Introduction
In the last two decades, metabolomics are considered important tools to be applied
and utilized to understand the biology of an organism and its response to environmental
stimuli or genetic perturbation. Metabolomics (a flowchart is shown in [Fig. 1]) was originally proposed as a method of functional genomics [1], but its utility extends well beyond that – it is useful whenever an assessment
of changes in metabolite levels is needed. Metabolomics (or metabonomics) is used
for assessing responses to environmental stress [2], [3], comparing mutants [4] and different growth stages [18], [19], for drug [5] and natural products [22] discovery, toxicology [6], [7], nutrition [8], [9], [10], [11], [12], genetic manipulation [13], cancer [14], [15], [16], [17], and diabetes [20], [21]. Indeed, metabolomics is the study of global metabolite profiles in a system (cell,
tissue, or organism) under a given set of conditions and has its roots in early metabolic
profiling studies, but is now a rapidly expanding field of scientific research, which
has justifiably taken its place alongside genomics, transcriptomics, and proteomics
as one of the latest and most exciting “-omic” sciences [23]. The global metabolite profiling involves measuring low molecular-weight metabolites
(< 1 kDa) in complex biofluids/tissues to study perturbations in response to physiological
challenges, toxic insults, or disease processes [24]. Metabolites are the end products of cellular regulatory processes and can be regarded
as the ultimate response of biological systems to control genetic or environmental
changes [25]. A deeper understanding of global perturbations in biochemical pathways in complex
diseases could provide valuable insights about the mechanisms of disease. In addition,
metabolomics has the potential to enable mapping of early biochemical changes in disease
and hence provide an opportunity to develop predictive biomarkers that can trigger
earlier interventions.
Fig. 1 Flowchart of metabolomic analysis.
TCM, a complex medical science, reflects traditional Chinese culture and philosophical
principles, embodies rich dialectical thought, places the human body into a large
system for observation and adjusts humans to remain in a healthy status. Entering
the 21st century, TCM is getting more and more popular in the whole world for improving
health conditions of human beings and preventing or healing diseases, and is especially
showing great advantages in early and combination intervention, personalized therapy,
etc. The international community has attached increasing importance to TCM [26]. However, like almost all other ethnopharmacologies, TCM faces severe challenges
and suffers from insufficient scientific research owing to the lack of modern and
technologic approaches; this restricts the development of TCM in the world. Chinese
medicine is not yet an integral part of the standard healthcare system in Western
countries due to a lack of scientific evidence for its efficacy and safety as well
as a language and cultural barrier [27]. Fortunately, metabolomics adopts a “top-down” strategy to reflect the function
and metabolic changes of complete organisms caused by interventions in a holistic
context. This property coincides with the holistic thinking of TCM and has the potential
to impact our understanding of Chinese medicine theory. Recently, a wide range of
metabolomic analytical techniques are widely used in the modern research of TCM. Metabolomic
analysis will facilitate the modernized study of TCM syndromes, provide an in-depth
understanding of the TCM theory, help predict the disease on-set and achieve a comprehensive
evaluation of clinical efficacy, safety, and action mechanisms of TCM formulae. Metabolomics
combined with TCM methodologies will provide a new pathway and methodology for the
study of complicated systems theory of TCM and its modernization. Especially the advances
in the high-throughput and comprehensive research technologies and the idea of metabolomics
provides new strategies for the analysis of active components in the formulae of TCM
in vivo. Furthermore, the initiatives of metabolomics may pave a new way to explain the action
mode of TCM in light of modern sciences and so contribute to establish a new technique
platform for evaluating the efficacy of the TCM formulae. It is believed that with
the further development of metabolomic analytical techniques, especially multi-analysis
techniques, metabolomics will greatly promote TCM research and be beneficial to its
modernization as well as establish international standards for it. Thus, we draw attention
to applicable approaches of using metabolomic techniques to resolve special TCM issues
and focuse particularly on the potential of metabolomics to contribute to biomarker
discovery in TCM research.
Advantages of Metabolomics
The study of biological systems in a holistic manner (systems biology) is increasingly
being viewed as a necessity to provide qualitative and quantitative descriptions of
the emergent properties of the complete system. Systems biology performs studies focused
on the complex interactions of system components, emphasizing the whole system rather
than the individual parts. Metabolomics is one functional level tool being employed
to investigate the complex interactions of metabolites with other metabolites but
also the regulatory role metabolites provide through interaction with genes, transcripts,
and proteins (e.g., allosteric regulation). Technological developments are the driving
force behind advances in scientific knowledge. Recent advances in the two analytical
platforms of mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy
have driven forward the discipline of metabolomics [28], [29], [30], [31], [32]. Technological advances in NMR and mass spectrometry have opened a new chapter in
biochemistry by using metabolomics as an approach to study metabolism and its regulation
in relation to genetic, disease, and environmental factors. NMR is one of the most
commonly used technologies in metabolomics research, providing detailed information
on the molecular structure and probing metabolite molecular dynamics and mobility
[33]. The high selectivity of mass spectrometry with low-detection limits makes mass
spectrometry an ideal tool for metabolomic applications. It requires separation of
the metabolic components using either gas chromatography after chemical derivatization
or ultra-performance liquid chromatography (UPLC). UPLC/MS is often used to obtain
the largest possible biochemical profile information subset. It is a sensitive tool
that can be used to characterize, identify and quantify a large number of compounds
in a biological sample where metabolite concentrations might cover a broad range of
information with regard to disease pathophysiology [34], [35], [36]. All metabolomic studies result in complex multivariate datasets that require visualization
software as well as chemometric and bioinformatic methods for interpretation. The
application of software tools for the analysis of the information contained in a database
can identify the signature of a disease and predict its risk and progression.
One of the major benefits of metabolomics in the study of disease and drug therapy
is that metabolic profiling can usually be achieved using urine or plasma samples.
The accessibility of urine and plasma clearly makes these samples ideal for large-scale
research [37]. Metabolomics data can be analyzed with a range of statistical and machine-learning
algorithms. These algorithms can be classified within two major classes: unsupervised
and supervised [38]. They can be useful in the identification of biomarkers [39], [40]. Examples of unsupervised methods that have been routinely used in analyzing molecular
fingerprinting data include principal component analysis (PCA) and self-organizing
maps [41].
Metabolomics has become useful in many medicine areas as an aid to disease diagnosis
or staging and as a tool to predict or monitor treatment response or toxicity. Considerable
interest in the field of metabolomics is that of personalized health care, whereby
an individual's drug treatment is tailored so as to achieve maximal efficacy while
avoiding adverse drug reactions. Another more recent approach has been to use metabolomics
to predict the metabolism of a dosed substance based on a pre-dose metabolic profile
[42]. Metabolomics holds a comprehensive and non-invasive analysis of metabolic biomarkers
that could detect early-stage disease, identify residual disease post-surgery and
help to monitor treatment response. Additionally, metabolomics can be seen as bridging
the gap between genotype and phenotype, providing a more comprehensive view of how
cells function, as well as identifying novel or striking changes in specific metabolites
[25], [43]. In the future, metabolomics may enable us to develop new approaches for improving
the health conditions of human beings and preventing or healing diseases, and is especially
showing great advantages in early and combination intervention, personalized therapy,
etc. [44], [45].
Metabolomic Applications of TCM
Metabolomic evaluation of formulae
Wang et al. evaluated metabolomic characters of the hepatotoxicity induced by alcohol
and the intervention effects of Yin Chen Hao Tang (YCHT), a classic traditional Chinese
medicine formula composed of Flos Artemisiae, Gardeniae Jasminoidis, and Fructus and
Radix et Rhizoma Rhei for the treatment of jaundice and liver disorders in China [46]. The greatest difference in metabolic profiling was observed in alcohol-treated
rats compared with control and YCHT-treated rats ([Fig. 2]). The number of positive ions, m/z 664.3126 (9.00 min), was elevated in the urine of alcohol-treated rats, whereas ions
with m/z 155.3547 (10.96 min) and 708.2932 (9.01 min) were at a lower concentration compared
with those in the urine of control rats. However, the number of these ions was not
statistically different between control and YCHT-treated rats ([Fig. 3]). The ion with m/z 664.3126 was found to correspond to ceramide (d18:1/25 : 0),
providing further support for an involvement of the sphingomyelin signaling pathway
in alcohol hepatotoxicity and the intervention effects of YCHT ([Fig. 4]). More recently, Wang et al. explored the excretion pattern of low molecular mass
metabolites in the male Wistar-derived rat model of kidney yin deficiency induced
with thyroxine and reserpine as well as the therapeutic effect of Liu Wei Di Huang
Wan (LW), one of the most important Chinese formulas consisting of six herbs including
Rehmannia glutinosa Libosch. (family: Scrophulariaceae), Cornus officinalis Sieb. (family: Cornaceae), Dioscorea opposite Thunb. (family: Dioscoreaceae), Alisma orientale (G. Samuelsson) Juz (family: Alismataceae), Poria cocos (Schw.) Wolf (family: Polyporaceae), and Paeonia suffruticosa Andrews (family: Paeoniaceae), and widely used in eastern Asia for treating kidney
yin deficiency in China [47]. The results showed that the changes in metabolic profiling were restored to their
baseline values after treatment with LW according to the PCA score plots ([Fig. 5]), indicating 20 ions (8 in the positive mode, 14 in the negative mode, and 2 in
both) as “differentiating metabolites” ([Fig. 6]). PCA score plots separated urine samples into different blocks, and samples subjected
to the same treatment were located on the same trajectory, indicating that treatments
have greatly disturbed the normal urine metabolic profiles of rats. There were different
phenotypes of metabolites based on HPLC‐UV urinary profiling after administration
of LW pills or carrageenan-stimulated inflammation model, and those could be conveniently
discriminated by PCA. In addition, the results also showed that LW Pills could restore
the metabolite network that was disturbed by inflammation, which would be a proof
of the therapeutic efficacy of LW pills on inflammation from a metabolomics study
[48]. A urinary metabolomics method based on the UPLC‐MS was used to evaluate the efficacy
and study the action mechanism of Xindi soft capsules, consisting of sea buckthorn
flavanoids (e.g., quercetin, kaempferol, and isorhamnetin), which is a TCM preparation
against blood stasis [49]. Compound Danshen tablets, a herbal (Salvia miltiorrhiza Bge.) compound preparation, presented protective effects on myocardial ischemia by
reversing potential biomarkers to sham levels, especially for the four metabolites
in the pathway of purine metabolism (hypoxanthine, xanthine, inosine, and allantoin)
[50]. Damage of mitochondria, disorder of energy metabolism, and osmoregulation were
observed in a cyclophosphamide-caused blood deficiency model by an NMR-based metabolomics
method. Siwutang, a novel prescription consisting of Paeonia lactiflora Pall, Angelica sinensis, Rehmannia glutinosa, and Rhizoma Chuanxiong could improve these effects [51].
Fig. 2 Resulting score plots from PCA of reversed-phase UPLC‐ESI‐TOF‐MS data obtained from
(A) control rats, (B) alcohol-treated rats, (C) YCHT-treated rats urine samples collected from day 7. Symbols: control rat (□),
alcohol-treated rat (×), and YCHT-treated rat (+).
Fig. 3 Resulting loadings plot from PCA of reversed-phase UPLC‐ESI‐TOF‐MS data obtained
from control rats, alcohol-treated rats, and YCHT-treated rats urine samples collected
from day 7.
Fig. 4 The proposed metabolic pathway for explanation of the relationship between alcohol
hepatotoxicity and the increased content of ceramide (d18 : 1/25 : 0) in urine.
Fig. 5 PCA score plots of urine samples collected from different treatment groups of rats
(a) in negative ESI mode and (b) in positive ESI mode. Key: █ Control; █ LW; █ Model 1; █ Model 2; █ Sanbu; █ Sanxie.
Fig. 6 Component loadings in (a) negative ESI mode and (b) positive ESI mode.
Metabolomic analysis of CHM
CHM has long been used for disease prevention and therapy in China and is becoming
increasingly important in the West [52]. Herbal medicine has been the source of many drugs used in modern therapeutics,
and particularly in the case of anticancer drugs, more than 50 % originally came from
natural products. However, due to the painstaking way of conventional lead-finding,
the attention towards CHM has been deviated in the last decades. A new strategy for
the detection of active compounds is necessary to get natural product research out
of its stalemate. Metabolomics, with its holistic approach and the possibility it
provides for the simultaneous detection of all sorts of metabolites, has the potential
to be instrumental for this new approach. A combined GC/MS and LC/MS metabolic profiling
strategy indicates that Tripterygium wilfordii Hook. F caused a time-dependent toxic effect at a high dose as revealed by the perturbed
metabolic regulatory network involving disorders in energy metabolism, elevated amino
acid, and choline metabolism pathways, as well as altered structure of gut flora [53]. Urinary metabolic perturbations associated with liver toxicity induced by Huang-yao-zi
(root of Dioscorea bulifera L.) were studied using NMR to determine the correlations between metabolomic profiling
and histopathologic/biochemical observations and to discover biomarkers for liver
toxicity, indicating metabolic changes observed in urine samples in response to Huang-yao-zi
treatment. In addition, the mechanism associated with oxidative injury of hepatic
mitochondria was investigated [54]. HPLC‐MS/MS-based metabolomics method was used to find the possible biomarker of
Rhizoma Coptidis in rat urine. 169 kinds of biomarkers were found. The result was
consistent with the pharmacological effects of R. Coptidis, such as anti-inflammatory,
anticentral nervous system, and energy metabolism inhibition [55]. Berberine might play a pivotal role in the treatment of type 2 diabetes through
downregulating the high level of free fatty acids. Comprehensive metabolomic measurements
are potentially very useful for studying the mechanisms of action of traditional Chinese
medicines [56]. Combined NMR and LC‐DAD‐MS analyses reveal comprehensive metabolomic variations
for three phenotypic cultivars of Salvia miltiorrhiza Bunge [57]. NMR-based metabolomics is an attractive method for nonselective and comprehensive
analysis of Ginkgo extracts, which are very complex mixtures prepared from raw leaf extracts by a series
of extraction and prepurification steps [58]. Ginkgo biloba leaves exert multidirectional lipid-lowering effects on the rat metabonome, including
limitation of the absorption of cholesterol, inactivation of HMGCoA, and favorable
regulation of profiles of essential polyunsaturated fatty acid [59].
Recently, changes of metabolites in rat urine after treatment with Aristolochia fangchi decoction were studied by the metabolomic method [60]. High-dose Aristolochia fangchi can induce nephrotoxicity, and its seriousness corresponds to the duration of administration.
Aristolochia fangchi may also have toxicity on the liver. The results suggested that this metabolomic
approach is a promising methodology for the rapid in vivo screening of nephrotoxicity associated with ingesting multi-ingredient medicinal
herb supplements [61]. Aristolochic acids, naturally present in Aristolochia plant species that have been used in CHM containing a mixture of varying herb species,
were identified by UPLC‐MS-based methodology [62]. A LC/MS metabolomics approach was applied to characterize the aging of rats and
the antiaging effect of total flavones of Epimedium, a traditional Chinese medicine, indicating that aging could be characterized by
changes of lipid metabolism and accumulation of free radicals. The antiaging effects
of total flavones of Epimedium might be due to the intervention on lipid metabolism and its property of antioxidation
[63]. Phenotype of aging at different levels demonstrates a common age-dependent trend.
Epimedium flavanoids can reverse this age-dependent change at different levels in a synchronous
manner [64]. The total flavones of Epimedium administration can markedly influence the aging process and exert antiaging effects,
which might due to the melioration of pyruyate metabolism and oxidative phosphorylation
[65]. Significant differences in endogenous metabolite profiles were observed in the
intervention rats, and the abnormality of metabolism recovered towards the normal
level after administration with Epimedium brevicornum extract. Four active constituents of Epimedium brevicornum Maxim were found in the blood circulation of kidney-deficient rats and two of the
metabolites in the urine. It suggests that the metabolomic approach is a potentially
powerful tool to explore the therapeutic basis and to clarify the possible action
mechanism of TCM herbs [66].
Metabolic changes in Wistar rats caused by Aconitum alkaloids aconitine, mesaconitine, and hypaconitine, which are the main toxic components
of the traditional herbal medicine Fu Zi (Aconitum carmichaelii Debx.) were investigated by means of integrated analysis of two metabolomic approaches.
Metabolites with significant changes or with a tendency to change in the aconitine
and mesaconitine groups were dissimilar, suggesting a possible difference in the acute
toxicity mechanisms of these alkaloids [67]. A metabolomic investigation of intoxication with Aconitum sp. alkaloids was carried out: they can cause metabolic disorders in rats. The toxicity
and corresponding mechanism of hypaconitine was different from those of aconitine
and mesaconitine, based on the differences of perturbed metabolic patterns between
groups [68]. The effect mechanism and potential biomarkers of the toxicity of Hei-Shun-Pian,
the processed lateral root of Aconitum carmichaelii Debx. (Ranunculaceae), on the metabolic profile of rats, suggests a toxic effect
of Hei-Shun-Pian on the rat heart in a dose-dependent manner [69]. Artemisia afra has been used as an infusion to treat malaria throughout the southern parts of Africa,
in much the same way as the antimalarial plant Artemisia annua in China. Liu et al. had used metabolomics to investigate the ethnopharmacological
use of Artemisia afra with NMR spectroscopy and multivariate data analysis [70]. Their findings show that there is no in vitro activity, and a list of the identified metabolites causing the metabolic differences
is presented. Artemisinin has been proven to be an effective antimalarial compound,
especially for chloroquine-resistant and cerebral malaria. In order to get new clues
about artemisinin biosynthesis, metabolic profiling by GC and GC‐MS was applied to
compare the secondary metabolites of two Artemisia annua L. genotypes. It could be shown that there were clear differences in terpenoids and
artemisinin metabolism between different growth stages and genotypes [71].
Metabolite profiling of five medicinal Panax herbs, including Panax ginseng (Chinese ginseng), Panax notoginseng (Sanchi), Panax japonicus (Rhizoma Panacis Majoris), Panax quinquefolium L. (American ginseng), and P. ginseng (Korean ginseng), was performed using UPLC‐MS and a multivariate statistical analysis
technique. PCA of the analytical data showed that the five Panax herbs could be separated into five different groups of phytochemicals. The chemical
markers such as ginsenoside Rf, 20(S)-pseudoginsenoside F11, malonyl ginsenoside Rb1,
and ginsenoside Rb2, accountable for such variations, were identified through the
loadings plot of PCA, tentatively by the accurate mass of TOF/MS, and partially by
a reference standard [72]. Six different types of ginseng roots from China and Korea could be easily differentiated
by NMR-based metabolomics [73]. Pharmacodynamic effects of the ginsenoside Rg3 on the urine metabonomes of healthy
and liver-tumor-bearing rats have been investigated. Seventeen biomarker candidates
including three apolar metabolites were detected for global analysis of highly complex
biosamples [74]. This approach may not only increase the number of discovered biomarkers but consequently
improve the comprehensive information on metabolic changes.
Liang et al. used LC/MS to analyze 16 saponins simultaneously, and the developed methodology
could effectively break the application bottleneck on the quantitative analysis of
multi-component LC/MS data and be applied widely in related fields for multi-component
analysis, especially in CHM research [75]. Concurrently, a number of metabolites involved in glucose metabolism, citric acid
cycle, and amino acid metabolism were affected immediately after the intake of green
tea, and the proposed approach provided a more comprehensive picture of the metabolic
changes after intake of green tea in human urine [76]. Interestingly, green and black tea intake had a different impact on endogenous
metabolites in urine and plasma. Green tea intake caused a stronger increase in urinary
excretion of several citric acid cycle intermediates, which suggests an effect of
green tea flavanols on human oxidative energy metabolism and/or biosynthetic pathways
[77]. The metabolic strategy has shown its potential in optimization of harvest time
and chemical markers screening of tangerine peels, herbal materials of two coupled
traditional Chinese medicines, Pericarpium Citri Reticulatae and Pericarpium Citri
Reticulatae Viride, which will open a wide perspective in the analysis of “coupled
TCMs” [78]. Major metabolite of both arecoline and arecaidine, N-methylnipecotic acid, is a
novel metabolite arising from carbon-carbon double-bond reduction. Another unusual
metabolite found was the monoacylglyceride of arecaidine. It was shown which role
is played by these uncommon metabolites in the toxicology of arecoline and arecaidine
[79]. Evocarpine, from the Chinese herb Evodia rutaecarpa, is not transported by p-gp and showed only slight toxicity at the highest test concentration
of 30 µM [80]. Evodiae rutaecarpa has changed the endogenous metabolites of rats and can provide the base for the further
research on the interpretation of drug property [81]. Molecular compositions of rosemary (Rosmarinus officinalis L.) extracts and their dependence on extraction solvents, seasons, and drying processes
were systematically characterized using NMR spectroscopy and multivariate data analysis.
Results showed that the rosemary metabonome was dominated by 33 metabolites including
sugars, amino acids, organic acids, polyphenolic acids, and diterpenes, among which
quinate, cis-4-glucosyloxycinnamic acid, and 3,4,5-trimethoxyphenylmethanol were found
in rosemary for the first time [82]. It can be concluded that the metabolomics are a potentially powerful tool to explore
the therapeutic basis of TCM herbs.
Metabolomics in the study of mineral medicine
An important traditional Chinese mineral medicine, Cinnabar, has been widely used as a Chinese patent medicine ingredient for sedative therapy,
induced disturbance in energy metabolism, amino acid metabolism and gut microflora
environment, as well as slight injury in the liver and kidney, which might indirectly
result from cinnabar induced oxidative stress [83]. The time-dependent biochemical variations induced by realgar were achieved using
pattern recognition methods. They illustrated the high reliability of the NMR-based
metabolomic approach for the study of biochemical effects induced by mineral medicine
[84].
Metabolomics in acupuncture
Chinese acupuncture, handed down from the ancient times, still survives in clinical
practice nowadays. At present, acupuncture has attracted extensive attention in the
domestic and overseas circles in the TCM field. Numerous experimental studies have
demonstrated that acupuncture can correct various metabolic disorders such as hyperglycemia,
overweight, hyperphagia, hyperlipidemia, inflammation, altered activity of the sympathetic
nervous system, and insulin signal defect, all of which contribute to the development
of insulin resistance [85]. In addition, acupuncture has the potential to improve insulin sensitivity. These
studies have revealed the mechanisms responsible for the beneficial effects of acupuncture,
though further investigations are warranted. Functional dyspepsia acupuncture-treated
patients showed significantly changed levels of leucine/isoleucine, lactate, and glucose,
and a slightly changed lipids level towards those of the healthy controls, demonstrating
the therapeutic effects of acupuncture on the relief of functional dyspepsia symptoms;
this is a NMR-based metabolomic approach in the study of biological effects of acupuncture
[86].
Metabolomics in Chinese medicine syndromes
Metabolomics, the analysis of the metabolite profile in body fluids or tissues, is
being applied to the analysis of a number of different diseases as well as being used
in following responses to therapy. There is good reason to think that metabolomics
will find particular utility in the investigation of disease, which may have tissue
specific and systemic components. Metabolomic analysis can inform us about ocular
or other body fluids and can therefore provide new information on pathways and processes
involved in these responses.
Metabolomics has been used in several animal models of human disease. A recent study
was conducted by Qiu et al. to investigate the metabolic responses to Herba Cistanches
intervention in a rat model of hydrocortisone-induced “kidney-deficiency syndrome”.
It revealed that Herba Cistanches caused a systemic recovery from hydrocortisone-
induced metabolic perturbation in rats [87]. The TCM concepts of “Xinxueyuzuzheng (heart blood stasis obstruction pattern)”
and “Qiyinliangxuzheng (qi and yin deficiency pattern)” for myocardial ischemia rat
models were constructed by Yan et al. Endogenous metabolites in rat plasma were identified,
such as valine, serine, threonine, ornithine, hydroxyproline, lysine, 2-hydroxybutanoic
acid, 3-hydroxybutanoic acid, galactofuranose, and inositol. These compounds were
indicated as the potential biomarkers, suggesting that the two above-mentioned patterns
are involved in dysfunction, oxidative stress, energy metabolism, and amino acid metabolism
[88]. Chen et al. described the metabolomic study of a biochemical modification in vivo induced by a high dose of hydrocortisone, which led to a unique pathologic condition
similar to the “kidney deficiency syndrome”, an early stage of obesity and diabetes
in TCM [89]. In the study, 24-h urine was collected pre-dose and at days 1, 3, 7, and 10 post-dose
after rats were injected with hydrocortisone at 1.5 mg/100 g. The acquired data were
transferred into Matlab to be processed using PCA. Results indicated clear and consistent
biochemical changes following hydrocortisone intervention. It suggested that metabolomic
approach could be used as a potentially powerful tool to investigate the biochemical
changes of certain physiopathologic conditions such as metabolic syndrome, as an early
diagnostic means.
Luo et al. determined the changes of the plasma metabolic phenotype in rats with chronic
restraint stress (rats with syndrome of liver qi stagnation and spleen deficiency)
[90]. Metabolomic analysis of rat plasma revealed different metabolic spectra between
stress and control groups, which were consistent with alterations of in vivo metabolisms in rats under stress stimuli. Compared with the control group, rats with
repeated stress displayed significant changes in spectral peak shapes of acetate,
lactate, tyrosine, and low-density lipoprotein. These altered metabolites can be used
as biomarkers of syndrome of liver qi stagnation and spleen deficiency. Therefore,
metabolomics is an approach with good development prospects for studies of TCM syndromes.
