked protective effect on the reversible ischaemic myocardium and could demonstrate a weak fibrinolytic action.⁷² Andrographis alleviated myocardial ischaemia-reperfusion injury in vivo.⁷³ It upregulated cellular reduced glutathione and protected cardiomyocytes against hypoxia/reoxygenation injury in vitro.⁷⁴ The mechanism was probably via a decrease in the harmful effect of oxygen free radicals.⁷⁵ A study using rabbits found Andrographis alleviated atherosclerotic arterial stenosis induced by both de-endothelialisation and a high cholesterol diet. In addition, it lowered the restenosis rate after experimental angioplasty.⁷⁶ Andrographolide (5 mg/kg, presumably ip) suppressed the hyperplasia of arterial neointima (about a 60% reduction) in a murine model of arterial restenosis. This was via the downregulation of NF-kappaB target genes that are critical in thrombosis and inflammation.⁷⁷

An aqueous extract of Andrographis given by intraperitoneal infusion to rats exhibited a dose-dependent reduction in systolic blood pressure in spontaneously hypertensive rats and normotensive controls.⁷⁸ A crude water extract of Andrographis, and two semi-purified n-butanol and aqueous fractions, significantly reduced mean arterial blood pressure in anaesthetised rats without decreasing heart rate after ip administration. The hypotensive substance in the crude water extract appeared to be concentrated in the butanol fraction.⁷⁹

Following the observation that some patients exhibited a hypotensive response while taking Andrographis, the in vitro and in vivo actions of the herb and three of its diterpenoids were investigated.⁸⁰ The diterpenoid 14-deoxy-11,12-didehydroandrographolide (DIAP) was most active at reducing the chronotropic response of isolated rat atria and exerting spasmolytic activity in rat aortic rings. An Andrographis aqueous extract with the highest levels of DIAP (delivering 19 mg/kg of this compound, oral doses for 7 days) was the most potent (compared to two other extracts with lower levels of DIAP) at reducing systolic blood pressure in rats. Mechanistic studies suggested that vascular smooth muscle was the major site of the hypotensive effect.

Andrographolide inhibited PAF-induced human platelet aggregation⁸¹ and deoxyandrographolide antagonised PAF-mediated processes in neutrophils,⁸² as did andrographolide,⁸³ all in vitro.

Several early in vivo studies found antipyretic and anti-inflammatory effects for andrographolides (after oral administration or injection). The anti-inflammatory activity of the andrographolides may be due to the promotion of ACTH (adrenocorticotrophic hormone) and consequent enhancement of adrenocortical function.⁵ Andrographolide administered orally (30, 100 and 300 mg/kg) significantly reduced inflammation in a number of animal models including adjuvant-induced arthritis.⁸⁴ The addition of andrographolide to an endothelial cell culture together with TNF-alpha caused a concentration-dependent reduction of the enhancement of endothelial monocyte adhesion, which is part of the inflammatory process.⁸⁵ Another in vitro study found that andrographolide inhibited NF-kappaB binding to DNA, thereby reducing the expression of pro-inflammatory proteins such as COX-2 in neutrophils.⁸³ Andrographolide prevented oxygen radical production by human neutrophils in vitro.⁸⁶

As well as demonstrating antioxidant activity in vitro and in vivo, a 70% methanolic extract of Andrographis (10 mg/animal for 5 days, ip) completely inhibited carrageenan-induced paw oedema in mice.⁸⁷ Oral administration of neoandrographolide (150 mg/kg) also demonstrated anti-inflammatory activity in mice as well as in vitro, using several experimental models.⁸⁸

In a murine model of asthma, andrographolides (30 mg/kg, ip) inhibited the elevation of bronchoalveolar fluid (BAF) levels of TNF-alpha and GM-CSF, and almost abolished the accumulation in BAF of lymphocytes and eosinophils.⁸⁹ In a similar model, andrographolide (0.1, 0.5 and 1 mg/kg, ip) dose-dependently inhibited increases in total cell count, eosinophil count and IL-4, IL-5 and IL-13 levels in BAF.⁹⁰ It also attenuated IgE responses, eosinophilia and airway mucus production and hyper-responsiveness. Examination of lung tissue specimens and further in vitro investigations suggested that andrographolide might act by inhibiting the NF-kappaB pathway. An anti-inflammatory mechanism mediated by reduced NF-kappaB expression was also observed for andrographolide (2 mg/kg/day for 7 days, ip) in another study in mice, using a similar experimental model of asthma.⁹¹

Oral doses of andrographolide at 300 mg/kg demonstrated analgesic activity; at 100 and 300 mg/kg significant antipyretic effects were also observed after 3 h. In addition, this dose exhibited significant protective activity against aspirin-induced ulceration in rats.⁹² An aqueous extract of Andrographis (40 and 100 mg/kg, oral) and andrographolide (25, 50 and 100 mg/kg, oral), but not a 95% ethanolic extract (100 and 200 mg/kg, oral), demonstrated significant analgesic activity in mice.⁹³ The aqueous extract and andrographolide were also active at oral doses of 100 mg/kg in reducing carrageenan-induced rat paw oedema. A similar activity profile (analgesic and antioedema) was demonstrated for subcutaneous injection of andrographolide (10, 25 and 50 mg/kg).⁹⁴ Analgesia was probably mediated via non-opioid pathways, since naloxone failed to antagonise the activity of andrographolide. A methanolic extract of Andrographis (100 to 300 mg/kg, ip) slightly lowered body temperature, increased pentobarbitone sleeping time, demonstrated analgesic activity, reduced exploratory behaviour and curiosity, and exhibited some muscle-relaxing activity in mice.⁹⁵ The same doses in rats also reduced exploratory behaviour in the Y-maze test.

An aqueous extract of Andrographis (10 mg/kg) was found to prevent glucose-induced hyperglycaemia in rabbits, but failed to prevent glucose absorption from the gut.⁹⁶ In a screening of several traditional remedies, only Andrographis (as aqueous extract, and especially freeze-dried extract, administered at 50 mg/kg and 6.25 mg/kg body weight) significantly lowered blood glucose levels in streptozotocin-induced hyperglycaemic rats.⁹⁷ In a further study Andrographis decoction was orally administered to alloxan-induced diabetic rats, with a significant reduction in blood glucose levels observed compared with controls.⁹⁸

In a study comparing normal and streptozotocin-induced diabetic rats, an ethanolic extract of Andrographis not only demonstrated hypoglycaemic effects, but also reduced oxidative stress in the diabetic rats. Normal and diabetic rats were randomly divided into groups and treated orally with distilled water, metformin (500 mg/kg) or Andrographis (400 mg/kg) twice daily for 14 days. Both Andrographis and metformin significantly increased body weight and reduced fasting serum glucose in the diabetic rats, but had no such effects in the normal rats. Both treatments also significantly increased the activity of the antioxidant enzymes superoxide dismutase (SOD) and catalase in the diabetic rats, but again not in the normal rats.⁹⁹ In another study, Andrographis extract (400 mg/kg, oral) decreased blood glucose and increase activities of the antioxidant enzymes SOD and catalase in streptozotocin-induced diabetic rats.¹⁰⁰

Inhibition of the digestive enzymes alpha-glucosidase and alpha-amylase can significantly decrease the postprandial increase in blood glucose. In vitro testing demonstrated that a 20% ethanolic extract of Andrographis possessed an appreciable alpha-glucosidase inhibitory effect, but demonstrated only weak inhibition of alpha-amylase.¹⁰¹ Supporting this observation, a single oral dose of Andrographis extract (250, 500 or 1000 mg) dose-dependently and significantly reduced blood glucose in streptozotocin-induced diabetic rats challenged with starch and sucrose, but not after a glucose challenge.¹⁰¹ In contrast, andrographolide (0.5 to 1.5 mg/kg, oral) dose-dependently decreased plasma glucose in streptozotocin-induced diabetic rats, and at 1.5 mg/kg (oral) significantly attenuated plasma glucose after glucose challenge in normal rats.¹⁰²

As a possible contradiction of the above results for sucrose, oral administration of Andrographis extract and andrographolide produced a dose- and time-dependent activation of brush-border membrane-bound hydrolases (lactase, maltase, sucrase) in rats, suggesting it accelerated the intestinal digestion and absorption of disaccharides.¹⁰³

An aqueous extract of Andrographis (200 mg/kg, oral) largely countered the nephrotoxic impact of gentamicin in rats, in terms of tending to normalise serum creatinine and urea, blood urea nitrogen and urine volume.¹⁰⁴ Whether this effect was due to antioxidant activity or represented a specific renoprotective activity is not clear.

Three studies cited above attest to significant in vivo and in vitro antioxidant activity for the herb,^87,93,100 as do some of the hepatoprotection studies. Oral administration of an aqueous extract of Andrographis (10, 20 and 30 mg per mouse) caused a significant elevation of catalase, SOD and glutathione-S-transferase activities in lymphoma-bearing mice, as well as exhibiting some antitumour activity.¹⁰⁵

Other studies have demonstrated protection against toxins, in some cases linked to antioxidant activity. Andrographolide and an aqueous extract of Andrographis (250 mg/kg/day for 7 days, ip) protected against nicotine-induced neurotoxicity in rats by reducing oxidative stress.¹⁰⁶ A similar protective activity against nicotine-induced oxidative stress was demonstrated in vitro for lymphocytes.¹⁰⁷

A 70% ethanolic extract of Andrographis (10 mg/animal/day for 10 days, ip) was also shown to protect against cyclophosphamide toxicity in mice in two separate but similar studies conducted by the same research group.^108,109 The elevation of TNF-alpha induced by the drug was lowered by Andrographis treatment. Andrographolide was also active.¹⁰⁹

Topical application of Andrographis extract (10%) improved wound healing in rats.¹¹⁰ Wounds dressed with Andrographis showed markedly less scar width, higher fibroblast proliferation, more collagen, less angiogenesis and an absence of inflammatory cells.

A recent systematic review of two systematic reviews and eight clinical trials concluded that there was evidence that Andrographis was useful in the treatment of upper respiratory tract infections, but expressed concerns about publication bias and particularly the fact that most of the trials had been conducted in association with product manufacturers.¹²² The authors called for more independent clinical trials.

The two systematic reviews included in the above were as follows. In one, seven double blind, controlled trials (n=896) that met inclusion criteria for evaluation of efficacy were considered. All trials scored at least three (out of a maximum of five) for methodological quality on the Jadad scale. Collectively, the data suggested that Andrographis was superior to placebo in alleviating the subjective symptoms of uncomplicated upper respiratory tract infection. There was also preliminary evidence of a preventative effect. Adverse events reported following the herb administration were generally mild and infrequent.¹²³ In the second review, 433 patients from three trials were included in the meta-analysis. Andrographis either alone or in combination with Siberian ginseng was more effective than placebo in the treatment of uncomplicated acute upper respiratory tract infection.¹²⁴

In a general review of the literature for evidence of the efficacy and safety of complementary and alternative medicine for the prevention and treatment of upper respiratory tract infection in children, the authors concluded that Andrographis decreased nasal secretions (p<0.01), but not other symptoms.¹²⁵

The subjects of these reviews as well as other studies follow.

Uncontrolled early Chinese clinical studies in patients with bacterial and viral respiratory infections suggested beneficial effects after oral administration of Andrographis or andrographolides, implying an immune enhancing action.⁵ Investigations from the Sichuan Traditional Medicine Research Institution found Andrographis exerted a beneficial effect in the treatment of infectious diseases associated with cold symptoms. The major finding was the lowered body temperature within 48 h after treatment with Andrographis. Of 84 cases of common cold, 70 achieved normal body temperature within 48 h.¹²⁶

A randomised double blind study of 152 patients with pharyngotonsillitis found Andrographis (6 g/day) for 1 week to be as effective as paracetamol (acetaminophen) in relieving fever and sore throat. For both groups the difference between baseline symptoms and final evaluation was significant (p<0.0001). Lower doses of Andrographis were not as effective.¹²⁷

Tablets containing a total of 1200 mg Andrographis extract (standardised to 4% andrographolides) or placebo were given to 61 patients suffering symptoms of common cold in a double blind, placebo-controlled clinical trial. After 4 days of treatment, measured symptoms were significantly reduced in the Andrographis-treated group compared to placebo: strength of disease (p=0.0001), tiredness (p=0.0001), sweating/shivering (p=0.001), sore throat (p=0.0001) and muscular ache (p=0.0001). In terms of clinical signs (rhinitis, sinus pains and headaches, lymphatic swellings), there was no significant difference between the treated and placebo groups at day 4. However, when the groups were compared over time (specifically day 0 versus day 4), there was a significant decrease in the intensity of these signs only for the Andrographis group (p<0.05). The overall reduction in the symptom score over time was also significant (p<0.01). The authors concluded that, based on their findings, Andrographis can significantly reduce the symptoms and duration of the common cold.¹²⁸

In a randomised, double blind, placebo-controlled clinical trial, 107 healthy children received either Andrographis extract tablets (200 mg/day of extract, standardised to 11.2 mg andrographolide) or placebo for 3 months during the winter season. This dose corresponds to about 1 g of original herb. Analysis after the first month indicated no significant change for Andrographis treatment. However, by the third month there was a significant decrease in the incidence of colds compared with placebo (30% versus 62%; p<0.01). The relative risk of catching a cold was 2.1 times lower for the Andrographis group.¹²⁹ The same research team conducted a further randomised, placebo-controlled, double blind study of an Andrographis extract at 1200 mg/day over 5 days in 158 adult patients. Visual analogue scale evaluations of the intensity of headache, tiredness, earache, sleeplessness, sore throat, nasal secretion, phlegm and frequency and intensity of cough were performed by the patients at days 0, 2 and 4 of the treatment. Using a logistic regression model of assessment, there was a significant decrease in the intensity of the symptoms of tiredness, sleeplessness, sore throat and nasal secretion in the Andrographis group at day 2, as compared with the placebo group. By day 4, a significant decrease in the intensity of all symptoms was observed for the Andrographis group. No adverse effects were observed or reported.¹³⁰

In another randomised, double blind, placebo-controlled pilot study, 50 outpatients with symptoms of common cold were treated with tablets containing Andrographis extract (1020 mg/day, about 6 g of herb). The patients were advised to make their first clinic visit not later than 3 days after the occurrence of cold symptoms. After 5 days of therapy, subjective evaluation demonstrated a significantly reduced number of sick leave days (p<0.03), improved symptoms (p<0.025) and hastened recovery (p<0.05). Side effects were very few and mild.¹³¹

Recently, a randomised, double blind, placebo-controlled clinical trial observed that treatment with a standardised extract of Andrographis reduced the symptoms of uncomplicated upper respiratory tract infection.¹³² A total of 223 patients received either 200 mg/day of an Andrographis extract (about 2.5 g of herb, containing 60 mg of andrographolides) or a matching placebo for 5 days after experiencing the typical symptoms of a common cold. Nine self-evaluated symptoms were used to assess the efficacy of the herbal treatment: cough, expectoration, nasal discharge, headache, fever, sore throat, earache, malaise/fatigue and sleep disturbance. Both groups showed improvement in these scores from days 1 to 3. However, from days 3 to 5 most of the symptoms in the placebo group were unchanged, whereas symptom improvement continued for the Andrographis group. The difference in the overall symptom score between the two groups was significant at day 5 (p<0.05). For individual symptoms on day 5, all were significantly improved for the Andrographis group versus placebo (p<0.05), except for earache. The overall efficacy of Andrographis was a significant 2.1 times higher than placebo (p<0.05), and the herbal treatment was well tolerated. One weakness of the trial design was that patients were not treated for longer than 5 days. Hence the impact of Andrographis on shortening the duration of the common cold could not be assessed.

In two randomised, parallel-group clinical studies, a standardised extract of Andrographis (as the combination Kan Jang) was compared with amantadine in the treatment of diagnosed viral influenza infection.¹³³ Each tablet comprised 85 mg of Andrographis extract (containing 5 mg andrographolides) and 10 mg of Siberian ginseng extract (from 120 mg root). The typical acute dose was four tablets three times daily. In the first pilot study, 71 Kan Jang-treated patients were compared with 469 patients on amantadine; in the second phase 66 patients were enrolled. Duration of sick leave and frequency of post-influenza complications were used as outcome measures and indicated that the herbal combination contributed to a quicker recovery. It reduced the risk of post-influenza complications and was also well tolerated.

A three-arm study compared the efficacy of standard treatment, Kan Jang and a preparation containing Echinacea purpurea extract in patients with uncomplicated common colds.¹³⁴ Of the 130 children aged between 4 and 11 years studied over a period of 10 days, 39 patients received only standard treatment, 53 were treated with the Andrographis combination plus standard treatment and 41 were treated with Echinacea plus standard treatment. It was found that adjuvant treatment with the Andrographis combination was significantly more effective than the Echinacea preparation when started at an early stage of uncomplicated common colds. The effect was particularly pronounced in terms of the amount of nasal secretion and congestion. It also accelerated the recovery time compared to other treatments. The need for standard medication was significantly less in the Andrographis combination group compared with the others and it was well tolerated, with no adverse reactions reported.

The same Andrographis combination was also tested in a phase III randomised, double blind, placebo-controlled parallel group clinical trial in the treatment of uncomplicated upperrespiratory tract infections. After an initial pilot trial involving 46 patients over 3 to 8 days, 179 patients completed the 3-day study according to protocol. Both the total symptom score (from patients’ evaluation) and the total diagnosis score (from physicians’ evaluation) showed highly significant improvements (p<0.0006 and p=0.003, respectively), as compared with placebo. Throat signs and symptoms demonstrated the most significant improvement.¹³⁵

A double blind, placebo-controlled clinical study evaluated the impact of Kan Jang treatment for 5 days in the management of acute upper respiratory symptoms (including sinusitis) in 185 patients. At the end of the treatment, significant differences compared with placebo were in evidence for total symptoms in the group as a whole (p<0.001) and in the acute and recurrent sinusitis subgroups. In terms of individual symptoms in the whole group, significant differences against placebo (p<0.001) were observed for sore throat, headache, malaise and catarrh.¹³⁶

Many early Chinese studies used oral administration of Andrographis or andrographolides in acute bacillary dysentery and enteritis and observed a marked benefit.⁵ Patients with acute diarrhoea were treated with powdered leaves and stems of Andrographis. The Andrographis was more effective in reducing the number of Shigella, but was less effective for cholera compared with tetracycline. Oral administration of 1 g every 12 h for 2 days was more effective than giving a dose of 500 mg every 6 h for 2 days.¹³⁷

Familial Mediterranean fever (FMF) is a recessively inherited inflammatory disorder characterised by recurrent attacks of fever and serositis (inflammation of the serous tissues such as pleura, pericardium and peritoneum). A combination of Andrographis extract 600 mg/day (containing 48 mg andrographolide), Siberian ginseng extract 120 mg/day (standardised to >9.6 mg eleutheroside E), Schisandra chinensis extract 600 mg/day (standardised to >9.6 mg schisandrins) and licorice extract 120 mg/day (standardised to >7.2 mg glycyrrhizin) was assessed over 30 days in a pilot study involving 24 children with FMF.¹³⁸ Using a double blind, placebo-controlled design, it was found that duration, frequency and severity of attacks were all significantly less compared with placebo following the herbal treatment (p=0.0003). A separate publication investigated the impact of the herbal combination on plasma nitric oxide (NO) levels during the trial.¹³⁹ Additional control groups were used, consisting of healthy volunteers and FMF patients treated with colchicine. Basal levels of NO in FMF patients during attack-free periods over the 30-day trial were found to be no different to the healthy controls. Surprisingly, NO levels fell during attacks. The herbal formulation with Andrographis was found to normalise blood levels of NO and decrease IL-6 in FMF patients during attacks.

A 14-week randomised, double blind, placebo-controlled clinical trial in 60 patients examined the impact of a 75% ethanolic extract of Andrographis (300 mg/day corresponding to 3 g of herb and containing 90 mg of andrographolides) in active rheumatoid arthritis.¹⁴⁰ All trial patients were given methotrexate and were allowed to take prednisone or chloroquine in stable doses if already prescribed. Compared with baseline there were significant improvements observed in the Andrographis group by week 14 for tender joints (p=0.001), number of swollen joints (p=0.02), severity of swollen joints (p=0.01), severity of tender joints (p=0.002), levels of rheumatoid factor (p=0.01) and quality of life measures (p<0.001). However, compared with the placebo group these changes were not statistically significant. Perhaps a larger trial with a higher dose might have yielded results that were more definite.

A randomised, double blind trial was conducted at five centres in Shanghai to compare a standardised extract of Andrographis with the non-steroidal anti-inflammatory drug mesalazine (4.5 g/day, in slow release form) in patients with mildly to moderately active ulcerative colitis (confirmed by colonoscopy).¹⁴¹ One hundred and eight patients completed the trial. The aqueous-ethanolic extract provided about 108 mg/day of andrographolide. Treatment with Andrographis extract demonstrated similar efficacy to mesalazine. Scores for clinical symptoms (fever, stool frequency, stool consistency, stool blood, abdominal pain, mucous stool, tenesmus (straining) and abdominal tension) were assessed throughout treatment. Symptom scores decreased over time in both groups. Clinical efficacy was also assessed by the percentage of patients attaining remission, partial remission or improvement in symptoms. Mucosal healing was evaluated by colonoscopy and, in the 34 patients with biopsies available, histopathology was evaluated. Such outcomes were significantly better (p<0.001) for both groups compared with baseline, and there was no significant difference between the two treatment groups. Thirteen per cent of patients in the Andrographis group and 27% of patients in the mesalazine group had at least one adverse event. Most adverse events appeared to be related to the underlying disease.

In a phase I clinical trial, 13 HIV positive patients and 5 healthy volunteers took 5 mg/kg andrographolide for 3 weeks, escalating to 10 mg/kg for 3 weeks, which was then intended to rise to 20 mg/kg for a final 3 weeks. However, the trial was interrupted at 6 weeks due to adverse events, including an anaphylactic reaction in one patient. All adverse events had resolved by the end of observation. A significant rise in the mean CD4+ lymphocyte level in HIV patients occurred after administration of 10 mg/kg andrographolide (from 405 to 501 cells/mm³, p=0.002). There were no statistically significant changes in mean plasma HIV-1 RNA levels throughout the trial.¹⁴²

An open study in Thailand compared parameters of urinary tract infection in patients undergoing shock wave dissolution of kidney stones (lithotripsy). The study found that 1 g of Andrographis was as effective as the antibiotics co-trimoxazole and norfloxacin in reducing pyuria and haematuria.¹⁴³

A phase I clinical study of Kan Jang in healthy men revealed a slightly positive benefit on sperm count, sperm activity and other indices of fertility when it was taken at three times the normal dose.¹⁴⁴

Sixty-three patients with cardiac and cerebrovascular diseases were observed at 3 h and/or 1 week after taking Andrographis extract. Results showed that platelet aggregation induced by ADP was significantly inhibited (p<0.001). The aggregation rate was lower at 1 week. In other volunteers taking Andrographis, serotonin release from platelets was decreased (p<0.01), but plasma serotonin levels remained unchanged. A rise in platelet cAMP levels might be the mechanism behind the antiplatelet activity of Andrographis.¹⁴⁵

A majority of 20 patients with infective hepatitis showed marked improvement in symptoms after approximately 24 days of treatment with an Andrographis decoction (equivalent to 40 g of herb per day). Significant decreases in various liver function tests were also observed. Overall, 80% of cases were ‘cured’ and 20% were relieved.¹⁴⁶

The following LD₅₀ data have been recorded for Andrographis extract and its constituents:

In acute toxicity studies, no toxic effects were observed in mice after oral administration of a suspension of Andrographis leaf powder (2 g/kg), a suspension of leaf alcohol extract (2.4 g/kg) or andrographolide (3 g/kg).¹⁴⁹ Similarly, subcutaneous administration of Andrographis leaf decoction (0.33 g/kg) to rabbits did not exhibit toxic effects.¹⁹ Female rats treated for 14 days with an oral dose of 5 g/kg of a methanolic Andrographis extract also demonstrated no adverse effects.¹⁵⁰

No toxic effects were observed in subacute oral toxicity tests when either a leaf powder suspension (200 and 400 mg/kg) or straight leaf powder (50 to 150 mg/kg) was administered on alternate days for 4 weeks to mice or for 14 weeks to rats, respectively.¹⁴⁹ Also andrographolide (1 g/kg/day) administered to rats and rabbits for 7 days did not cause toxic effects.⁵ Rats administered Andrographis powder (part and route not specified) at dosages of 0.12, 1.2 and 2.4 g/kg/day for 6 months exhibited no abnormalities in growth rate, food consumption, clinical signs, serum biochemical parameters or histology.¹⁴⁸

Oral administration of andrographolide (2 g/day) for 4 days caused a transient elevation of the liver enzyme ALT in healthy volunteers. Levels normalised upon discontinuation of the andrographolide. Hepatic and renal functions were not impaired after doses of 0.9 g/day for 5 days.⁵

Effects of Andrographis on male fertility show conflicting results, but there is probably no cause for concern based on the more recent studies. Reduced fertility and prolongation of gestation were observed in mice when the male was fed Andrographis stem powder (0.75% of diet) prior to mating. As mating rates were not confirmed, these effects may have been due to a reduction in libido. Treated females mated with untreated males showed no appreciable change in fertility or gestational period.¹⁵¹ In contrast, antifertility effects were not observed in mice fed the powdered leaf or root (1% of diet; approximately 2 g/kg/day) for 2 weeks prior to mating and for 3 weeks during mating.^151,152

Oral administration of Andrographis leaf powder (50 and 100 mg/kg/day) for 24 to 60 days to male rats resulted in the cessation of spermatogenesis and biochemical and degenerative changes in the testes and male accessory organs.^153,154 Decreased sperm counts, spermatozoa abnormalities, histopathological changes in the testes and lack of fertility were observed after oral administration of high doses of andrographolide (25 and 50 mg/kg) for 48 days.¹⁵⁵ However, no significant differences were observed in reproductive organ weights, testicular histology or serum testosterone levels after oral administration of an Andrographis dried herb (5:1) ethanol extract (containing 5.6% andrographolide) at dosages of 20, 200 and 1000 mg/kg/day for 60 days. The authors concluded that the above variation in results might be due to differences in the preparation of the plant material used.¹⁵⁶

Recently Andrographis extract (20, 200 and 1000 mg/kg/day orally for 65 days)¹⁵⁷ and andrographolide (50 mg/kg/day for 2 to 8 weeks)¹⁵⁸ demonstrated no significant effects on sperm morphology, motility and counts and were without significant adverse effects. There is also the phase I clinical study in men cited earlier.¹⁴⁴

After nitrosation with nitrite under acidic conditions, an ethanol extract of Andrographis became mutagenic to strains TA 98 and TA 100 (Salmonella/microsome test) tested either in the presence or absence of S-9 mix.¹⁵⁹ A methanolic extract of Andrographis was devoid of significant genotoxic effects in three different in vitro models.¹⁵⁰

Like parthenolide from feverfew, helenalin and certain other SLs in Arnica exert potent anti-inflammatory activity in vitro by inhibiting the activation of the transcription factor nuclear factor (NF)-kappaB. This was first demonstrated in 1997 and found to be quite selective, as the activity of four other transcription factors was not affected.²⁶ Follow-up research indicated that helenalin acts by selectively alkylating the p65 subunit of NF-kappaB, rather than by inhibiting the degradation of IkappaB.²⁷ (NF-kappaB, composed of a p50 and a p65 subunit, is retained in an inactive cytoplasmic complex by binding to a third (inhibitory) subunit IkappaB. Degradation of IkappaB results in its activation.)

Inhibition of NF-kappaB has also been demonstrated in vitro for various extracts of Arnica species, including Arnica tincture.^28–30 Consistent with inhibition of NF-kappaB activation, an Arnica extract inhibited nitric oxide production and cyclo-oxygenase (COX)-2 activity in activated murine macrophages.³¹

Other related anti-inflammatory activity has been found for the SLs in Arnica. The transcription factor nuclear factor of activated T cells (NFAT) is also inhibited in vitro by Arnica extract.³⁰ Suppression of matrix metalloproteinase (MMP)-1 and MMP-13 mRNA levels in bovine and human articular chondrocytes was also observed.³² This latter effect may be due to inhibition of activation of the transcription factors activator protein (AP)-1 and NF-kappaB, and implies possible activity in osteoarthritis.

Arnica SLs have also demonstrated additional anti-inflammatory mechanisms in vitro. Two derivatives of 11alpha, 13-dihydrohelenalin inhibited the stimulated release of the pro-inflammatory enzyme neutrophil elastase from human neutrophils.³³ Helenalin provoked irreversible inhibition of leukotriene C4 (LTC4) synthase in human platelets and inhibited both 5-lipoxygenase and LTC4 synthase in human granulocytes.³⁴ Polymorphonuclear neutrophil chemotaxis was inhibited at low concentrations, whereas prostaglandin synthetase activity was inhibited at a higher concentration by a series of sesquiterpene lactones, including helenalin and dihydrohelenalin.³⁵

Helenalin also appears to modulate immune activity in vitro. It induced apoptosis in activated CD4+ T cells by triggering the mitochondrial pathway of apoptosis, or otherwise inhibiting their proliferation.³⁶ As previously demonstrated, activation of NFAT (specifically NFATc2) was also found.

Additional results from in vitro studies suggest that the mechanism of the anti-inflammatory activity of Arnica and its constituents also possibly occurs at several other sites:³⁵

Several sesquiterpene lactones, including those from Arnica, have demonstrated anti-inflammatory activity by intraperitoneal injection (2.5 mg/kg/day) in animal models such as carrageenan-induced paw oedema and chronic adjuvant arthritis. The alpha-methylene-gamma-lactone structure was required for inhibitory activity in both models and the 6-hydroxy group of helenalin was required for potency in the former model. Inhibition of writhing reflex (an indication of analgesic activity) was also demonstrated.³⁷

Topical Arnica was shown to be an effective anti-inflammatory in a rat model of acute muscle damage.³⁸ The Arnica gel (containing 200 mg/g of an unspecified Arnica tincture) was either applied by massage or ultrasound, with the latter failing to exert an anti-inflammatory effect above the control group.

Components of the essential oil of Arnica have demonstrated potent activity against Gram-positive and Gram-negative bacteria and against Candida spp. in vitro.³⁹ Helenalin and helenalin acetate were also active in vitro against Gram-positive bacteria and Proteus vulgaris.⁴⁰ The polyacetylenes have shown broad-range antimicrobial activity against pathogenic fungi (Trichophyton spp., Microsporum gypseum, Epidermophyton spp.) and bacteria (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli) in vitro.⁴¹ However, compared with propolis, an Arnica extract showed little antibacterial activity against oral pathogens.⁴² Some modest activity for Arnica was demonstrated against periodontal bacteria.⁴³ SLs including helenalin were also quite active against Leishmania mexicana with IC50 values of 2 to 4 μM.⁴⁴

Helenalin and related sesquiterpene lactones have anti-trypanosomal activity (with an IC50 as low as 0.05 μM for helenalin)⁴⁵ and the pseudoguaianolide SLs show high activity against asexual blood forms of Plasmodium falciparum.⁴⁶

Staph. aureus is a major udder pathogen causing bovine mastitis. Some pro-inflammatory cytokines, including tumour necrosis factor (TNF)-alpha, enhance its growth. Helenalin markedly reduced the growth of Staph. aureus in the presence of TNF-alpha in bovine mammary epithelial cells.⁴⁷ Helenalin also reduced Staph. aureus infection and associated inflammation in a murine model of mastitis following ip administration (20 mg/kg).

Like parthenolide from feverfew, helenalin has exhibited antitumour activity in vitro, although it has been less extensively studied. In a screening of 21 flavonoids and five SLs from Arnica species, helenalin demonstrated the strongest cytotoxic activity of the SLs when tested on lung carcinoma cells. The flavonoids showed moderate to low cytotoxicity compared with the reference compound.⁴⁸

Helenalin strongly induced apoptosis in leukaemia Jurkat T cells,⁴⁹ even if they lacked the CD95 death receptor or over-expressed antiapoptotic proteins.⁵⁰ It probably acts by inducing a mitochondria-dependent pathway. The cytotoxicity of helenalin against KB tumour cells was decreased by cysteine, but the cytotoxicity of 11alpha,13-dihydrohelenalin was not affected.⁵¹

The effect of flavones and flavonols from Arnica spp. on the cytotoxicity of helenalin was investigated in the human lung carcinoma cell line. At non-toxic concentrations, all flavonoids (except kaempferol) significantly reduced helenalin-induced cytotoxicity.⁵²

Arnica SLs inhibited Walker 26 carcinosarcoma and Ehrlich ascites tumour growth in vivo (2.5 mg/kg day for rats and 25 mg/kg/day for mice, both ip).⁵³

The stabilisation of the lysosomal membrane in liver cells by SLs was dependent on the alpha-methylene-gamma-lactone structure.³⁵

Helenalin and 11alpha,13-dihydrohelenalin in vitro have inhibited human platelet function via thiol-dependent pathways. The reduction in platelet sulphydryl groups is probably associated with reduced phospholipase A₂ activity.⁵⁴

Helenalin, dihydrohelenalin and epoxyhelenalin (20 mg/kg/day in mice) produced a lowering of serum cholesterol (30%) and serum triglycerides (25%). Thiol-bearing enzymes of lipid synthesis were inhibited by these compounds in vitro, suggesting they alkylate thiol nucleophiles by a Michael-type addition.⁵⁵

Astragalus markedly enhanced the cytotoxicity of natural killer cells,¹³ potentiated interleukin-2-generated LAK (lymphokine-activated killer) cell cytotoxicity manifested by tumour cell lysis¹⁴ and reversed tumour-associated macrophage suppression in urological tumours,¹⁵ all in vitro. Using an in vitro local graft-versus-host reaction as a test assay for T-cell function, Astragalus extract restored the reaction in cells taken from 9 of 10 cancer patients.¹⁶ Saponins from Astragalus stimulated the natural killer (NK) cell activity of human peripheral blood lymphocytes and restored steroid-inhibited NK cell activity, both in vitro.¹⁷ Astragalus saponins reduced nicotinic acetylcholine receptor antibodies in blood cell cultures from myasthenia gravis patients.¹⁸

More recently, Astragalus was found in vitro to correct the immunological dysfunction in peripheral dendritic cells taken from children with Henoch–Schönlein purpura¹⁹ and modified responses from lipopolysaccharide-stimulated macrophages, reducing cytokines in a dose-dependent manner.²⁰ Astragalosides significantly increased phagocytic activity against Mycobacterium tuberculosis in vitro.²¹ Peripheral blood mononuclear cells (PBMCs) from 27 children with recurrent tonsillitis were stimulated and cultured with Astragalus in vitro for 48 hours.²² Astragalus improved interferon-gamma output. Similar results were seen for Astragalus in PBMCs from 15 asthma patients, suggesting a reversal of Th2 predominant status.²³

Oral doses of Astragalus in mice (200 mg/kg) enhanced several aspects of immunity, including increased thymus weight, potentiation of phagocytic function, superoxide anion production by peritoneal macrophages and proliferation of splenocytes.²⁴ Protective effects on immune suppression in mice were observed after co-administration of Astragalus with a carcinogen. Macrophage numbers and white cell function were raised to the same as or greater than normal levels.²⁵ Co-administration of Astragalus with an antitumour agent resulted in protection against the immunosuppression induced by the antitumour agent.²⁶ Oral administration of Astragalus (5 g/kg/day for 7 days) increased phagocytic activity²⁷ and significantly increased the lymphocyte transformation rate in a suppressed cellular immunity model.²⁸ Astragalus promoted interleukin-2 production in splenic lymphocytes of blood-deficient mice.²⁹ A protective effect of Astragalus extract after oral administration against Japanese encephalitis virus infection in mice was demonstrated. The authors proposed that the protective effect of Astragalus is based on a non-specific mechanism during the early stage of infection, before shifting to antibody production, and that macrophages play an important role by inducing the production of active oxygen.³⁰

Astragaloside IV (50 mg/kg/day, iv) mitigated the development of the characteristic features of ovalbumin-induced chronic experimental asthma in mice.³¹ Astragalus saponins (400 mg/kg, oral) exerted a protective effect against microbial sepsis in mice.³² An ethanolic extract of Astragalus modified Th1/Th2 cytokine secretion patterns in vitro and in vivo (1.25 g/kg/day, oral for 7 days), favouring Th2 responses.³³ In contrast, oral doses of Astragalus attenuated the expression of IgA nephropathy in rats, possibly by diminishing Th2 cytokine responses.³⁴ Astragalus improved the phagocytic activity of peritoneal macrophages in mice.³⁵ The effect of ip injection was higher than for oral doses, but the difference was not significant. The immune response of carp (including phagocytic activity) was enhanced after including Astragalus at 0.5% of diet.³⁶

Astragalus injection (at the equivalent of 6 g/kg/day of root by oral gavage) protected against cyclophosphamide-induced thymus injury in mice.³⁷ In other experiments, injection of Astragalus protected the immune organs of rats with obstructive jaundice,³⁸ and improved haematopoiesis in myelosuppressed mice.^39,40 Astragalus extract by injection demonstrated potential as a vaccine adjuvant⁴¹ (this is a common property of saponins, including those in Astragalus).⁴² The injection of an Astragalus decoction (10 g/kg) prevented airway hyper-reactivity in the ovalbumin mouse model of chronic asthma by inhibiting Th2 cytokine release.⁴³

High oral doses of Astragalus decoction given to healthy subjects (15.6 g/day for 20 days) significantly increased serum IgM, IgE and cAMP.⁴⁴ Two months of oral treatment in people susceptible to the common cold greatly increased levels of IgA and IgG in nasal secretions, and administration for 2 weeks or 2 months enhanced the induction of interferon by peripheral white blood cells.⁴⁵ Healthy human volunteers treated with Astragalus fresh root tincture (15 mL/day, equivalent to 1.23 g of root for 7 days) demonstrated significant increases in white blood cell CD69 expression (a marker for lymphocyte activation) after 1 and 7 days compared with placebo in a small, double blind trial.⁴⁶ Patients receiving chronic haemodialysis treated with intravenous Astragalus injection (30 mL two to three times a week for 2 months) exhibited significantly greater levels of interleukin-2 than patients in the control group in a small trial involving 31 people, suggesting a favourable effect on immune function.⁴⁷ The expression of CD25 on T cells was also significantly increased in a similar clinical study using the same doses after 24 hours (p<0.02), but not after 7 days.⁴⁸

There is a substantial number of in vitro studies on the immune effects of Astragalus polysaccharides of varying composition. Astragalus polysaccharides in vitro potentiated the immune-mediated antitumour activity of interleukin-2⁴⁹ and the activity of monocytes.⁵⁰ Astragalus polysaccharides improved the in vitro responses of lymphocytes taken from normal volunteers and cancer patients⁵¹ and enhanced the NK cell activity in vitro of blood samples from normal volunteers and systemic lupus erythematosus (SLE) patients.⁵² The polysaccharide fraction F3 potentiated the lymphokine-activated killer cell-inducing activity of interleukin-2 in blood taken from cancer and AIDS patients.⁵³ Other studies have shown that Astragalus polysaccharides in vitro activated and exerted mitogenic activity on B cells,^54,55 promoted neutrophil adhesion to vascular endothelial cells,⁵⁶ activated macrophages^55,57 and dendritic cells,^58,59 improved the innate immune response of bladder epithelial cells,⁶⁰ and induced differentiation of splenic dendritic cells, followed by the shifting of Th2 to Th1 balance, with enhancement of T cell function.⁶¹

Injection of Astragalus polysaccharides downregulated the immune response in rats with glomerulonephritis⁶² and mice with type 1 diabetes,⁶³ augmented antibody responses⁵⁴ and restored the lymphocyte blastogenic response in older mice.⁵⁴Astragalus polysaccharides (2 g/kg/day, oral) also prevented the development of type 1 diabetes in NOD mice by correcting the Th1/Th2 cytokine imbalance.⁶⁴ Oral doses of Astragalus polysaccharides (2 mg per chick) improved immune responses in chickens vaccinated against Newcastle disease⁶⁵ and at 220 mg/kg of feed acted synergistically with probiotic bacteria in modulating chick immune responses.⁶⁶ The innate immune response (phagocytic activity) was enhanced in sea cucumbers fed Astragalus polysaccharides (0.3% of feed) and superfine root powder (3.0% of feed).⁶⁷

A comparison of oral doses of the whole extracts of Astragalus membranaceus, A. membranaceus var. mongholicus and Hedysarum polybotrys (all at 560 mg/kg) and their various chemical fractions (all at 560 mg/kg) demonstrated that the polysaccharide fractions of all three herbs were the most active fractions and exerted similar levels of immune enhancement in two separate assays in mice.⁶⁸ Another study found that the polysaccharides from four Chinese herbs, including Astragalus, were all active at inducing serum antibodies and promoting T cell proliferation following ip injection in chickens.⁶⁹ These studies highlight a key paradox of the research into the contribution of polysaccharides to the immune activity of herbs, namely that the polysaccharide fraction from any plant is likely to have immune-enhancing activity. A key question is: why has a particular herb, such as Astragalus, developed a strong traditional reputation as an immune-enhancing herb if such activity is solely due to similarly acting components common to most other plants?

The antiviral activity of Astragalus is most likely to be due to increased immunity and possibly enhanced interferon production.² In support of this, Astragalus demonstrated slight inhibitory activity against adenovirus type 7 in vitro. Natural and recombinant interferon enhanced the inhibitory activity of Astragalus.⁷⁰ It also promoted the production of interferon by mouse lung against parainfluenza virus type I and Newcastle disease virus in vitro.⁷¹ Astragalus exhibited potent hepatitis B surface antigen-inactivating activity in vitro,⁷² inhibited the activity of murine retroviral reverse transcriptase and human DNA polymerases⁷³ and had a protective effect on cultured beating heart cells infected with coxsackie B2 virus.⁷⁴

Astragalus extract exhibited activity against herpes simplex virus type 1 in vitro⁷⁵ and countered the growth-inhibitory effect of cytomegalovirus infection on human cord blood progenitor cells.⁷⁶ Calycosin-7-O-beta-glucopyranoside, a major isoflavonoid in Astragalus, inhibited coxsackie virus B3 (CVB3) in vitro and was also active (24 mg/kg, oral) in an acute myocarditis model in mice.⁷⁷

Oral or intranasal administration of Astragalus decoction protected mice from infection with parainfluenza virus type I.^24,45,78 Results from a series of in vivo experiments indicated that the effect of Astragalus resembled that of both bronchitis vaccine and the interferon mediator tilorone.⁴⁵ Astragalus polysaccharides had a weak inhibitory effect on hepatitis B virus in mice,⁷⁹ but astragaloside IV was quite active against the virus in vitro.⁸⁰

Astragalus increased the survival rate and improved some abnormal electrophysiological parameters in acute CVB3 viral myocarditis in vivo.⁸¹ In vitro and in vivo studies indicate Astragalus may act by decreasing the secondary damage caused by calcium ion influx, thereby improving abnormal myocardial electric activity, as well as inhibiting the replication of CVB3 virus RNA in the myocardium.^82,83

Astragalus feeding (2.2 mg/kg/day) increased survival rate, alleviated pathological changes and reduced markers of cardiac damage in mice with CVB3 myocarditis.⁸⁴ In contrast to other work, an inhibitory effect on virus RNA replication in vivo was not correlated with the induction of beta-interferon,⁸⁵ but was greater than a calcium channel blocker (verapamil) and a steroidal anti-inflammatory drug (dexamethasone) in vitro.⁸⁶ Routine therapy combined with oral administration of Astragalus to viral myocarditis patients significantly enhanced immune parameters when compared with patients receiving routine therapy alone.⁸⁷

Addition of Astragalus enhanced growth, metabolism and longevity in cell cultures.² It lowered oxygen consumption in mitochondria, enhanced tolerance to stress and prolonged the life of human embryonic kidney cells in culture.⁸⁸ Two isomers of the molecule HDTIC extracted from Astragalus extended the lifespan of human fetal lung diploid fibroblasts in vitro by slowing telomere shortening, reducing DNA damage and improving DNA repair.⁸⁹ They also delayed replicative senescence in the same model.⁹⁰ The telomerase activator cycloastragenol (TA-65, found at low levels in Astragalus) elongated short telomeres in mouse embryonic fibroblasts and improved some healthspan indicators in adult/old mice without increasing cancer incidence.⁹¹ Such observations have been interpreted by some as suggestive that Astragalus or these components will promote human longevity, but this is clearly premature on current limited evidence.

Administration of Astragalus over 2 weeks to mice markedly increased plasma cAMP.⁹² Astragalus decoction improved learning performance in animal maze tests⁹³ and improved memory in two models (50 g/kg/day for 7 days, oral).⁹⁴ Administration over 15 days inhibited field search behaviour, decreased spontaneous activity and increased sleep time.⁹³ Decoction of Astragalus improved endurance in mice and increased weight gain compared with controls.¹ A mixture of ginseng and Astragalus demonstrated antifatigue activity in mice. This activity was partly due to an improvement of energy metabolism.⁹⁵ Astragalus extract (400 mg/kg/day) countered the adverse effects of repeated restraint stress in rats, improving spatial learning and memory and reducing anxiety.⁹⁶ Oral administration of Astragalus increased the turnover of proteins in serum and liver in animals treated daily for 10 days.⁹⁷ Astragalus lowered collagen content in the aorta and lung of old rats to near levels found in young animals⁹⁸ and improved the density of M-cholinergic brain receptors.⁹⁹ In early research, oral administration of Astragalus increased plasma cAMP in healthy subjects.¹⁰⁰

Astragalus saponins demonstrated a positive inotropic action on isolated heart and decreased the resting potential of cultured myocardial cells, suggesting an inotropic effect exerted through modulation of Na⁺,K⁺-ATPase.¹⁰¹ Astragalosides reduced intracellular calcium overload in rat cardiomyocytes and enhanced free radical removal, both suggestive of a protective effect against myocardial injury.¹⁰² Astragaloside IV improved post-ischaemic heart function and ameliorated reperfusion arrhythmias in rat hearts in vitro,¹⁰³ and improved intracellular calcium handling in hypoxia-reoxygenated cardiomyocytes.³⁴ Astragalus extract prevented daunorubicin-induced apoptosis of cultured cardiomyocytes by decreasing free radical release¹⁰⁴ and demonstrated a cardiotonic effect in isolated beating rabbit atria.¹⁰⁵ Astragaloside IV improved homocysteine-induced endothelial dysfunction in rat aortic rings via antioxidant activity.¹⁰⁶

Oral administration of aqueous extract of Astragalus countered the rise in blood pressure and plasma renin activity in a hypertensive model.¹⁰⁷ Intragastric administration of Astragalus produced a hypotensive effect in another experimental model.¹⁰⁸ Gamma-aminobutyric acid was isolated as a potential hypotensive constituent.¹⁰⁹ Oral Astragalus improved impaired endothelial-dependent vasodilation in obese rats.^110,111 Astragaloside IV has exhibited vasodilatory effects in vivo,^112,113 as has the whole extract.¹¹⁴

Improvement of cardiac function has been demonstrated for Astragalus or its isolates in a number of in vivo models. In several such studies Astragalus or Astragalus components were given by injection. Examples of oral dose studies include the inhibitory effect of Astragalus (5, 10 and 20 g/kg/day) on left ventricular hypertrophy induced by pressure overload in rats.¹¹⁵ Astragalus (20 g/kg, oral) also counted abnormal cardiac function in rats with pressure overload-induced heart failure, and at oral doses of 3.3 or 10 g/kg/day improved cardiac function in doxorubicin-injured rat hearts.¹¹⁶ All these doses are rather high. Astragalosides have also demonstrated cardioprotective activity in vivo following administration by injection,^103,117 as has Astragalus injection.¹¹⁸

Cardiac output increased in 20 patients with angina pectoris after 2 weeks of treatment with Astragalus.¹¹⁹ Astragalus strengthened left ventricular function and had an anti-OFR (oxygen free radical) effect in acute myocardial infarction patients compared with controls. The decrease in the pre-ejection period:left ventricular ejection time ratio was closely correlated with the increase in superoxide dismutase activity of red blood cells and the decrease in lipid peroxidation of plasma. This anti-OFR activity of Astragalus may be one of the mechanisms behind its cardiotonic activity.¹²⁰

Astragalus extract demonstrated a protective effect on erythrocyte deformability in vitro for blood taken from normal subjects and patients with SLE.¹²¹ Astragalus significantly enriched the blood, as measured by improvement in haemorheological indices,¹²² and, in a ‘blood stagnation’ experimental model, decreased whole-blood specific viscosity and increased plasma specific viscosity.¹²³

Astragalus saponins were protective against chemically induced liver injury in vitro and in vivo.¹²⁴ Oral doses of Astragalus polysaccharides (200 mg/kg/day) also demonstrated hepatoprotective activity in rats.¹²⁵ Astragalus extracts exhibited hepatoprotective activity and increased the activity of hepatic lysozymes, tissue dehydrogenase and liver glycogen.^126–128 Oral doses of an extract combination of Astragalus and Paeonia lactiflora (60, 120 and 240 mg/day) protected against immunological liver injury in mice.¹²⁹ The total flavonoids of Astragalus (100 mg/kg, oral) protected against paracetamol liver damage in mice.^130,131

Some studies suggest that Astragalus in combination might protect against the hepatic fibrosis associated with chronic liver damage. Oral doses of the Astragalus and Paeonia combination mentioned above (80 and 160 mg/kg/day) reduced liver damage and fibrosis in rats with carbon tetrachloride-induced liver injury and decreased the elevation of tumour growth factor (TGF)-beta1.¹³² Oral doses of a combination of Astragalus and Salvia miltiorrhiza (60, 120 and 240 mg/kg/day) also demonstrated antifibrotic activity in a similar model.¹³³

A 2009 systematic review examined the published in vivo studies for Astragalus in early diabetic nephropathy in rats.¹³⁴ Of 41 articles identified, 13 reports that fulfilled the inclusion criteria were reviewed. Meta-analysis revealed that Astragalus extract (orally or by injection depending on the study, but mainly the former) significantly reduced fasting blood glucose, glomerular filtration rate, urinary albumin excretion and thickness of the glomerular basement membrane (p values ranging from <0.00001 to 0.03).

Several other in vivo studies have demonstrated protective effects on renal function in a variety of models, including IgA nephropathy.^135–140 Doses used were often relatively high and the route of administration was not always specified.

See also the Clinical trials section for diuretic activity in a human study.

In addition to the studies included in the meta-analysis above, the effects of Astragalus or its components have also been examined in several diabetic models. For example, oral doses of Astragalus polysaccharides (400 to 2000 mg/day) alleviated glucose toxicity and restored glucose homeostasis,¹⁴¹ lowered blood glucose and reduced insulin resistance^142–144 and reduced cardiomyopathy¹⁴⁵ in various diabetic models. Other components have also been shown to have antidiabetic activity, including the isoflavonoids¹⁴⁶ and astragaloside IV, which protected against diabetic neuropathy in rats (3, 6 and 12 mg/kg twice a day, oral).¹⁴⁷

Astragalus saponins,^148–150 Astragalus injection^151,152 and an Astragalus extract¹⁵³ have all shown growth inhibitory activity against a variety of tumour cell lines in vitro. Mechanisms included promotion of apoptosis,^148,149,153 growth inhibition¹⁵⁰ and downregulation of Akt phosphorylation.¹⁵¹

Astragalus saponins (100 and 200 mg/kg/day for 20 days, oral) reduced tumour volume in a mice xenograft model of colon cancer, demonstrating activity comparable to 5-fluorouracil.¹⁵⁰ Various fractions of Astragalus polysaccharides (prepared by decoction followed by stepwise ethanol precipitation) demonstrated weak cytotoxic activity in vitro, but promoted immune response in a mouse cancer model after ip injection.¹⁵⁴ Astragalus extracts demonstrated chemopreventative activity in mice (10, 20 and 40 mg/kg/day, oral)¹⁵⁵ and rats (90 and 180 mg/kg/day, oral).¹⁵⁶

Astragalus flavonoids demonstrated a protective effect on mammalian cell damage caused by the hydroxyl radical, inhibited lipid peroxides and increased superoxide dismutase activity in vitro.¹⁵⁷ Three Astragalus saponins demonstrated superoxide anion scavenging activity in vitro.¹⁵⁸ Astragalus extract reduced free radical-mediated injury to renal tubules in rabbits (2.4 g/kg, iv).¹⁵⁹ Astragalus injection reduced measures of oxidative stress and inflammation in a controlled trial involving 60 haemodialysis patients.¹⁶⁰

An aqueous extract of Astragalus demonstrated a broad anti-inflammatory effect on human amnion cells in vitro.¹⁶¹ Astragalosides inhibited the formation of advanced glycation endproducts during the incubation of bovine serum albumin with ribose.¹⁶²

Astragalus polysaccharides (250, 500 and 1000 mg/kg/day, oral) were anti-inflammatory in rats with adjuvant arthritis¹⁶³ and the 8:1 dried aqueous extract of Astragalus (30 and 100 mg/kg/day, oral) reduced the expression of various inflammatory mediators in zymosan air-pouch mice.¹⁶⁴ Astragalus decoction (100 mg/kg/day, oral) also inhibited the development of experimental atopic dermatitis in mice.¹⁶⁵ High oral doses of an aqueous extract of Astragalus either before (2 and 4 g/kg/day) or after (4 and 8 g/kg/day) hapten provocation reduced experimental colitis in rats.¹⁶⁶ Oral (4 and 8 g/kg/day) or intracolonic (200 and 800 mg/kg/day) doses of this Astragalus extract also demonstrated similar therapeutic activity in the same colitis model, but by modulation of different colonic cytokines, depending on the mode of administration.¹⁶⁷

Astragalus extract,¹⁶⁸ astragaloside IV¹⁶⁹ and Astragalus isoflavones¹⁷⁰ exhibited neuroprotective activity in various in vitro models. Peripheral nerve regeneration was demonstrated in vitro and in vivo (in rats via local administration) for an Astragalus extract.¹⁷¹ An Astragalus extract (150 and 300 mg/kg, oral) also protected against pentylenetetrazole-induced seizures in mice.¹⁷²

Astragalus inhibited aldose reductase,¹⁷³ promoted the replication of hepatic DNA,¹⁷⁴ inhibited mitochondrial oxygen consumption caused by lipid peroxidation¹⁷⁵ and stimulated the motility of human sperm,¹⁷⁶ all in vitro. Incubating sperm from infertile men with Astragalus decoction significantly enhanced motility.^177,178 Intraperitoneal injection of Astragalus extract into rats made infertile by cadmium resulted in significant increases in sperm counts and reduced sperm malformation, compared with untreated controls.¹⁷⁹

A methanolic extract of Astragalus inhibited the growth of the human intestinal bacterium Clostridium perfringens in vitro.¹⁸⁰ Oral administration of Astragalus normalised the imbalance in intestinal flora in an experimental model of senility.¹⁸¹

Oral administration of a concentrated solution of Astragalus strengthened small intestine movement and muscle tonus, especially in the jejunum.¹⁸² This activity supports its traditional use in organ prolapse.

Astragaloside IV (3, 10 and 30 mg/kg/day, oral) exerted gastroprotective effects in rats with ethanol-induced gastric mucosal damage.¹⁸³

In an open, randomised clinical trial, 115 patients with leucopenia received a high dose of a concentrated Astragalus preparation (equivalent to 30 g/day of Astragalus) or a low dose (equivalent to 10 g/day) over a period of 8 weeks. There was a significant rise of average white blood cell (WBC) counts in both groups after treatment (p<0.001). The average WBC count for the high-dose group was significantly higher than for the low-dose group (p<0.05). On the basis of these findings, the author suggested that Astragalus is an effective treatment for leucopenia and increasing the dosage could enhance its effectiveness.¹⁸⁶ In an open study, 1000 volunteers received Astragalus either orally, as a nasal spray or in a compound formula. A prophylactic effect for the common cold was observed, as evidenced by decreased incidence and shortened duration of infection.⁴⁵

A combination of Astragalus (320 mg/day of an 18:1 extract containing 40% polysaccharides) and calcium-aluminium silicate as a mineral carrier and possible synergist was investigated in a 6-week double blind, placebo-controlled clinical trial involving 48 patients with seasonal allergic rhinitis.¹⁸⁷ Both the attending physicians and the patients judged the active treatment as more efficacious than placebo (p=0.003 and p=0.026, respectively). However, no statistically significant differences were detected between the groups for serum IgE, IgG and nasal eosinophils, although there was a trend to lower IgG for Astragalus plus minerals (p=0.18). In a post hoc analysis of 21 patients with weed pollen allergy, Astragalus plus minerals significantly improved total symptom score (p=0.022) and quality of life (p<0.001) compared with placebo. All adverse events were mild and not considered to be connected to the active treatment.

Astragalus in combination with other herbs has also been investigated for clinical effects on immune function. In a double blind, placebo-controlled trial, 85 children aged 7 to 15 years with asthma were randomly assigned to receive either an oral herbal formula (0.619 g/day, comprising equal weights of dried aqueous extracts from Astragalus, Cordyceps, Stemona, Fritillaria and Scutellaria baicalensis), or placebo for 6 months.¹⁸⁸ There was no significant difference recorded for any of the trial endpoints between the active treatment and placebo groups. The impact of Compound Astragalus Recipe was investigated in 60 patients with myasthenia gravis in a controlled trial.¹⁸⁹ While clinical outcomes were similar in the herbal and control groups, the herbal treatment significantly lowered the CD4+/CD8+ ratio (p<0.05).

Astragalus injection has demonstrated a range of effects on immune function in open label clinical trials. For example, it improved cellular immunity in patients with serious abdominal trauma,¹⁹⁰ promoted recovery of haematopoietic function in patients with chronic aplastic anaemia,¹⁹¹ improved immune function in patients with congestive heart failure¹⁹² and enhanced the efficacy of conventional treatment for SLE.¹⁹³

Administration of Astragalus to a large number of patients with chronic viral hepatitis resulted in a success rate of 70% in an open trial. In most cases, elevated serum ALT levels returned to normal after 1 to 2 months.¹⁹⁴

In a double blind clinical trial, 235 patients with typical chronic cervicitis (associated with viral infection) received one of the following treatments applied locally by gauze: recombinant interferon-alpha1 (at 5 or 10 mg), combined interferon (5 mg) and Astragalus (0.5 mL of a 1:1 extract), or Astragalus alone (0.5 mL of a 1:1 extract). These treatments were applied twice per week for 3 weeks. The Astragalus plus interferon group showed a similar improvement to the higher dose interferon group, with approximately 60% of patients demonstrating complete resolution or marked improvement. Only 8% of patients treated with Astragalus alone exhibited marked improvement and no patients were completely cured. These results suggest that Astragalus acted synergistically with interferon therapy.¹⁹⁵ An earlier double blind trial showed a similar result for 164 patients with cervical erosion associated with herpes simplex virus infection.¹⁹⁶ In 106 patients with herpes simplex keratitis, those randomly assigned to Astragalus treatment (dose and route not specified in the English abstract) exhibited significant improvement in the rebalancing of Th1/Th2 cytokines, an effect not observed in the comparative control group receiving ribavirin.¹⁹⁷

An infused formulation of six herbs with Astragalus as the main component was significantly better than a control treatment (including silymarin) in terms of clinical improvement and greater negative seroconversions in an open label trial involving 208 patients with chronic viral hepatitis B.¹⁹⁸

A formulation containing extracts of Astragalus combined with Glycyrrhiza glabra, Artemisia capillaris, Morus alba and Carthamus tinctorius was investigated in HIV positive patients in Thailand. In an open label study, 28 HIV-1 infected adults (CD4 count >200 cells/mm³ and HIV-1 RNA >20 000 copies/mL) received 5 g/day of the combination plus sulfamethozaxole and trimethoprim for 12 weeks.¹⁹⁹ Up to 36% of patients demonstrated a reduction in plasma HIV-1 RNA of more than 0.5 log during the trial, but CD4 counts were unchanged. In a subsequent randomised, double blind, 24-week clinical trial, the efficacy of the herbal combination (7.5 g/day) given with zidovudine and zalcitabine was compared against the two drugs plus a herbal placebo in 60 HIV-1 positive patients.²⁰⁰ The decline in HIV RNA was significantly greater in the group receiving the herbal combination (p<0.001) and CD4 cell counts were higher compared with baseline (p<0.05), versus no change in the control group. Serious adverse events were not observed.

A 2004 Cochrane review of herbal medicines for viral myocarditis found 40 randomised trials involving 3448 patients.²⁰¹ Twenty-five different herbal treatments were tested in the included trials. Astragalus was given as a single treatment in 10 trials (oral doses in three, by injection in seven) and as part of a herbal combination in another 13 trials (all doses oral). Analysis of the 10 Astragalus-only trials (plus one where the formulation was mainly Astragalus) found Astragalus significantly improved premature beat, cardiac output, ejection fraction and some levels of myocardial enzymes (indicating cardiac damage). Overall the authors stressed the low methodological quality, but suggested some herbal medicines such as Astragalus deserve further examination in this context in rigorous trials.

In a comparative trial, 92 patients suffering from ischaemic heart disease were treated with Astragalus, Salvia miltiorrhiza or the anti-anginal drug nifedipine. Results were superior for the Astragalus-treated group, as demonstrated by marked relief from angina pectoris and improvement in several objective clinical parameters. Treatment of ischaemic heart disease by Astragalus was significantly more effective compared with the control group (p<0.05).²⁰² In an open label trial in 20 patients with angina pectoris, Astragalus (60 g/day, presumably by decoction) significantly improved cardiac output by 16.9% after 2 weeks (p<0.01), with no improvement of left ventricular diastolic function.²⁰³ Unlike digitalis, the herb did not inhibit ATP activity.

Forty-five patients with chronic heart failure (defined in TCM (traditional Chinese medicine) terms by Xin-qi or Xin-yang deficiency) were randomised to receive conventional medicine or conventional medicine plus 4.5 g/day (oral) of an Astragalus granule containing only Astragalus.²⁰⁴ While improvements from baseline in TNF-alpha, left ventricular ejection fraction (LVEF) and walking distance were observed in both groups, they were all significantly higher in the group receiving Astragalus (p<0.05).

Ninety chronic heart failure patients (TCM diagnosis as above) were randomly assigned to receive perindopril (4 mg/day) and one of three different doses of Astragalus granule (15, 9 or 4.5 g/day).²⁰⁵ Clinical improvements, including for LVEF, walking distance and quality of life, were greatest in the highest dose group (p<0.01).

Ninety-four patients with vascular disease secondary to type 2 diabetes mellitus were randomly assigned to either Astragalus with saponins from Panax notoginseng, or simvastatin, in an open label trial. Blood cholesterol and triglyceride levels and a measure of vascular inflammation (MMP-9) improved similarly in the two groups.²⁰⁶

Astragalus injection has demonstrated a range of effects in cardiac patients in clinical trials. For example, it improved cardiac function and haemodynamics in children with tetralogy of Fallot,²⁰⁷ improved heart function parameters in patients with congestive heart failure^208,209 and acute myocardial infarction,²¹⁰ and decreased inflammatory cytokine production in patients undergoing heart valve replacement.²¹¹ Injected doses typically contained high levels of the crude herb (up to 80 g).

A randomised, double blind, placebo-controlled crossover study in 12 healthy men assessed the impact of a single oral dose of Astragalus (0.3 g/kg of a dried 4:1 aqueous extract) on diuresis.²¹² Compared with placebo, Astragalus markedly increased urinary sodium excretion, fractional sodium excretion and urinary excretion of chloride during the first 4 hours. The authors concluded that Astragalus induces marked natriuresis in healthy men and attributed this to enhanced renal responses to endogenous ANP (atrial natriuretic peptide).

A US group of doctors described two separate cases (published 3 years apart) of remission of idiopathic membranous nephropathy (IMN, probably autoimmune in origin) after therapy with Astragalus. The first case described a 77-year-old woman with nephrotic syndrome secondary to IMN who was largely unresponsive to conventional treatments.²¹³ After beginning Astragalus (15 g/day as part of the formulation Shen-Yan Siwei Pian) there was a marked decrease in proteinuria. Nephrotic syndrome recurred after a temporary cessation of the formulation, with complete remission after its reintroduction. Remission persisted even after stopping the herbal treatment. The second case was a 63-year-old man with nephrotic syndrome due to IMN.²¹⁴ In addition to conventional treatments (which had not resolved his proteinuria) he took Astragalus (15 g/day herb equivalent of a 4:1 extract – presumably aqueous) for around 12 months, after which he experienced complete remission of nephrotic syndrome.

In an open clinical trial, a combination of Arctium lappa fruit and tincture of Astragalus orally for 3 months with losartan was compared with losartan alone in 54 patients with diabetic nephropathy. The herbal combination reduced hyperlipidaemia, proteinuria and postprandial hyperglycaemia significantly compared with losartan alone.²¹⁵ Patients with diabetic nephropathy were randomly assigned to take simvastatin or a combination of Astragalus and Panax saponins. Both treatments reduced hyperlipidaemia and various markers of renal damage.²⁰⁶ In an open label trial, 21 patients with type 2 diabetes and microalbuminuria received 150 mL four times a day of an unspecified Astragalus and Ligusticum decoction for 6 months.²¹⁶ The herbal combination improved both urinary albumin excretion and endothelial dysfunction.

Astragalus injection improved renal tubular function in patients with IgA nephropathy²¹⁷ and primary nephrotic syndrome.²¹⁸

A meta-analysis of 34 randomised clinical trials involving patients with non-small-cell lung cancer treated with platinum-based chemotherapy and Astragalus-based Chinese herbs suggested a benefit from the combination.²¹⁹ Most trials involved formulas featuring Astragalus, but two were of Astragalus alone. The herbs were administered by injection in around one-third of the trials. Twelve trials measuring such outcomes reported significantly lower mortality rates after 12 months when Astragalus was combined with chemotherapy (risk ratio 0.67). Nine studies reported significantly lower mortality rates after 24 months when Astragalus was combined with chemotherapy (risk ratio 0.73). Most of the studies included were of low methodological quality. A Cochrane review indentified four relevant trials where a decoction of Astragalus and a formulation featuring Astragalus was combined with chemotherapy regimens in patients with colorectal cancer.²²⁰ Chemotherapy-induced nausea, vomiting and leucopenia were all decreased by concomitant administration of Astragalus decoction, and immune function was improved. The trials were of low quality, suggesting larger, more rigorous trials are needed to confirm these results.

A systematic review and meta-analysis of Chinese herbs in the treatment of hepatocellular carcinoma found that formulations containing Astragalus had a larger treatment effect than the pooled broad estimate (odds ratio 1.35, p=0.048).²²¹ Products containing Astragalus significantly improved 12-month survival rates (odds ratio 1.28, p<0.0001). While there were questions over trial design, the authors considered this to be compelling evidence that needs to be evaluated in high-quality clinical trials. The herbal treatments were often administered by injection.

In an open label trial, 20 children with acute leukaemia in remission received Astragalus (90 g/day, mode of administration not specified in the English abstract) plus chemotherapy for 1 month, while another 24 in the control group received chemotherapy alone.²²² The herbal treatment increased dendritic cell induction of peripheral mononuclear cells and enhanced antigen-presenting ability.

In a double blind, placebo-controlled clinical trial involving 507 elderly people, oral administration of Astragalus in combination with Polygonum multiflorum and Salvia miltiorrhiza demonstrated significant antiageing effects. Improvements were noted in vigour, strength, vision, cellular immunity and serum lipofuscin levels. The total effective rate was 76.6% compared with 34.5% for placebo (p<0.001).²²³

Eighty-four patients with liver cirrhosis and portal hypertension were randomly assigned to receive either conventional treatment or a combination of Astragalus and Salvia miltiorrhiza (dan shen) for 3 months in an open label trial.²²⁴ There were significant improvements in haemodynamic measures and indices of liver fibrosis in the group treated with the herbal combination compared with the control group.

A 4-week randomised, double blind, placebo-controlled clinical trial was conducted with 36 adults with chronic fatigue (but not necessarily chronic fatigue syndrome).²²⁵ The trial participants were divided into a control group (receiving a ‘placebo’ Chinese herbal formulation known as Hyangsapyunweesan (in Korean), 3 g/day) or 3 or 6 g/day of a 3.3:1 concentrated aqueous extract from equal parts of Astragalus and dan shen. This combination significantly decreased subjective fatigue severity scores compared with the control group (p<0.05).