A survey of Chinese herbal ingredients with liver protection activities

A literature survey was conducted on herbs, their preparations and ingredients with reported liver protection activities, in which a total of 274 different species and hundreds of active ingredients have been examined. These ingredients can be roughly classified into two categories according to their activities: (1) the main ingredients, such as silybin, osthole, coumarin, glycyrrhizin, saikosaponin A, schisandrin A, flavonoids; and (2) supporting substances, such as sugars, amino acids, resins, tannins and volatile oil. Among them, some active ingredients have hepatoprotective activities (e.g. anti-inflammatory, anticancer, antioxidant, immunomodulating and liver cirrhosis-regulating effects). Calculation of physicochemical parameters indicates that the main ingredients with negative and positive Elumo values possibly display their hepatoprotective effects through different mechanisms, such as antioxidative, anti-inflammatory and immunomodulating effects. As the combination of herbs may achieve some treatment effects synergistically and/or additively, it is common in Chinese medicine to use mixtures of various medicinal herbs with pharmacologically active compounds to have synergistic and/or additive effects, or to reduce harmful effects of some pharmacologically active compounds. In particular, the active compounds with Clog P around 2 are suitable for passive transport across membranes and accessible to the target sites. Thus, Elumo and Clog P values are good indicators among the calculated parameters. Seven different physicochemical parameters (MW, Clog P, CMR, μ, Ehomo, Elumo and Hf) and four major biological activities (antioxidant, anti-inflammatory, antiviral/antitumor and immunomodulating) are discussed in this review. It is hoped that the discussion may provide some leads in the development of new hepatoprotective drugs.


Background
It is well recognized that liver is one of the most important organs in the biotransformation of food, drugs, endogenous and exogenous substances. Profuse supply of blood and the presence of many redox systems (e.g. cytochromes and various enzymes) enable liver to convert these substances into different kinds of inactive, active or even toxic metabolites. The burden of metabolism and exposure to dangerous chemicals make liver vulnerable to a variety of disorders, such as acute or chronic inflammation, toxin-/ drug-induced hepatitis, cirrhosis and hepatitis after viral infection.
For centuries, many herbs have been used as natural remedies for the prevention and/or treatment of liver diseases. Various herbs and herbal products are believed to have liver protective functions and widely used in clinical practice in the West as well as East. In the First Edition of PDR for Herbal Medicine [1], for example, a total of 49 herbal products were listed under the preparations for clinical use in liver and gall bladder complaints and 32 products for liver disorders. Some complex Chinese herbal formulae, such as Pro-liver Pill (Yanggan Wan) [2], Liver Care (Himalaya Drug Co, Bangalore, India), Liv-52 [3], Jianpi Wenshen Pill (Jianpi Wenshen Wan) [4], Binggan capsules (Binggan Jiaonang) [5], Binggan Tang [6], Yizhu decoction(Yizhu Koufuye) [7], Yiergan Tang [8] and Xiaochaihu Tang (Sho-saiko-to or SST,) [9], have been reported to have significant therapeutic effects on liver protection or treatment of liver diseases. It is yet to be established as to which of the herbs and their active ingredients in these formulae contribute most to the activities of liver protection and treatment.
Both manual and electronic search of literature and/or biological information in herbal medicine was carried out in order for us to examine which structural and physicochemical factors probably affect hepatoprotection. Based on two reference books, namely Zhongcaoyao Xiandai Yanjiu [10][11][12] and The Chemical Constituents of Oriental Herbal Drugs (Vol 1 & 2) [62,63], electronic literature search was performed with "ovid"(an online resource with access to multiple databases such as Medline, Biosis, Embase, Current Contents, as well as journals and books), by the key words of "herbal medicine with hepatoprotective activity" and was mainly focused on the literature from 1995 to 2005. In this review, most of the reported ingredients of the heptoprotective herbs are summarized and listed under family, genus and species names of the herbs (Additional file 1) and seven physicochemical parameters of the hepatoprotective compounds, namely molecular weight (MW), calculated octanol/water partition coefficient (Clog P), molar refraction (CMR), dipole moment (μ), energy of the highest occupied molecular orbital (E homo ) , energy of the lowest unoccupied molecular orbital (E lumo ) and heat of formation (H f ), have been calculated and reported (Additional file 3). Chemical structures of some of the representative compounds together with their respective calculated physicochemical parameters are also provided (Additional file 2).

Biological activities of extracts and ingredients of medicinal herbs
According to the results from our literature survey, nearly 300 species of plants belonging to 92 different families have been reported to have liver protection or therapeutic effects on hepatic disorders (Additional file 1). Families of Compositae, Labiatae and Leguminosae cover the largest numbers of species.
These plant species usually contain phytochemicals such as phenols, organic acids, flavonoid glycosides, tannins, resins, amino acids, sugars, alkaloids, saponins and volatile oil as well as their own unique constituents. It is not clear whether one, several or all of these components are active ingredients for liver protection. It is likely that some specific ingredients of each herb play a vital role in liver protection/treatment, thereby contributing to their therapeutic effects. Structures of some representative liver protective compounds  and key physicochemical parameters which may affect their pharmacokinetic and pharmacodynamic effects are provided (Additional file 2).
Silymarin, a mixture of flavonolignans extracted from the seeds of Silybum marianum, is most widely used as a remedy for liver diseases. It is composed of four isomers: the major active component silybin, isosilybin, silychristin and silydianin ( Figure 1). Moreover, other seed constituents of Silybum marianum such as betaine, other flavonolignans and essential fatty acids from the same plant have been shown to have anti-inflammatory properties [37]. The beneficial effects of silymarin are most often observed in patients with cirrhosis as a result of alcohol abuse [38]. But it does not reduce mortality; nor does it improve liver histology or biochemical markers of liver function among patients with chronic liver diseases [35]. It was reported that treatment with Silybum marianum and its constituents appears to be safe with no signs of adverse effects [15].
Structure-bioactivity relationship studies indicate that GA is the most active ingredient against platelet aggregation, tumor cell lines, rotavirus infection and growth of Helicobacter pylori, followed by GAMA and GL [41]. Thus, GA is a better hepatoprotective drug than GAMA or GL. It has    Glycyrrhizin(GL) GAMA GA + -D-glucuronidase in intestines and liver been suggested that GL (a natural prodrug and sweet principal of Glycyrrhiza) and its hydrolytic metabolites (GAMA, GA) bind to hepatocytes to modify the expression of HBV (hepatitis B virus)-related antigens and suppress sialylation of hepatitis B virus surface antigen (HbsAg) [34,42]. Clinical studies showed that GL has comparable antiviral effects to those of interferon and that it is effective in reducing complications of chronic hepatitis C without apparent side-effects [43,44].

Structures of the four isomers in silymarin
Schisandra chinensis (Wuweizi), traditionally used as a tonic, is now widely used for the treatment of chemical/ viral hepatitis. Biological studies indicate that seed extract of Schisandra chinensis can enhance hepatic GSH (glutathione) antioxidant/detoxification system and facilitate both processes in the liver. It may be a promising agent for improving phase I oxidative metabolism in liver damaged by CCl 4 [45]. Chemical investigations on the extract have revealed the presence of lignans with a dibenzocyclooctadiene skeleton such as schisandrin, gomisin A, deoxyschisandrin (schisandrin A), γ-schisandrin and wuweizisu C [46] (Figure 3). These components are believed to be active in hepatoprotective, anti-inflammatory, anticancer, anti-HIV and immunomodulating effects [47][48][49][50].
Structure-activity relationship (SAR) analyses suggest that the presence of methylenedioxy/a hydroxyl group in the dibenzocyclooctadiene skeleton is an important structural feature for hepato-and neuro-protective activities [51,52]. Osthole, a coumarin derivative contained in many medicinal plants especially in Boenninghausenia albiflora, was reported to protect against liver injury, and these effects are related to the presence of 7-methoxy and 3-methyl-2butenyl groups in the structure [53,54]. Due to its low toxicity, osthole appears to be suitable for the treatment of chronic hepatitis and can be used as a lead compound for further development of orally ingested hepatoprotective drugs.
Apart from single herbal medicine, 'Kampo medicine' which utilizes combinations of raw herbs, was widely used in Japan, among which SST is the most well-known. ( Figure 4). Comparative experiments showed that the liver concentrations of the active constituents of SST extract in the liver-injured rats increased significantly after the animals received SST extract, as compared with the groups receiving the respective purified constituents. Thus, administration of SST extract was more useful than that of its active ingredients individually [56].
In summary, the ingredients as stated above possess hepatoprotective activities, ranging from antiviral, antiinflammatory, anticancer, antioxidant to immunomodulating and liver cirrhosis regulating effects. Why do the active constituents with different structural features have common biological activities? There may be some innate factors affecting liver protective activities. (1) E lumo values can be divided into three subsets ranging from -1.12 ~ -0.10, and 0.03 ~ 1.07, to 1.10 ~ 2.12 (Additional files 2 and 3). As E lumo values reflect electron affinity of the respective compounds [57], these positive and negative ranges may indicate different mechanisms of actions in hepatoprotection. For example, the E lumo values of Silybum marianum, osthole, coumarin, as well as flavonoids and their glucosides, ranging from -1.03 to -0.83, suggest that their hepatoprotective activities may be achieved via their antioxidative properties, while glycyrrhizin, Schisandra chinensis and Bupleurum chinense, with their E lumo values ranging from 0.03 to 1.07, possibly exert their liver protection through other mechanisms (e.g. anti-inflammatory and/or immunomodulating).   Although biological effects of these ingredients have not been fully elucidated by these data, the antioxidant activities are indicated in their E lumo values. It is speculated that the compounds with negative E lumo values may act as free radical scavengers by having one electron from the highest occupied molecule orbital (HOMO) changed to the lowest unoccupied molecule orbital (LUMO), thereby eliminating the damaging effects of free radicals. The damaging effects of oxidation on the biological system via radical mechanism and the hepatoprotective effects by antioxidants and/or free radical scavengers are illustrated in Figure 5 [61].

Conclusion
Herbal medicine has been used in China for more than two thousand years and is now being gradually accepted worldwide, particularly in Asia, Europe and the North America. Differences, however, do exist between the East and West in the areas such as philosophical view point, concept of diseases and treatment approaches. In the West, people tend to use a single herb in therapies and try to identify active ingredients and therapeutic mechanisms, whereas in Chinese medicine it is common to use mixtures of various medicinal herbs which contain a range of pharmacologically active compounds working additively/synergistically to treat patients or to reduce harmful effects of some chemical compounds. It is our hope that the integration of Chinese and Western medicine will not only identify different ingredients with similar bioactivities or similar ingredients with different bioactivities, but also provide us with methods for quality control in herbal medicine and clues to elucidate action mechanisms of active ingredients.
This review is a summary of 274 species (in 216 genera among 92 families) of herbs that have been reported to have positive effects on liver protection and treatment, as well as hundreds of active ingredients. These ingredients can be roughly classified into two categories: (1) the main ingredients, such as silybin, osthole, coumarin, glycyrrhizin, saikosaponin A, schisandrin A, flavonoids, and (2) the supporting substances, such as sugars, amino acids, resins, tannins and volatile oil. Calculations of molecular parameters suggest that negative and positive E lumo values in the main ingredients represent two different hepatoprotective mechanisms. The main ingredients with negative E lumo values may display their hepatoprotective effects through an antioxidative mechanism, whereas other ingredients with positive E lumo values may act via an antiinflammatory and/or immunomodulating mechanism. Moreover, the Clog P values of many pharmacologically active compounds are around two and other ingredients with higher or lower Clog P values usually go through metabolic processes in which some hydrophilic groups are added or removed so that their Clog P values are around two. E lumo and Clog P values, therefore, should be useful in the future development of hepatoprotective drugs. Damaging effects of oxyradical on biological systems and sites of blockades ( ) by antioxidants/free radical scavengers (adapted and expanded from reference [61]).