Fitoterapia 92 (2014) 133147

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Fitoterapia
journal homepage: www.elsevier.com/locate/fitote

Review

Synergy effects of herb extracts: Pharmacokinetics and

pharmacodynamic basis
Yong Yang a,, Zaiqi Zhang a, Shuping Li a, Xiaoli Ye b, Xuegang Li c, Kai He a,
a
Biomedical Research Center, Huaihua Medical College, Huaihua 418000, China
b
School of Life Science, Southwest University, Chongqing 400715, China
c
College of Pharmaceutical Sciences, Southwest University, Chongqing 400716, China

a r t i c l e i n f o a b s t r a c t

Article history: Herbal medicine, especially traditional Chinese medicine and Ayurvedic medicine have played and
Received 20 July 2013 still play an important role in fighting against various diseases. Emerging clinical studies regarding
Accepted in revised form 17 October 2013 traditional Chinese medicine have provided convincing evidence for the first time to gain credibility
Available online 28 October 2013 and reputation outside China. Although synergistic therapeutic actions of herbal ingredients have
been frequently reported, few reports have offered clear underlying mechanisms. This might be the
Keywords: main reason for the conflicting views with respect to the therapeutic efficacy of medicinal herbs.
Traditional Chinese medicine Therefore, this paper reviews the herb synergisms reported in the recent literature and discusses
Synergy effects thoroughly the mechanisms underlying synergistic actions of herbal ingredients. The authors
Mechanisms
conducted an electronic literature search to detect articles published mainly in the last five years.
Transporters
Articles were included if they pertained to synergy research of ethnomedicines or the active
Drug resistance
Pharmacokinetics and pharmacodynamic compounds derived from them, included verification of synergy effects using modern analytical tools
basis and molecularbiological methods. Results have revealed that the multi-component nature of
medicinal herbs makes them particularly suitable for treating complex diseases and offers great
potential for exhibiting synergistic actions. The mechanisms underlying synergistic therapeutic
actions of herb medicines are (1): different agents may regulate either the same or different target in
various pathways, and therefore cooperate in an agonistic, synergistic way; (2): regulate the
enzymes and transporters that are involved in hepatic and intestinal metabolism to improve oral
drug bioavailability; (3): overcome the drug resistance mechanisms of microbial and cancer cells;
and (4): eliminate the adverse effects and enhance pharmacological potency of agents by
processing or by drugdrug interaction. The exploration of synergistic mechanisms of herbal
ingredients will not only help researchers to discover new phytomedicines or drug combinations but
also help to avoid the possible negative synergy. Further clinical research is required for verifying
these reported drug combinations and discovered synergistic mechanisms.
2013 Elsevier B.V. All rights reserved.

Abbreviations: TCM, traditional Chinese medicine; RC, Rhizoma Coptidis; HPLCSPENMR, high-performance liquid chromatographysolid-phase extraction
nuclear magnetic resonance spectroscopy; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling; 5-HT, 5-hydroxytryptamina; PKC, protein kinase C; PKA,
protein kinase A; MAPK, mitogen-activated protein kinase; ERK, extracellular signal-regulated kinase; PC, phosphatidylcholine; FDA, US Food and Drug Administration; DTL,
docetaxel; YCHT, yin-chen-hao-tang; ALT, alanine amino transferase; AST, aspartate amino transferase; ALP, alkaline phosphatase; MRT, mean residence time; SLC, solute
carrier family; ABC, ATP-binding cassette; OAT, organic anion transporter; OATPs, organic anion-transporting polypeptides; P-gp, p-glycoprotein; BCRP, breast cancer
resistance protein; CYPs, cytochrome P450 enzymes; MDR, multidrug resistance; ROS, reactive oxygen species.
Corresponding author. Tel.: +86 13974530358; fax: +86 0745 2380023.
Corresponding author. Tel.: +86 15115162159; fax: +86 745 2380023.
E-mail addresses: hnyyong@163.com (Y. Yang), hekai69@126.com (K. He).

0367-326X/$ see front matter 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.tote.2013.10.010
134 Y. Yang et al. / Fitoterapia 92 (2014) 133147

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 134
2. Synergistic multi-target effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
3. Improvement of oral bioavailability . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 137
3.1. Drug targeting to intestinal transporters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 137
3.2. Inhibition and induction of cytochrome P450 enzymes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 138
4. Agents that reverse drug resistance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139
4.1. Interactions of agents with resistance mechanism of microorganism . . . . . . . . . . . . . . . . . . . . . . . . 139
4.2. Interactions of agents with multidrug resistance mechanism in cancer . . . . . . . . . . . . . . . . . . . . . . . 139
5. Eliminate the adverse effects and enhance pharmacological potency of agents in herb extracts . . . . . . . . . . . . . . . 141
6. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143
Conict of interest . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143
Acknowledgement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143

1. Introduction due to its potential treatment effects by synergy. For instance,

The last decade has witnessed an emergence and rapid shift
of the paradigm in chemotherapy, involving the gradual
transition from the mono-substance therapy that had long
been advocated with great vehemence to a multidrug therapy.
This is mainly because of the ineffectiveness, resistance
problems and side effects may appear when using synthetic
mono-drugs, especially in the treatment of chronic diseases
such as cancer, atherosclerosis, diabetes and inflammation [1].
The basis of using multidrug therapy for various disorders is the
recognition that for each, more than one mechanism and gene
is identified. For instance, a global genomic analysis has
demonstrated 12 partially overlapping processes that are
genetically altered in the great majority of pancreatic cancers.
In addition, the pathway components that are altered in any
individual tumor vary widely [2]. This has also been docu-
mented in the context of human glioblastoma multiforme,
which is the most common and lethal type of brain cancer [3].
These results indicated that it is difficult to suppress cancer and
other diseases by targeting a single gene or a single pathway
that may alter amongst patients and that can be subject to
mutations. These assays also lend credence to the use of a mixture
of several drugs or herbs to achieve an overall therapeutic
biological effect. Since many leading researchers have advocated
using combination approaches to pursue the optimum therapeu-
tic efficacy and to improve the patient's overall health status [4,5],
the utilization of herbal medicine, which is considered as a
multi-target herb [6], should be optimized.
As one of the largest biodiversity regions in the world, China
has abundant medicinal and aromatic plant species, well
documented traditional knowledge, a long-standing practice of
traditional medicine, and the potential for social and economic
development of medicinal and aromatic plants. Owing to their
reduced side effects, high efficacy and a wide range of pharma-
ceutical activities, the use of TCM is gaining a reputation as a
modern alternative to western medicine or as a complementary
product to maintain health or treat aspects of diseases. Because
the herb extracts consist of complex mixtures of major com-
pounds, concomitant agents and other substances, the complex
multi-component nature of medicinal herbs may serve as a
valuable resource for network-based multi-target drug discovery
polyphenols and terpenoids are two groups of constituents
which are contained in many plant extracts; the former possess a
strong binding ability to different molecular structures like
proteins or glycoproteins, while the latter have great affinities for
cell membranes and therefore, a high potential to permeate
through cell walls of the body or bacteria [7]. Epigallocatechin
gallate (EGCG), the most abundant catechin in tea, is able to
enhance the therapeutic efficacy of temozolomide in patients
with glioblastoma. Chen et al. [8] have revealed that EGCG can
cross the bloodbrain barrier enough to cause chemo-
sensitization in a mouse glioma model. When combined with
temozolomide, EGCG could significantly reduce the expression
levels of glucose-regulated protein 78 in temozolomide-treated
animals, which is a key pro-survival component of the
endoplasmic reticulum stress response system (EGCG alone did
not provide survival benefit). Another interesting finding is that
terpenes provided significant enhancements for the flux and
cumulative amounts of the four model drugs. Among the tested
terpenes, nerolidol provided the highest increase in the flux of
the evaluated drugs. The flux of nicardipine hydrochloride,
hydrocortisone, carbamazepine and tamoxifen were increased
approximately 135-fold, 33-fold, 8-fold and 2-fold respectively
[9]. Hence, under certain conditions, even polyphenols and
terpenoids have not contributed directly to the pharmacological
therapy achieved by the total herb extracts, but the polyvalence
of these compounds can enhance the overall efficacy when
co-administrated with other active compounds. Rhizoma
Coptidis (RC), also known as Huang Lian, is extensively used by
Chinese people for clearing heat as well as purging fire, resolving
phlegm to activate meridians and promoting blood circulation to
remove blood stasis. It is estimated that 1760 prescriptions, in 13
ancient prescription books before Chinese Song Dynasty,
contained RC for the treatment of various diseases [10].
Previously, we recognized that the anti-diabetic effects of RC
total extract were greater than alkaloid-rich fraction, thus the
alkaloids including palmatine, berberine, coptisine, epiberberine,
columbamine, jatrorrhizine and the minor polar constituents
such as magnoflorine, ferulic acid and choline was purified from
RC and the bioactivity of these compounds were preliminarily
studied [11]. Modern pharmaceutical research have demon-
strated that each of these compounds could mediate multiple
 Y. Yang et al. / Fitoterapia 92 (2014) 133147
signaling pathways (summarized in Fig. 1). The possible
synergistic mechanisms underlying this effect will be discussed
in the following section.
Although synergistic therapeutic actions of herbal ingredi-
ents have been frequently reported, few reports have offered
clear underlying mechanisms [12,13]. This phenomenon has
resulted from the absence of high-tech analytical methods and
advanced molecular biological methods. Now, thanks to the
development and the wide use of various detection techniques
as well as the new biological technology, it is more convenient
and effective to elucidate the physical foundation, action
mechanism and prescription compatibility of TCM. For instance,
the introduction of high throughput technologies provides the
opportunity to determine profiles of plants and to systemati-
cally explore the mode of action of combinatory drug regimes
[14]. The hyphenated HPLCSPENMR technique provided an
effective approach for the structural identification of complex
compounds [15]. Besides, the penetration of molecular biology,
gene chip, fluorescence probe, TUNEL assay and differential
mRNA display technology etc. also offer new perspectives on
TCM research at the level of organs, cells and molecules.
Recently, Wagner and Ulrich-Merzenich have systematically
reviewed the synergistic effects of herbal combinations, and
they concluded that the synergy effects can be achieved by
multi-target effects, enhancing the bioavailability, inhibiting the
Fig. 1. Schematic of proposed biological pathways that can be mediated by the main active compounds in Rhizoma Coptidis.
135
antibiotic resistance of bacteria and eliminating the adverse
effects in herb extracts [7]. In this review, based on our early
results, the herb synergisms reported in the recent literature
were summarized and notably, the pharmacokinetics and
pharmacodynamic basis underlying synergistic actions of herbal
ingredients were discussed.
2. Synergistic multi-target effects
When an extracellular signaling molecule activates a cell
surface receptor, signal transduction occurs, and this process
involves the numerous actions of cellular messengers. Though
different constituents may affect various cellular messengers, the
same response may appear in a cell. On the other hand, different
agents may regulate the same target, and therefore cooperate in
an agonistic, synergistic way. RC was used for clearing heat as
well as purging fire, resolving phlegm to activate meridians,
promoting blood circulation to remove blood stasis, removing
dampness and nourishing the kidney to replenish yin by
Chinese people for thousands of years. The pharmacologically
active constituents of RC mainly consist of alkaloids, including
palmatine, berberine, epiberberine, coptisine, jatrorrhizine etc.
exhibited antihyperglycemic [16], anti-inflammatory [17], anti-
Alzheimer [18] and antibacterial activity [19]. However, emerg-
ing evidence suggested that alkaloids from RC also showed
136 Y. Yang et al. / Fitoterapia 92 (2014) 133147
unexpected side effects such as severe arrhythmia, extrapyrami-
dal system reactions, liver function injury and even death
[20,21]. In previous study, we noticed that the combination of
berberine, coptisine, palmatine and epiberberine showed greatly
enhanced anti-diabetic activity and decreased toxicity in HepG2
cell and diabetic mice than berberine used alone [22]. Even so,
the median lethal dose (LD50) in mice of berberine, coptisine,
palmatine and epiberberine (713.57, 852.12, 1533.68 and
1360 mg/kg, respectively.) was smaller than the LD50 value of
RC total extract (2.95 g/kg) [21,23] and the antihyperglycemic
ability of RC total extract was greater than alkaloid-rich fraction
[24]. This raises the question of what contributed to the
synergistic activity of RC total extract. In fact, RC was commonly
consumed as a boiled medicinal soup or medicinal tea to treat
various diseases, and few side effects have been reported.
Therefore, we speculated that the compounds of high polarity,
which appeared in RC water solution, improved the anti-diabetic
activity of RC alkaloids and then showed a tremendous
synergistic activity. This was demonstrated by our further
study: except for four main RC alkaloids, ferulic acid and
choline were separated from the polar parts of RC total extract,
and the combination of alkaloids and these minor constituents
in RC showed synergistic anti-hyperglycemic effects and low
cytotoxicity of RC extract in HepG2 cells and rats [11] (Fig. 2).
Pharmacological studies of ferulic acid revealed that it
has a wide range of biological activities, such as DNA
protection by diminishing the lipid peroxidation and
Fig. 2. The combination of Rhizoma Coptidis alkaloids and minor constituents showed synergistic anti-hyperglycemic effects and low cytotoxicity in rats.
positively modulates the antioxidant status [25], antidiabet-
ic effect by inhibiting monocyte chemoattractant protein-1,
NF-kB expression and by reducing oxidative stress [26,27].
Moreover, it has very recently been shown that administra-
tion of ferulic acid increases levels of 5-hydroxytryptamina
(5-HT), norepinephrine, dopamine and activates 5-HT1A and
5-HT2A receptors in mice. The antidepressant-like effect
of ferulic acid may be mediated through the regulation
of protein kinase C (PKC) as well as protein kinase A
(PKA), Ca2+/calmodulin-dependent protein kinase II (CaMKII),
mitogen-activated protein kinase/extracellular signal-regulated
kinase (MAPK/ERK) and phosphatidylinositol-3-kinase (PI3K)
signaling pathways [28,29]. Most of these pathways play a
significant role in the progression of diabetes and diabetes-
associated complications. Choline, an essential nutrient, can be
obtained from diet or by synthesis de novo in tissues. Recent
studies have indicated that dietary choline could improve animal
growth, enhance digestive and absorptive ability by regulating
target of rapamycin signaling pathway and 4E-BP2 gene
expression in tissues [30]. Besides, choline can alleviate the
cytotoxic effects of interaction between folate deficiency and
radiation exposure [31]. Choline therapy in asthma patients
significantly reduced IL-4, IL-5 and TNF- level, suppressed
cysteinyl leukotriene, leukotriene B4 and 8-isoprostanes (a
biomarker for oxidative stress) as compared to baseline. It
should be noted that phosphatidylcholines also play a major role
in delivering poorly soluble compounds and several other
 Y. Yang et al. / Fitoterapia 92 (2014) 133147
benefits, which gives a rationale for selecting choline as an
adjunct therapy for various diseases [32]. Therefore, it can be
concluded that ferulic acid and choline in RC extracts could
enhance the anti-hyperglycemic effects and reduce the cytotox-
icity of alkaloids in RC extract. More importantly, the use of
mono-compound purified from RC like berberine, palmatine,
coptisine, ferulic acid, etc, has proved to be a very powerful agent
in modulating the numerous cellular signaling pathways
including AMPK signaling [33], NF-kB signaling [34,35], PPAR
signaling [36,37], JNK signaling [38], etc [29,3943]. (summa-
rized in Fig. 1). Taken together, it is not surprising that RC
extracts exhibited greatly amplified pharmaceutical effects
compared with RC alkaloids.
Docetaxel (DTL): the US Food and Drug Administration (FDA)
approved drug for the treatment of metastatic androgen-
independent carcinoma of prostate, has contributed to improved
survival and quality of life in patients suffering from prostate
cancer. Nevertheless, the dose limiting adverse effect of DTL is
febrile neutropenia and anemia. Nagaprashantha et al. [44]
investigated the anticancer effects of the active compound
vicenin-2 from an Indian herb Ocimum Sanctum as well as the
potential synergistic tumor regression ability due to oral DTL and
vicenin-2 combination. The results indicated that vicenin-2 and
DTL co-administration caused greater decrease in the levels of
proliferation marker, Ki67 and angiogenic marker-CD31, while
increasing the tumor suppressor E-cadherin expression to a
greater extent than either of the agents alone. These results were
parallelly demonstrated by in vivo mice xenografts study:
combination of vicenin-2 (1 mg/kg) and DTL (0.01 mg/kg)
induced synergistic effect with significant tumor regression and
reduction in tumor-weight as compared to the vehicle treated
group. Critical signaling proteins like pIGF-1R which is important
for androgen-independent carcinoma of prostate survival and
progression [45], and pAkt, proliferating cell nuclear antigen
(PCNA), cyclin D1, fibronectin were also remarkably inhibited by
co-administration of vicenin-2 and DTL. The author concluded
that due to multi-specific anticancer effects, vicenin-2, either
alone or in combination with other antitumor agents, could
provide a promising treatment for pharmacological intervention
in prostate cancer.
3. Improvement of oral bioavailability
A majority of herb medicines and drugs marketed worldwide
are administered orally. The efficacy of these drugs is dependent
on the oral bioavailability, which in turn, is dependent on drug's
physicochemical properties, formulation design and physiolog-
ical conditions of gastrointestinal tract [46]. Yin-Chen-Hao-Tang
(YCHT), which consists of Artemisia annua L(A), Gardenia
jasminoides Ellis(G), and Rheum Palmatum L(R), has been used
to treat jaundice and liver disorders. According to Chinese
medicinal practice, A used in YCHT is the monarch herb, while G
and R are the minister herb and the assistant or servant herb
respectively. To validate the therapy efficiency of YCHT, the
efficacy of combination therapy with A, G and R (AGR) was
compared to efficacy of mono-therapy with each of the three
components in a rat model of hepatic injury. Compared to the
single substance or partial combination, administration of AGR to
model rats produced significant increase in maximal concentra-
tion (Cmax) and area under the concentrationtime curve (AUC),
while the half-life (t1/2), mean residence time (MRT) were partly
137
prolonged, total body clearance (CL) were reduced markedly.
Simultaneously, the expression of various biochemical indicators
such as alanine amino transferase (ALT), aspartate amino
transferase (AST), and alkaline phosphatase (ALP), malondialde-
hyde, GSH-PX and SOD, etc. were restored to normal levels in
hepatic injury rats by AGR combination. Pharmacokinetic study
further revealed that the therapeutic effect of A on rats with liver
injury can be improved by the addition of G and R: both G and R
can increase the Cmax, prolong MRT and reduce the elimination
rate constant (Ke) and CL of A when compared with A treated
rats. While, as a principal herb in this formula, A can prolong the
time for maximal concentration (Tmax) and MRT, increase the
Cmax and reduce Ke and CL of G. Besides, in the A + R treated
rats, the t1/2 and MRT of R were significantly shorter; the AUC
was obviously decreased; the Ke and CL were increased slightly
by comparison with R group. Taken together, these results
demonstrated that administration of AGR could significantly
increase the bioavailability, slow elimination rate, prolong the lag
time of A and exert a more robust therapeutic effect than any one
or two of the three individual compounds by hitting multiple
targets in a rat model of hepatic injury [47]. The underlying
mechanism of this effect remains unknown, but it is demon-
strated that both the contribution of hepatic metabolism and
intestinal metabolism have been recognized as being clinically
important factors in the pharmacokinetics of orally administered
drugs [48,49]. Strategies aimed at improving the drug absorption
mainly through regulating the physiology conditions of gastro-
intestinal tract, especially by altering the expression of intestinal
drug transporters as well as by inhibiting or inducing cyto-
chrome P450 enzymes (CYPs), which expressed in the human
hepatic and intestinal.
3.1. Drug targeting to intestinal transporters
Targeting to transporters not only helps us understand
previous puzzles on clinical pharmacokinetics but also provides
a unique strategy to improve drug absorption or overcome
unfavorable absorption barrier. Membrane transporters can be
divided into two main classes: solute carrier (SLC) family and
ATP-binding cassette (ABC) transporters. The SLCs are mainly
responsible for the uptake of amino acids, peptides, ions,
xenobiotics, endobiotics, sugars and other biologically active
compounds [50]. Clinical and pre-clinical studies have es-
ablished that co-administration of drugs and organic anion
transporter (OAT, belong to SLC transporters) inhibitor may
result in longer plasma half-life and reduced renal clearance. For
instance, co-administration of probenecid, a known OAT
inhibitor has been demonstrated to diminish the renal clearance
of benzyl penicillin and ciprofloxacin [51]. Yuan et al. [52] found
that the drugdrug interaction between gemcabene and
quinapril involving inhibition of renal OAT and subsequent
elevation in plasma concentrations of quinaprilat (the pharma-
cologically active metabolite of quinapril) is responsible for the
synergistic blood pressure reduction observed with these
compounds. Recently, the main active components of Salvia
miltiorrhiza (Dan Shen) including lithospermic acid, rosmarinic
acid, salvianolic acid A, salvianolic acid B and tanshinol were
demonstrated to elicit significant competitive inhibition on
OAT1 and OAT3-mediated substrate uptake at clinically relevant
concentrations. The results indicated that these compounds
may interact with therapeutic drugs known to be OAT1/OAT3
138 Y. Yang et al. / Fitoterapia 92 (2014) 133147
substrates and enhance their pharmacological activity [53].
Organic anion-transporting polypeptides (OATPs), also belong
to the SLC family, are present in the lipid bilayer of the cell
membrane, acting as the cell's gatekeepers. Each OATP is
predicted to contain 12 transmembrane (TM) domains and
mediates the transport of organic anions across the cell
membrane. Citrus juices have been reported to reduce the
bioavailability of orally administered fexofenadine and increase
the bioavailability of atorvastatin. These interactions are
considered to be caused by the inhibition of intestinal OATPs
[54,55]. Fuchikami et al. [56] evaluated the effects of 15 herbal
extracts on the function of human OATP-B, which is expressed
on human intestinal epithelial cells and involved in the
intestinal absorption of various drugs. They suggested that 7
herbal extracts potently inhibited the function of OATP-B and the
inhibitory effects may be primarily attributable to flavonoid
components. Consistent with this view, recent studies have
indicated that the apigenin, kaempferol and quercetin can inhibit
the OATP1A2 and OATP2B1-mediated uptake of substrates
(atorvastatin and fexofenadine) into HEK293 cells by a compet-
itive mechanism [57]. Since flavonoids are abundantly occurring
in plants and can reach high concentrations in the gut lumen, the
inhibition of OATP-mediated drugs by flavonoids should be
considered as a possible new mechanism for fooddrug or drug
drug interactions. ABC efflux transporters move a wide range of
substrates out of cells. The important ABC transporters in
human include p-glycoprotein (P-gp), multidrug-resistance
associated proteins (MRPs), breast cancer resistance protein
(BCRP), cholesterol transporter (ABCG5/8) and bile salt
efflux pump transporter (BSEP) [58]. Among them, P-gp is
one of the most prevalent efflux transporters expressed in a
number of cancer cells as well as in several organs such as
intestine, liver, kidney and the bloodbrain barrier [59]. P-gp
plays an important role in limiting the intestinal absorption
of its substrates in vivo and inhibition of P-gp leads to the
improvement of bioavailability of orally administrated drugs
and therapeutical agents. Ginsenosides have been found to
be the main components responsible for ginseng's pharma-
cological activity. However, the low oral bioavailability of
ginsenosides has presented a major barrier to the utilization
of those drugs [60]. Recent work has revealed that the active
metabolite of ginsenoside, compound K is a solid substrate of
P-gp, and P-gp mediates the efflux of compound K in vitro
and in vivo. Using P-gp inhibitor verapamil and cyclosporine
A substantially decreased the efflux ratio of compound K in
Caco-2 cells. Administration of compound K to P-gp
deficiency MDR1a/b/ FVB mice also lead to large
enhancement of its absorption and bioavailability [61]. This
synergistic reaction also occurs between phytomedicines.
Sinomenine which derives from the stem of Sinomenium
acutum could significantly improve the bioavailability of
paeoniflorin (derived from the root of Paeonia lactiflora) in rats.
By investigating the intestinal kinetic absorptive characteristics
of paeoniflorin as well as the absorptive behavior influenced by
co-administration of sinomenine and P-gp inhibitors, Chan et al.
[62] suggested that sinomenine in a pattern, which influenced
paeoniflorin's absorption, manifested as similar to that of P-gp
inhibitors. In addition, the inhibition of intestinal breast cancer
resistance protein (BCRP), which restricts the absorption of
xenobiotics, may also increase the systemic availability of
its substrates and various phytochemicals. Natural bioactive
compounds such as flavonoids, chalcones, terpenoids, isothio-
cyanates and nonprenylated rotenoids, are known to inhibit
BCRP [6365]. Tamaki et al. [66] have evaluated the inhibitory
effects of 9 herbal extracts and 23 isoflavonoids on BCRP-
mediated methotrexate (MTX) transport. The results indi-
cated that extracts of soybean, Gymnema sylvestre, black
cohosh, passion flower, rutin and all isoflavonoids strongly
inhibited BCRP-mediated transport of MTX. Most impor-
tantly, the author found that the addition of a 5-hydroxyl or
6-methoxyl moiety tended to potentiate the inhibition
activity [66].
3.2. Inhibition and induction of cytochrome P450 enzymes
Inhibition and induction of CYPs is another main mechanism
of pharmacokinetic drugdrug interactions affecting oral bio-
availability of agents. The cytochromes P450 constitute a
superfamily of heme-containing mono-oxygenases that catalyze
the oxidative metabolism of a wide variety of xenobiotics,
including drugs, plant derived or fungal-derived secondary
metabolites and are therefore of particular relevance for clinical
pharmacology [67]. Only about a dozen enzymes belonging to
the 1, 2, and 3 CYP-families are responsible for the metabolism of
the majority of drugs and other xenobiotics. It is reported that
CYP3 enzymes metabolize more than 50% of the currently
marketed drugs [68]; nevertheless, CYP3 have a strong effect in
reducing the oral bioavailability and in mediating the elimination
of numerous drugs. For example, docetaxel is metabolized by
CYP3A to several metabolites that are all considered to be
therapeutically less effective [69]; CYP3A inhibitors could
improve the bioavailability of docetaxel [70]. CYP enzymes can
also be inhibited by natural products and synergistic therapeutic
actions can frequently arise when one ingredient inhibits the
metabolic clearance of a specific CYP enzyme, the bioavailability
and exposure of other herbal ingredients or co-administered
drugs that act as substrates for the same CYP enzyme is increased
[71]. A recent study has suggested that treatment with
Kaempferia parviflora extract on mice showed various effects to
CYP enzymes: short-term treatment significantly induced
CYP1A1, CYP1A2 enzyme activities; CYP2B enzyme activities
were markedly increased during all Kaempferia parviflora extract
treatment time points, whereas CYP2E1 activity was enhanced
only after long-term treatment. However, Kaempferia parviflora
extract did not affect the CYP3A enzyme activity [72]. However,
the authors have not further investigated which constituent is
responsible for the inhibition of CYP enzymes. The following
researches may provide some useful clues: naturally occurring
flavonoids that constitute the most abundant class of dietary
natural products and herbal remedies, were found to have CYP1
inhibition activities [73,74]. In addition to the inhibition of CYP1,
flavonoids can also exert chemotherapeutic roles by undergoing
CYP1-mediated oxidative metabolism to conversion products
that inhibit tumor cell growth [75]. Thus there is strong
indication that flavonoids act as CYP1 inhibitors and CYP1
substrates. The other secondary metabolites such as alkaloids
(berberine, corydine) and lactones were also reported to have
CYPs inhibitory activities [76,77]. Therefore, it is imperative to
consider the possible herb-transporter or herb-CYPs interactions
before we have a thorough knowledge of the mechanisms of
synergistic actions in herbal ingredients.
 Y. Yang et al. / Fitoterapia 92 (2014) 133147
4. Agents that reverse drug resistance
Considering the problems with ever growing drug resis-
tance [78], it is urgent to investigate the resistance mechanisms
and to discover new combination therapies for treating
infections and cancers. Drug resistance, a phenomenon that
reduces the effectiveness of a drug in curing a disease or
improving patient symptoms can develop against antibiotics,
antivirals or chemotherapeutic agents for cancers. Drug
resistance to chemotherapy remains a major challenge in the
treatment of cancer as well as various infections caused by
fungus and bacteria. Agents that reverse drug resistance may
encompass two aspects: in the first place, antimicrobials
combined with herbal agents that are able to suppress
microbial resistance mechanisms. In the second place,
multidrug resistance (MDR) exists against every effective
anticancer drug and can develop by numerous mechanisms
and targets, while herbal compounds used alone or in
combination with anticancer drugs represent a useful
means of overcoming MDR.
4.1. Interactions of agents with resistance mechanism
of microorganism
There are mainly three defense mechanisms employed by
bacteria against the accumulation of antimicrobial drugs inside
the cell [79]. One is to introduce mutations in the target site,
which often result from spontaneous mutation of a bacterial
gene on the chromosome and selection in the presence of the
antibiotic [80], leading to a reduction in the activity of the drug
towards the microbe. Methicillin-resistant Staphylococcus aureus
(MRSA), which emerged due to transfer of the mecA gene
encoding the novel penicillin-binding protein 2A [81], is
spreading at an alarming rate among infected patients. Previous
study suggested that baicalein exhibits synergy effects with
tetracycline and -lactam antibiotics against MRSA by inhibiting
the outwards transport of tetracyclin of bacteria due to
intervention with the responsible flavonoid-borne gene Tet K
[82]. Chan et al. [83] further demonstrated that baicalein could
restore the antibacterial actions of ciprofloxacin against the NorA
efflux pump overexpressed SA-1199B, as well as inhibit the
enzymatic activity of MRSA specific pyruvate kinase. Another
falconoid, curcumin could markedly reduce the minimal inhib-
itory concentrations of the antibiotics oxacillin, ampicillin,
ciprofloxacin, and norfloxacin used against MRSA [84].
The second is by the synthesis of enzymes that selectively
target and destroy or modify the antibiotics. For instance, the
most common mode of resistance to aminoglycosides is
enzymatic modification of the drug by acetyltransferase from
Escherichia coli, resulting in reduced affinities for binding their
therapeutic target, the bacterial r RNA aminoacyl-tRNA site
[85]. Besides, inactivation of antibiotic by -lactams which
mediated by the overproduction of -lactamases is probably
the most widespread example of enzymatic degradation
resulted in bacterial resistance [86]. Clavulanic acid, which
binds with high affinity to many bacterial -lactamases, has
exhibited great potential to reverse the extended-spectrum
-lactamases and plasmid AmpC -lactamases in combination
with ceftibuten or cephalosporin [87].
The final approach is to reduce accumulation of the
antibiotic inside the microorganism via reducing permeability
139
of the drug through the outer membrane of the cell or via
effluxing the accumulated drug out of the cell. In this context,
natural products also offer a promising strategy to overcome
bacterial resistance mechanisms: 2,6-dimethyl-4-phenyl-
pyridine-3,5-dicarboxylic acid diethyl ester, isolated from
Jatropha elliptica, acts as an inhibitor of the NorA efflux pump
and restores the level of intracellular drug concentration [88].
Artesunate enhances the antibacterial effect of -lactam
antibiotics against Escherichia coli by increasing antibiotic
accumulation via inhibition of the multidrug efflux pump
system AcrABTolC [89]. The major catechin present in green
tea extracts, epigallocatechin-gallate enhances the activity of
tetracycline in staphylococci by inhibiting Tet(K) and Tet(B)
efflux pumps [90]. Another example of the synergistic action
betweenherbalingredientsandantibioticsisgivenbyEumkebet
al.[91].Apigenin and ceftazidime have a synergistic effect
on reversion of bacterial resistance by means of inhibiting
peptidoglycan synthesis and the activity of certain -lactamases,
as well as altering the permeabilization of outer membrane
and cytoplasmic membrane of Enterobacter cloacae. The
authors claimed that the 5, 7-OH group of A ring and one
4-OH group of the B ring in apigenin and naringenin are
important for synergistic activity.
Recently, aside from the known microbial resistance
mechanisms, chemogenomic profiling has become a power-
ful tool that predicts synergy between antifungal or antibac-
terial drug combinations. Strains with reduced fitness in the
presence of a compound are identified, and the genes deleted
in those strains provide information on cellular pathways
targeted by the compound as well as pathways that are
required for conferring sensitivity to that compound [92,93].
If the combination of two synergizing drugs generates a
unique chemogenomic profile of sensitive mutants, then
genes deleted in those mutants provide information on the
drug synergy mechanism. Using this approach, Spitzer et al.
[94] have identified the synergistic interaction of six
compounds with the antifungal drug fluconazole. The result
suggested that fluconazole alone could perturb the genes
which are required for ergosterol biosynthesis (the pathway
targeted by azole antifungal), vesicle-mediated transport,
and membrane organization. Of the six compounds analyzed,
five inhibited the growth of strains carrying deletions in
genes involved in membrane function, vesicle trafficking and
lipid biosynthesis, suggesting that these five compounds cause
general membrane perturbation. Thus, these compounds may
exert their synergistic effect on fluconazole by altering its import
or export, or they may impair the import of exogenous
ergosterol. The sixth compound indicated that it interfered
with sphingolipid biosynthesis, thus suggesting an interplay
between the ergosterol and sphingolipid biosynthetic pathways.
4.2. Interactions of agents with multidrug resistance mechanism
in cancer
The development of MDR to chemotherapy remains a
major challenge in the treatment of cancer. Complex
mechanisms are involved in the resistance of cells to
chemotherapy, but they may be grouped into the following
categories [95,96]: (1) drug uptake reduced by drug efflux
pumps and modification of the lipid bilayer; (2) altered drug
metabolism and drug sequestration; (3) alterations in cell
140 Y. Yang et al. / Fitoterapia 92 (2014) 133147
cycle, increased repair of DNA damage, reduced apoptosis;
(4) abnormally expressed MDR-linked genes; and (5) altered
signal transduction pathways.
Of these mechanisms, the one that is most commonly
encountered is the increased efflux of a broad class of hydro-
phobic cytotoxic drugs that is mediated by ABC transporters. As
discussed in Section 3.1, although MRP2MRP5 (ABCC2ABCC5)
and BSEP (ABCB11) are capable of transporting drugs, the other
ABC transporters, including MDR1 (ABCB1), MRP1 (ABCC1) and
ABCG2, can confer MDR to cancer cells in vitro and play a critical
role in the development of MDR in cancer cells [97]. Emerging
evidences have demonstrated that herb compounds could act
synergistically with anticancer agents and reverse the MDR in
cancer cells. Paclitaxel was approved as a mitotic inhibitor used
to treat patients with lung, ovarian, breast, head and neck cancer.
A preclinical study has demonstrated that P-gp inhibitor
zosuquidar trihydrochloride could increase penetration of
paclitaxel into the brain by 2.15.6 fold [98]. A major ingredient
in Ganoderma lucidum, ganoderic acid, reduces the mRNA and
protein expression level of ABCB1 by inhibiting the activity of
ABCB1 promoter, and also inhibits the expression levels of MRP1
and MRP2. Meanwhile, ganoderic acid stimulates the decline in
mitochondrial membrane potential and up-regulates p-p53, p53,
Bax, caspase-3, and caspase-9, leading to apoptosis in MDR cells
[99]. Moreover, P-gp and MRP1-mediated MDR can also be
reversed by gypenoside aglycon separated from Gynostemma
pentaphyllum [100], Kuguacin J isolated from Momordica
charantia [101], and plant sterols [102].
Drug metabolism enzymes are the second line of cellular
resistance. This process involves three phases. Phase I is
mediated mainly by CYPs and epoxide hydrolases [103]. Drug
species are metabolized and converted into highly mutagenic
aromatic metabolites that can be conjugated by phase II
enzymes including GSTs, UGTs, SULTs, and NATS [104]. These
conjugated metabolites are then effluxed by members of the
ABC transporters, which can be considered as phase III of drug
metabolism [105]. Inhibition of the drug metabolism enzymes
could become a promising strategy for reversing MDR. Zou et
al. [106] evaluated the effects of 25 purified components of
commonly used herbal products on the catalytic activity of
cytochrome P450 isoforms. The results showed ginkgolic acids,
dihydromethysticin, methysticin, hyperforin and quercetin,
etc, significantly inhibited one or more of the human P450
isoforms at concentrations of less than 10 M. Quercetin has
been demonstrated to increase the bioavailability of pioglita-
zone in rats by inhibiting CYP3A [107]. Rosemary extract
enhanced antitumor effect of 5-fluorouracil by down regula-
tion of thymidylate synthetase enzyme and TK1 genes, which is
essential for the synthesis of dTMP and related to 5-fluorouracil
resistance [108]. Other enzymes including CYP1A, CYP2C and
UDP-glucuronosyltransferases could also be inhibited by herb
ingredients or herb extracts [109111].
Chemotherapeutic drugs have been employed to kill
proliferating cells, causing extensive DNA damage that
ultimately leads to cell cycle arrest and cell apoptosis.
However, the efficacy of these therapeutic agents would be
compromised by the ability of cells to repair DNA [95]. The
DNA repair protein, O6-alkylguanine-DNA alkyltransferase
and DNA methyltransferase are important cellular resistance
mechanisms [112,113], their inactivation can reverse resis-
tance to such agents [114,115]. The herb extracts including
Salvia miltiorrhiza, Panax notoginseng, Crataegus pinnatifida
and Polygonum multiflorum [116,117], as well as the pure
herbal-derived compound protocatechuic aldehyde can
weaken the ability of cells to repair damage by inhibiting
DNA synthesis [118].
Numerous MDR-linked genes such as ABC transporters, Bcl2
family genes, metallothionein genes and altered signal transduc-
tion pathways were involved in MDR. In this context, the
dominant approach to resolve MDR is based on reactivation of
apoptotic signaling [119], including: (1) modulating the expres-
sion of inhibitor of apoptosis protein and apoptogenic factors, i.e.,
cytochrome c, smac; (2) strategies for targeting the Bcl2 family
proteins; (3) p53 reactivation; (4) modulation of protein kinase
signaling, mainly Ras/Raf/MEK/ERK and Ras/PI3K/PTEN/Akt
pathwaysbecause both of these pathways are thought to have
anti-apoptotic and drug resistance effects on cells [120]; and
(5) activation of death receptor pathways [121]. Furthermore,
recent data demonstrated that drug resistance is regulated not
only by genetic and epigenetic changes, but also by microRNAs
(miRNAs) [122]. It has been shown that miRNAs play a critical
role in modulation of survival and apoptosis pathways, drug
targets and DNA repair as well as drug transport and MDR (see
details in the review of Giovannetti) [123]. Herb ingredients can
modulate multiple pathways that are related to MDR, thus
resulting in the synergistic therapeutic response. Honokiol, an
active component isolated from TCM magnolia, was demon-
strated to inhibit growth and induce apoptosis of various cancer
cell lines. Combination of honokiol with lapatinib as well as
rapamycin synergistically enhanced the anticancer effects of
honokiol. This may be achieved through the following mecha-
nisms: firstly, honokiol interrupted the cell cycle at G0/1-phase
to S-phase via down-regulating cyclin D1, cyclin E and CDK-2
which have been shown to be crucial for cell cycle progression
through this period [124], and resulted to cancer cell apoptosis.
Additionally, honokiol significantly induced the activation of
caspases-3, -6, and -9, these events then triggered apoptotic
pathways. Secondly, when honokiol combined with human
epidermal growth receptor 2(HER-2) inhibitor lapatinib is able
to overcome therapy resistance which is frequently observed
clinically in HER-2 over-expressed breast cancer and is believed
to be one of the reasons for the resistance to an array of anti-
cancer agents [125]. Lastly, the expression of activated phos-
phoinositide 3-kinases (Akt) has been linked to the promotion of
survival and growth of breast cancer cells. Honokiol treatment
remarkably down-regulated Akt signaling and up-regulated the
expression of phosphatase and tensin homolog deleted on
chromosome ten (PTEN) which negatively regulates Akt activity
[126]. 7, 8-dihyroxycoumarin (Daphnetin) also inhibits human
renal cell carcinoma proliferation by induction of a differentia-
tion response that is mediated by p38 MAPK signaling [127]. Yi
Gan Kang (YGK), which consists of 15 crude herb ingredients,
was recently approved by the FDA phase II clinical trials for the
treatment of hepatitis and non-small cell lung cancer [128].
Studies have demonstrated that YGK significantly repressed E6/
E7 oncogenes at the transcriptional level, with subsequent
reactivation of p53 and p21 expression [129]. Ashwagandha
extracts and its components kill cancer cells by at least five
different pathways: p53 signaling, granulocytemacrophage
colony stimulating factor (GMCFS) signaling, apoptosis signal-
ing, G2-M DNA damage regulation pathway and death receptor
signaling [130]. By activating the death receptor pathway, herb
 Y. Yang et al. / Fitoterapia 92 (2014) 133147
compounds inducing cancer cell apoptosis were frequently
observed. Curcumin induced cell apoptosis in chondrosarcoma
cells through the extrinsic death receptor pathway: Fas, FasL,
and DR5 expression were up regulated and animal studies
revealed a dramatic 60% reduction in tumor volume after
21 days of treatment [131]. Solanum incanum extract containing
solamargine alkaloid as the main active ingredient, induces
apoptosis in human squamous cell carcinoma by up-regulating
the expressions of tumor necrosis factor receptors (TNFRs) and
Fas, and downstream adaptors FADD/TRADD of the TNF-a and
Fas ligand signaling cascades [132].
5. Eliminate the adverse effects and enhance pharmaco-
logical potency of agents in herb extracts
Though TCMs have been used in clinical practice for
several thousands of years, Chinese practitioners also noticed
that some herbal medicine contained toxic compounds and
adverse effects may occur when the herb medicines were
misused. Modern pharmacology studies have identified some
herbal ingredients that are responsible for clinical side
effects. For example, aristolochic acid in Saristolochia fangchi
may lead to renal failure [133]. Dysosma versipellis, which
contains the toxic compound podophyllotoxin, has caused
several cases of thrombocytopenia and liver damage [134].
Despite the limited information available on the chemical
structures and properties of the toxic compounds, our
ancestors have invented a large number of pretreatment
methods to reduce the toxicity of crude drugs, which is called
processing of Chinese materia medica. Processing refers
to any physical and/or chemical treatment by which crude
natural drugs are transformed into materia medica. Addi-
tionally, processing can moderate drastic action, diminish
side effects, modify the energetic properties (flavor, nature,
action), dissipate disagreeable odors and flavors, and so on.
The processing methods of TCM are mainly divided into
cleaning, cutting, and processing practices which include
stir-frying, charring, steaming, boiling, calcining, etc [135].
For instance, using processing methods (Pao zhi), the toxic
ingredients in Aconitum including aconitine, mesaconitine
and hypaconitine can be decomposed into less or non-toxic
derivatives [136]. Despite these methods, the adverse
reactions of herbal compounds can also be eliminated by
co-administration with other phytomedicines. Estragole and
methyleugenol belong to alkenylbenzenes, are found in
many herbs and spices. These two compounds have been
shown to induce hepatomas in rodent bioassays, which can
be ascribed to their bioactivation to alkenylbenzene metab-
olites then resulting in DNA adduct formation. Nevadensin, a
major flavonoid constituent in the Lamiaceae family from
the genus Ocimum basilicum, has been shown to inhibit
sulfotransferase-mediated DNA adduct formation in HepG2
cells and rat hepatocytes exposed to the proximate car-
cinogen 1-hydroxyestragole and 1-hydroxymethyleugenol
[137]. The structureactivity relationship study of nevadensin
has revealed that the presence of C5 and C7 hydroxyl
substituent on the A ring and a C23 double bond in con-
junction with the C4 carbonyl group on the C-ring are required
for the inhibitory action of nevadensin on sulfonation [138].
PHY906, a pharmaceutical grade of traditional Chinese herbal
formulation Huang-Qin-Tang (HQT), composed of four distinct
141
herbs: the roots of Scutellaria baicalensis (S), Glycyrrhiza
uralensis (G), Paeonia lactiflora (P) and the fruit of Ziziphus
jujuba (Z), has been documented for nearly 1800 years for
treating common gastrointestinal distress, including diarrhea,
abdominal spasms, fever, headache, etc. Notably, all four herbs
of the PHY906 formulation are necessary for its biological
effects to be manifested. Experiments were conducted in which
the effectiveness of herbal formulations that lacked one of the
constituent herbs were compared with PHY906 itself in
affecting the antitumor therapeutic efficacy of CPT-11. The
results showed that both S and P play significant roles in
enhancing the antitumor activity of CPT-11, whereas G and Z
play only minor roles. In terms of protecting animals from body
weight loss induced by CPT-11, S and G appear to be the most
important herbs in the PHY906 formulation followed by Z; P
plays no role [139]. Clinical studies for colorectal, liver and
pancreatic cancers have shown that PHY906 enhances the
therapeutic indices and reduces chemotherapy-induced toxic-
ities of a broad spectrum of anticancer agents, such as
5-fluorouracil, etopophos, CPT-11, troxacitabine, clevudine,
paclitaxel, etc. [140]. Previous investigations of the mecha-
nisms of PHY906's antitumor function have explored that
PHY906 can suppress the CPT-11-induced inflammatory re-
sponse by decreasing the infiltration of neutrophils/macro-
phages, decreasing TNF- expression in the intestine, and
decreasing pro-inflammatory cytokine levels in plasma. The
mechanism could be mediated through the inhibition of the
NF-kB pathway and direct inhibition of Cox-2 and iNOS
activities by various constituent chemicals or metabolites of
PHY906. Pharmacogenetic studies suggested that CPT-11 was
found to have a major impact on the gene expression of the
NF-kB family as well as on apoptosis, whereas PHY906 alone
had no significant effect on such pathways. However, PHY906
was shown to reverse the down-regulation of the expression of
genes related to innate immune responses resulting from
CTP-11 treatment [139]. Another example is glycyrrhizin,
which constitutes 1025% of licorice root extract and is
considered as the primary active ingredient in licorice. In vivo
and clinical studies have reported beneficial effects of both
licorice and glycyrrhizin consumption including anti-ulcer,
anti-viral, and hepatoprotective responses [141]. However,
glycyrrhizin also possesses side-effects including hypertension
and edema. Cantelli-Forti et al. [142] have studied the effects of
an aqueous licorice root extract on the pharmacokinetics of the
active component glycyrrhizin, in rats and humans. The results
showed that when glycyrrhizin was provided as a component
of the licorice extract to rats, both the AUC and the Cmax for
plasma levels were significantly reduced. Besides, the results of
clinical trials on aqueous licorice root extracts were similar to
those noted in rats: the use of licorice extracts is safer than
glycyrrhizin is administered alone.
Nevertheless, it should be noted that in most cases, the
adverse effects of TCMs are related to their desired pharmaco-
logical actions. A good example is arsenic, one of the oldest
drugs in the world and has been hailed as a miracle medicine: it
kills people, but saves lives; it can cause some cancers but
cures others such as the acute promyelocytic leukemia (APL).
Molecularly, arsenic binds thiol residues and induces the
formation of reactive oxygen species(ROS), thus affecting
numerous signaling pathways. It also can directly bind
the C3HC4 zinc finger motif in the RBCC domain of PML

Table 1
Examples of pairs of herb compounds or extracts reported to exhibit synergistic effects.
Number Herb 1 (active ingredients)
1 Actein (black cohosh)
2 3-(8(Z),11(Z)-pentadecadienyl) Doxorubicin
catechol (Semecarpus
anacardium)
3 Coumarins and volatile oil
(Angelicae dahuricae)
4 Berberine (RC)
5 Bisbenzylisoquinoli-ne
6 Paris polyphylla
7 Oxyresveratrol (Artocarpus
lakoocha)
8 Berberine (RC)
142
Herb 2 Reported synergistic effect Possible molecular mechanism of synergy Type of synergism Reference
(active
ingredients)
or drugs
Digitoxin Actein enhances the growth inhibitory Both actein and digitoxin could inhibit Na+K+-ATPase activity, Pharmacodynamic synergy [151]
effect of digitoxin on human breast actein potentiated digitoxin's inhibitory effect and the expression because facilitating actions
cancer cells of NF-B promoter, p-ERK, p-Akt and cyclin D1 protein levels could on the same target
be affect by actein
synergistic cytotoxicity with doxorubicin 3-(8(Z),11(Z)-pentadecadienyl) catechol inhibited the Pharmacodynamic synergy [152]
checkpoint kinases; caused blockage of cells at S-phase and G2/M because facilitating actions
Y. Yang et al. / Fitoterapia 92 (2014) 133147
phases. The decrease in the percentage of G0/G1 population was on the same target
also noticed.
Baicalin Coumarins and volatile could increase Both of the two compounds could increase the apparent Pharmacokinetic [153]
the intestinal absorption of baicalin permeability values and absorption rate constant of baicalin and potentiation because of
carrier-mediated active transport may involved in the absorption enhanced bioavailability
process of baicalin rather than P-gp-mediated efflux systems
Fluconazole Berberine exhibits synergy effect with Combination of berberine and fluconazole increased mitochondrial Pharmacokinetics [154]
fluconazole against fluconazole-resistant membrane potential, decreased intra-cellular ATP level, inhibited potentiation because of
Candida albicans ATP-synthase activity, and increased generation of endogenous facilitating actions on
ROS in fluconazole-resistant strains different targets and
pathways
Vinblastine, Bisbenzylisoquinoli-ne reverses the MDR Bisbenzylisoquinoline is able to inhibit the ABC transporters Pgp Pharmacokinetic [155]
paclitaxel in SW620 Ad20 cells. and MRP1 potentiation because of
and eliminated MDR
depsipeptide
5-fluorouracil Paris polyphylla exerts a synergistic Paris polyphylla caused S phase cell cycle arrest, the activation of Pharmacokinetics [156]
and antiproliferative effect by a combination pro-caspase 3, upregulation of Bax and down-regulation of Bcl-2 potentiation because of
Oxaliplatin with 5-fluorouracil and Oxaliplatin proteins expression. Combination of Paris polyphylla and facilitating actions on
5-fluorouracil or Oxaliplatin suppressed the mRNA levels of different targets and
thymidylate synthase and human excision repair pathways
cross-complementing
Acyclovir Synergistic antivirus effects as tested in Oxyresveratrol exhibited the inhibitory activity at the early and late Pharmacodynamic synergy [157]
Vero cells infected with herpes simplex phase of viral replication and inhibited the inhibited late protein because facilitating actions
virus synthesis on the same target
ER Treatment of ER antagonists and the Berberine down regulated, EGFR, HER2, Bcl-2 and COX-2, while, Pharmacokinetics [158]
antagonists crude extract of coptis or berberine up-regulated IFN-b and p21. potentiation because of
conferred synergistic growth inhibitory facilitating actions on
effect on MCF-7 cells different targets and
pathways
 Y. Yang et al. / Fitoterapia 92 (2014) 133147
(promyelocytic leukemia) and PML-RAR, inducing their
homodimerization and multimerization [143]. In addition,
when the herbs are boiled together or alone for medicinal
usages, the hydrophobicity and ion concentration of the
decoction may change. These changes may increase extraction
of constituents from other herbs within the formula and herbs
may react within the decoction medium to create new
chemical structures [144]. Astragali Radix (AR) and Rehmanniae
Radix (RR) are two TCMs widely used in China for treating
diabetes mellitus and its complications. In the foot ulcer animal
model, neither AR nor RR at clinical relevant dose (0.98 g/kg)
promoted diabetic wound healing. However, when they were
used in combination in the ratio of 2:1, they could significantly
reduce the wound area of rats when compared to water group
[145]. As mentioned above, this synergistic interaction may
attribute to the new compounds that were generated during
the boiling process. Lastly, eukaryotic organelles can be used as
an intracellular reaction chamber, in which two compounds
react together to form a novel product [146], or to generate a
bioactive molecule within the organelle that can then diffuse
out of it to improve the efficacy or duration of its action on
other targets [147]. So there is a great possibility that new
chemical drugs can be generated by the reaction of compounds
in herb medicine during enterohepatic circulation.
6. Conclusions
The synergistic effect of complex agents and herb formulas
has already been noticed and applied in clinical practice. Its
most famous example is a cocktail of drugs that is now
commonly employed against HIV infection, because a vaccine
comprising a single antigen will fail to generate broadly reactive
B-cell and T-cell responses that are able to confer protection
against the diverse isolates of HIV-1[148]. Moreover, the
multidrug approach has also been applied in the treatment of
other complex diseases such as cancer, hypertension and
diabetes [149,150]. This strategy is in accordance with the
TCM theory which emphasizes the attainment of health by
maintaining a state of equilibrium between the various systems
and functions of the person. To achieve this goal, Chinese people
use herb prescriptions-in which each medicine can provide its
own special function and form an integrated function for treating
diseases. However, due to the lack of sufficient synergy research
and the specific explanation for the underlying mechanisms,
there have been conflicting views with respect to the therapeutic
efficacy of medicinal herbs. Therefore, this paper reviews the
recent literaturesreported herb synergism and discusses
thoroughly the pharmacokinetics and pharmacodynamic basis
underlying synergistic actions of herbal ingredients. Moreover,
extra examples of herbal synergism are listed in Table 1.
The multi-component nature of medicinal herbs makes
them particularly suitable for treating complex diseases. How-
ever, under no circumstance should the safety issues of herbal
medicine be neglected, since negative interactions among herbs
or between herb and synthetic drugs have been reported
[159,160]. The most common mechanism of drug interactions is
the inhibition of CYP enzymes, which are a major source of
variability in drug pharmacokinetics and responsible for the
biotransformation of most foreign substances including 7080%
of all drugs in clinical use [161,162]. When drugs are co-
administered or one drug inhibits the metabolic clearance of
143
another drug by inhibition of a specific CYP enzyme, drug inter-
actions can arise. For instance, co-administration of the azole
antifungal, ketoconazole, a potent inhibitor of CYP3A, can cause
marked elevations in systemic exposure to compounds meta-
bolically cleared by this enzyme family. Since the increased use
of herbal products worldwide provides another opportunity for
interactions with any co-administered synthetic drugs, it is
imperative to consider the potential herbdrug interaction
before we use CYP-metabolized drugs, especially these with a
narrow therapeutic index.
One of the most important approaches for the moderniza-
tion of TCM is to elucidate the active component in each herbal
medicine, especially the material foundation of compound
prescriptions and their pharmacodynamic mechanisms. After
numerous efforts, a great number of drugs that originated from
plants, including metformin from French lilac [163], colchicine
from Colchicum autumnale, quinine from Cinchona, etc. [164]
have been developed and applied in clinical use. However, the
absence of holistic approaches and integrated evaluation
models can result in studies that are inappropriately designed
or give incorrect results. This is because synergetic effects of
components will be lost in a target driven single lead discovery
program of TCM. For example,5methoxyhydnocarpin, an
amphipathic weak acid, has no antimicrobial activity alone but
potentiated antimicrobial activity with the alkaloid berberine,
which is contained in the same plant by 100 fold [165]. To identify
the bioactive components, one should bear in mind the concept of
systems biology, emphasizing a holistic perspective during
evaluation of the pharmaceutical activity of natural products.
The exploration of synergistic mechanisms of herbal ingredients
will help researchers to discover both new phytomedicines and
drug combinations. Finally, it should be noted that further clinical
research is required for verifying these reported drug combina-
tions and discovered synergistic mechanisms.
Conict of interest
The authors declare that they have no conflicts of interest
to disclose.
Acknowledgement
Thanks are due to Hunan Provincial Science and Technology
Department (2013FJ3062) and Hunan Provincial Education
Department (13B088) for financial assistance and we would
like to thank Beth Sample for a critical reading of the manuscript.
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