Access to this full-text is provided by PLOS.
Content available from PLOS One
This content is subject to copyright.
RESEARCH ARTICLE
Integrated global and unique metabolic
characteristics to reveal the intervention
effect of Yiyi decoction on acute pancreatitis
Guanwen Gong
1☯
, Yongping Wu
2☯
, Yanwen Jiang
3
, Yuan CaoID
3
*
1Department of General Surgery, Affiliated Hospital of Nanjing University of Chinese Medicine, Jiangsu
Province Hospital of Chinese Medicine, Nanjing, China, 2Laboratory of Chemistry, Jiangsu Provincial
Institute of Materia Medica, Nanjing, China, 3Department of Pharmacy, Affiliated Hospital of Nanjing
University of Chinese Medicine, Jiangsu Province Hospital of Chinese Medicine, Nanjing, China
☯These authors contributed equally to this work.
*yfy0012@njutcm.edu.cn
Abstract
Yiyi decoction is a Chinese herbal formula for the treatment of acute pancreatitis that has
been used in clinical practice for decades. A previous study has suggested that resveratrol,
emodin, rhein and their derivatives might be the potential pharmacodynamic components in
Yiyi decoction, and researchers have proposed that resveratrol, emodin and rhein are candi-
date markers for quality control. The present study investigated the intervention effect of
Yiyi decoction and its effective components on murine acute pancreatitis using metabolomic
approach that integrated global and unique metabolic characteristics. First, serum metabo-
lomics based on the platform of ultra-high performance liquid chromatography coupled with
quadrupole time-of-flight mass spectrometry was performed to assess metabolic changes
in experimental acute pancreatitis. Second, an in-depth analysis of bile acid metabolism
was performed based on an in-house database. Finally, an integrated analysis of the inter-
vention effect of Yiyi decoction and its effective components in response to these metabolic
perturbations was performed. As a result, 39 potential biomarkers for the pathogenesis of
acute pancreatitis, mainly phospholipids, fatty acids, bile acids and lipoylcarnitines, were
screened and annotated. Integrated analysis revealed that the metabolic disorders in acute
pancreatitis mice were reversed by Yiyi decoction primarily via regulating glycerophospholi-
pid metabolism, bile acid biosynthesis, carnitine synthesis and fatty acid metabolism. Yiyi
decoction components may effectively target the migratory metabolome. Histopathological
and biochemical analyses suggested that Yiyi decoction maintained the gut barrier function
and inhibited inflammatory cytokines, thus exert anti-acute pancreatitis effects. The present
study utilized an approach that integrated global and unique metabolic characteristics to elu-
cidate the underlying mechanisms of Chinese herbal formulas from a metabolomics
perspective.
PLOS ONE
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 1 / 17
a1111111111
a1111111111
a1111111111
a1111111111
a1111111111
OPEN ACCESS
Citation: Gong G, Wu Y, Jiang Y, Cao Y (2024)
Integrated global and unique metabolic
characteristics to reveal the intervention effect of
Yiyi decoction on acute pancreatitis. PLoS ONE
19(11): e0310689. https://doi.org/10.1371/journal.
pone.0310689
Editor: Chun-Hua Wang, Foshan University, CHINA
Received: April 27, 2024
Accepted: August 30, 2024
Published: November 21, 2024
Copyright: ©2024 Gong et al. This is an open
access article distributed under the terms of the
Creative Commons Attribution License, which
permits unrestricted use, distribution, and
reproduction in any medium, provided the original
author and source are credited.
Data Availability Statement: All relevant data are
within the paper and its Supporting Information
files.
Funding: National Natural Science Foundation of
China (Nos. 81704083 and 82073983) Natural
Science Foundation of Jiangsu Province (Nos.
BK20161086), Special Research Project of Jiangsu
Provincial Administration of Traditional Chinese
Medicine (ZT202108). The funders had no role in
study design, data collection and analysis, decision
to publish, or preparation of the manuscript.
Introduction
Acute pancreatitis (AP) is a common gastrointestinal condition of the abdomen that leads to
hospitalization, it is characterized by premature activation of pancreatic enzymes and local
and/or systemic inflammation [1]. The global incidence ranges from 13–45 cases per 100 000
individuals annually, and has continued to increase during the last decade [2]. The typical clin-
ical symptoms of AP include persistent and sharp abdominal pain, accompanied by distension,
nausea and vomiting [3]. Approximately 20% of patients with severe AP may develop pancre-
atic necrosis, sepsis, and multiple organ dysfunction syndrome (MODS), which is the most
severe complication in terms of mortality [4]. Specific agents for the treatment of AP are still
unavailable despite the substantial burden of this disease [3,5]. Therefore, effective drugs that
can combat AP and ameliorate its clinical symptoms are urgently needed.
The therapeutic effects of rhubarb-based Chinese herbal formulas (CHFs) such as Dachengqi
decoction and Qingyi decoction have been validated in clinical practice, and recommended in
the updated guidelines for the diagnosis and treatment of AP in China (2021) [3]. Yiyi decoc-
tion (YYD) is a CHF that has been used to manage AP in clinical practice for decades, in which
rhubarb (Rhei Radix Et Rhizoma, RRR) represents a monarch drug and Polygoni Cuspidati
Rhizoma et Radix (PCRR), Piperis Kadsurae Caulis and Hirudo serve as adjuvant drugs to
improve the effects of rhubarb. Metabolome-oriented network pharmacology research has
revealed the potential of resveratrol, emodin, rhein, and their derivatives as efficacious compo-
nents of YYD, most of which are derived from the RRR monarch drug and the PCRR minister
drug [6]. Resveratrol, emodin, and rhein (RER) exhibit multiple beneficial effects, including
antimicrobial, anti-inflammatory, and antioxidant effects [7–9], and the anti-AP effects of these
components have been well-documented [10–16], suggesting that they are candidate markers
for quality control [6]. However, the overall efficacy and underlying mechanism of YYD in AP
remain to be clarified. Additionally, the efficacy and mechanism of the combination of resvera-
trol, emodin, and rhein in a similar ratio found in YYD warrant further verification.
Metabolomics provides a promising analysis methodology to investigate the global metabolic
impact after treatment by CHFs. Metabolomics involves an integrated analysis of metabolites in
aparticular compartment of an organism and focuses on the physio-pathological conditions of
biological systems. Under AP conditions, metabolites involved in different biochemical pathways
become dysregulated. Gas chromatography/liquid chromatography-mass spectrometry (GC/LC–
MS) or nuclear magnetic resonance (NMR) based metabolomics studies have been performed
using biofluids from AP patients or murine models to analyze global metabolic changes and iden-
tify significantly altered metabolites [17–20]. An in-depth understanding of disrupted metabolic
profiles will help revealing the potential biomarkers for pathogenesis and uncovering the underly-
ing mechanism, thus contributing to the discovery of specific drugs for AP treatment.
In the present study, a metabolomics strategy integrating global and unique metabolic char-
acteristics was utilized to explore the underlying mechanism of the effects of YYD and RER on
AP. First, serum metabolomics was performed to identify the metabolic changes in experimen-
tal AP. Second, an in-depth analysis of bile acid metabolism was performed based on an in-
house database. Finally, an integrated analysis of the intervention effects of YYD and RER on
the metabolic perturbations was performed.
Methods
Materials and reagents
Resveratrol, emodin, and rhein were purchased from Chengdu Alfa Biotechnology Co., Ltd,
and the purity of each compound was greater than 98%, as determined by HPLC analysis. MS-
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 2 / 17
Competing interests: The authors have declared
that no competing interests exist.
Abbreviations: AMS, amylase; AP, acute
pancreatitis; BA, bile acid; CA, cholic acid; CHFs,
Chinese herbal formulas; DCA, deoxycholic acid;
DU, dihydroxycholic acid; OPLS-DA, orthogonal
partial least-squares discriminant analysis; PCA,
principal component analysis; PCRR, Polygoni
Cuspidati Rhizoma et Radix; PLS-DA, partial least-
squares discriminant analysis; QC, quality control;
RER, resveratrol-emodin-rhein; RRR, Rhei Radix Et
Rhizoma; T, taurocholic acid sulfate; TCA cycle,
tricarboxylic acid cycle; TCDCA,
taurochenodeoxycholic acid; TDCA,
taurodeoxycholic acid; THDCA,
taurohyodeoxycholic acid; TU, trihydroxycholic
acid; TUDCA, tauroursodeoxycholic acid; UCA,
ursocholic acid; UDCA, ursodeoxycholic acid; YYD,
Yiyi decoction.
grade methanol and acetonitrile were obtained from Merck Company Inc. (Darmstadt, Ger-
many). MS-grade formic acid was obtained from Fisher Scientific Company Inc. (Fairlawn,
NJ, USA). All other reagents were of analytical grade. Fresh ultrapure water (18.2 MO) was
prepared using a Milli-Q water purification system (Millipore, Milford, MA, United States).
TNF-αand IL-6 assay kits were purchased from Nanjing Jiancheng Bioengineering Institute
(Nanjing, China), while amylase (AMS) was purchased from Nanjing Jin Yibai Biological
Technology Co. Ltd. (Nanjing, China). 2-Chloro-l-phenylalanine, which was used as the inter-
nal standard (IS), was purchased from J&K Scientific Ltd. (Beijing, China).
Preparation of YYD
YYD is composed of Rhei Radix et Rhizoma (Lot. 20201203), Polygoni Cuspidati Rhizoma et
Radix (Lot. 20201202), Piperis Kadsurae Caulis (Lot. 20201201), and Hirudo (Lot. 20201201)
at a ratio of 3:4.5:1:5, and these herbs were purchased from Anhui Wansheng Chinese Herbal
Pieces Co., Ltd (Anhui, China). The quality of the above herbs was verified according to the
Chinese Pharmacopeia (China Pharmacopoeia Committee, 2020). Briefly, the four herbs were
soaked in distilled water for 3 h, and then extracted twice with a 12- fold volume of boiling
water for 1 h. The extracted solutions were concentrated under reduced pressure to give a con-
centration of 1.08 g/mL and then stored at 4˚C. UPLC-UV was used to determine the contents of
resveratrol, emodin, and rhein in YYD, which indicated contents of 1.20, 2.01, and 1.14 μg/mL
respectively.
Animal experiment
Twenty-eight male SPF C57BL/6 mice (24 ±1 g) were maintained under controlled tempera-
ture (23 ±2˚C), humidity (55 ±5%) and lighting (12 h light-dark cycles) conditions. After 3
days of acclimatization, the mice were randomly divided into the following four groups: con-
trol group (Control), AP group (Model), YYD group (YYD) and active ingredient-treated
group (RER) (n = 7 in each group). The AP model was induced by intraperitoneal injection of
8% L-arginine (3.5 g/kg, twice, hourly interval), and the control group was injected with the
same volume of sterile normal saline. Then, YYD was intragastrically administered to mice at
a dosage of 19.6 g crude drug/kg/d, and RER was intragastrically administered to mice at a
dosage of 50 mg/kg/d, which is in accordance with the proportions of the original YYD (res-
veratrol at 12.5 mg/kg/d, emodin at 25 mg/kg/d, and rhein at 12.5 mg/kg/d). Drugs were
administered twice a day with an interval of 8 h. The animal protocol was approved by the Ani-
mal Ethics Committee of the Affiliated Hospital of Nanjing University of Chinese Medicine
(2018DW-05-06). All efforts were made to minimize animal suffering and the number of ani-
mals used for the studies.
After 72 h, the mice were anesthetized by intraperitoneal injection of 1% pentobarbital solu-
tion (50 mg/kg), and sacrificed by cervical dislocation under anesthesia. Blood samples were
collected, and centrifuged for 10 min at 3000 rpm. The serum was transferred and then stored
at -80˚C until use. Pancreatic and ileal tissues were collected and fixed in 4% paraformalde-
hyde solution for histopathological staining.
Histopathological examination and biochemical analysis
Tissues were embedded in paraffin, stained with hematoxylin and eosin (H&E), and then
observed under an optical microscope (Olympus, Japan) at a magnification of 200×. The histo-
pathological scores of pancreatic tissue and ileal tissue were evaluated by two blinded patholo-
gists, using the scoring system reported previously [21,22], and the data were presented as the
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 3 / 17
mean ±SD (n = 4). The serum levels of TNF-α, IL-6 and AMS were determined according to
the instructions of the assay kits.
Metabolome analysis
Serum preparation. Serum (200 μL) was fortified with 600 μL of acetonitrile (containing
20 ng/ml IS) and vortexed for 3 min. After centrifugation at 13000 rpm and 4˚C for 10 min,
the supernatant was dried under nitrogen gas. Before analysis, the residues were reconstituted
in 100 μL of acetonitrile/water (1:1, v/v).
Instruments and conditions. Chromatographic separation was performed on an
ExionLC system (AB Sciex, Foster City, CA, USA) equipped with a Waters Acquity BEH C18
column (2.1×100 mm, 1.7 μm) at 35˚C. In brief, 2 μL of the samples was injected, and a flow
rate of 0.3 mL/min was used. Mobile phase A consisted of water with 0.1% formic acid (v/v),
and mobile phase B consisted of acetonitrile. The following gradient was utilized: 0–8 min
from 5% to 60% mobile phase B, 8–18 min from 60% to 97% mobile phase B, 18–21 min at
97% mobile phase B; and back to the initial ratio of 5% mobile phase B, which was maintained
for an additional 4 min for re-equilibration.
A 5600 Q-TOF mass spectrometer (AB Sciex, Foster City, CA, USA) with an electrospray
ionization source (Turbo Ionspray) was used for detection under negative and positive ion
modes. The parameters of the mass spectrometer were as follows: gas1 and gas2, 45 psi; curtain
gas, 35 psi; heat block temperature, 550˚C; ion spray voltage, -4.5 kV in negative mode and 5.5
kV in positive mode; declustering potential, 50 V; collision energy, ±35 V; and collision energy
spread (CES), ±15 V. A pooled sample, which consisted of a mixture of small aliquots of each
sample, was used as the quality control (QC). The QC specimens were analyzed after every five
or six samples throughout the entire analysis procedure.
Data processing and analytical strategy. All of the raw data files were uploaded using the
SCIEX OS software for data extraction. A three-dimensional matrix consisting of sample
names (observations), annotated peak indices (RT-m/z pairs), and peak intensities was
obtained. The peak intensity was calibrated to that of the internal standard. Variants with an
RSD�40% in QCs were excluded. The pretreated data were imported into SIMCA-P (version
14.1, Umetrics, Umea, Sweden) for OPLS-DA and PLS-DA. Before biomarker filtration and
identification, a Pareto-scaled PCA-Class was performed to exclude abnormal samples (out of
the 2nd line) and improve the reliability of the data. The results were considered statistically
significant at P<0.05. Potential markers were annotated by HMDB (http://www.hmdb.ca/)
and LIPID MAPS (https://www.lipidmaps.org/). The results from pathway analysis are pre-
sented graphically by MetaboAnalyst 5.0 (http://www.MetaboAnalyst.ca/). Metabolite set
enrichment analysis (MSEA) and pathway analysis, which are integrated enrichment analyses
and pathway topology analyses, were used to visualize the important metabolic pathways. A
small P value and large pathway impact factor revealed that the pathway was significantly
affected. The correlation and interaction between biomarkers and the network analysis were
established based on the pathway analysis and references.
Results
Pathological parameters
Histopathological examination revealed that compared with the NS-treated mice and drug-
treated mice, the AP model mice had markedly shorter villi and fewer necrotic epithelial cells
in the distal ileum (Fig 1A). The ileal histological scores of Control, Model, YYD and RER
group were 0, 1.2±0.2, 0.3±0.1, and 1.1±0.2, respectively. Moreover, histiocyte edema, pancre-
atic cell necrosis, inflammatory cell infiltration, and pancreatic lobule fuzziness, were obviously
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 4 / 17
observed. Conversely, there was mild edema among tissue cells and small inflammatory cell
infiltration in the YYD and RER groups (Fig 1B). The pancreatic histological scores of Control,
Model, YYD and RER group were 2.2±0.6, 12.5±4.1, 2.5±0.5, and 2.0±0.3, respectively. In addi-
tion, abnormal increases in the serum TNF-α, AMS and IL-6 levels (Fig 1C–1E), which are
essential for the diagnosis of AP, were detected in the AP model mice. The level of the AMS
digestive enzyme was measured to assess the severity of pancreatic injury. A 3-fold increase in
the serum AMS level is among the diagnostic criteria for AP, whereas the TNF-αand IL-6 levels
are consistently associated with increased severity of AP. Treatment with YYD and RER signifi-
cantly reversed the increase in the levels of anti-inflammatory cytokines in the AP model mice.
Data processing
As shown in S1 and S2 Figs (Supplemental information), the overlapping total ion current
(TIC) chromatograms of the QC samples demonstrated that acceptable variations occurred
during large-scale sample analysis, and all of the samples were in the second line in both posi-
tive and negative modes. Therefore, the results were meaningful for further analysis.
Metabolomic analysis of acute pancreatitis
Orthogonal partial least-squares discriminant analysis (OPLS-DA) was utilized to identify dif-
ferent metabolites and screen potential biomarkers, to provide a comprehensive
Fig 1. The effects of YYD and RER on 8% L-arginine induced AP in mice. (A) Representative HE-stained histological
sections of the ilea. (B) Representative HE-stained histological sections of the pancreas. (C) Serum TNF-alpha level; (D)
Serum AMS level; (E) Serum IL-6 level. Data are presented as the mean ±SD (n = 7), *P<0.05 vs. Model, **P<0.01 vs.
Model.
#
P<0.05 vs. Control,
##
P<0.01 vs. Control.
https://doi.org/10.1371/journal.pone.0310689.g001
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 5 / 17
understanding of the AP mechanism. Metabolites with variable importance in the projection
(VIP) value >1 and |p(corr)|>0.3 were further subjected to the Mann-Whitney U test to deter-
mine the significance of each metabolite, and the P<0.05 was considered to indicate statistical
significance. As shown in Fig 2, the control group was clearly distinguished from the model
group in the negative mode (Fig 2Ai) and positive mode (Fig 2Bi). Moreover, the permutation
test confirmed the satisfactory validity of the OPLS-DA models (Fig 2Aii and 2Bii). Volcano
plots (S3 Fig) showing the fold change (FC) and Pvalue were generated using RStudio, and the
metabolites were annotated. The significantly altered compounds included 14 phospholipids,
8 unsaturated fatty acids, 5 lipoyl-carnitines, 4 bile acids, 4 phenols, 3 organic acids and 1
amino acid (Table 1).
Metabolomic analysis of the effects of YYD and RER on acute pancreatitis
Partial least-squares discriminant analysis (PLS-DA) was also performed to observe the dis-
tances among the groups. As shown in Fig 3, the model and control groups were differentiated
in both positive and negative modes. The RER group was closer to the control group than to
the YYD group, which indicated that RER might be more effective at regulating the migratory
metabolome. The YYD points were scattered far from the other groups, which may have been
due to the detection of xenobiotics in the YYD group.
Fig 2. Orthogonal partial least-squares discriminant analysis (OPLS-DA) on the data from ultra-performance
liquid chromatography quadrupole/time-of-flight mass spectrometry (UPLC-Q/TOF MS) profiling data from
control vs model groups. (A) Negative ion mode (i) Score plot, (ii) Permutation plot (R2X 0.346, R2Y 0.9937, Q2
0.334); (B) Positive ion mode (i) Score plot, (ii) Permutation plot (R2X 0.435, R2Y 0.862, Q2 0.364).
https://doi.org/10.1371/journal.pone.0310689.g002
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 6 / 17
Table 1. Identification results of significantly changed metabolites in serum of AP mice.
No. Name Formula Rt ESI- ESI+ VIP p
corr
p
value
Fold
Adduct m/z calm/z ppm Fragments Adduct m/z calm/z ppm Fragments
1 3-Hydroxyvaleric acid C
5
H
10
O
3
2.84 M-H 117.0559 117.0557 1.54 91 - - - - - 4.74 0.68 0.0243 1.44
2 Pyrocatechol sulfate C
6
H
6
O
5
S 2.98 M-H 188.9863 188.9863 -0.11 188, 109, 108,
79
3.49 0.47 0.0243 1.73
3 Phenol sulphate C
6
H
6
O
4
S 3.26 M-H 172.9919 172.9914 2.89 172, 93, 79, 65 - - - - - 16.77 -0.61 0.0193 0.60
4 Phenylglucuronide C
12
H
14
O
7
3.41 M-H 269.0668 269.0667 0.45 175, 113, 93,
85, 59
- - - - - 1.62 -0.60 0.0380 0.56
5 Hippuric acid C
9
H
9
NO
3
3.61 M-H 178.0513 178.0510 1.85 178, 134, 77 M+H 180.0655 180.0655 -0.11 105, 77 2.33 0.58 0.0380 2.06
6 Hydroxyferulic acid C
10
H
10
O
5
4.13 M-H 209.0460 209.0456 2.15 209, 165, 121 M+H 211.0603 211.0601 0.95 149, 121, 65 2.86 0.52 0.0380 1.34
7 Equol 4’-sulfate C
15
H
14
O
6
S 5.39 M-H 321.0441 321.0438 0.84 321, 241, 135,
121, 119, 79
4.41 0.54 0.0193 1.63
8 N-Heptanoylglycine C
9
H
17
NO
3
5.39 M-H 186.1136 186.1136 0.16 74 M+H 188.128 188.1281 -0.53 95, 67 1.03 -0.51 0.0380 0.65
9 3-Oxocholic acid C
24
H
38
O
5
7.90 M
+FA-H
451.2696 451.2701 -1.17 451, 423, 405,
387, 353
M+H 407.2794 407.2792 0.49 407, 389, 371,
353, 335, 159,
145, 135
5.42 -0.54 0.0305 0.48
10 Ursocholic acid C
24
H
40
O
5
8.01 M
+FA-H
453.2849 453.2858 -1.94 407, 345, 343,
289, 233
- - - - - 2.93 -0.58 0.0469 0.43
11 Ursodeoxycholic acid C
24
H
40
O
4
8.26 M
+FA-H
437.2899 437.2909 -2.20 391, 195 - - - - - 1.51 -0.40 0.0380 0.59
12 Dodecanoylcarnitine C
19
H
37
NO
4
8.89 - - - - - M+H 344.2800 344.2795 1.34 344, 285, 183,
144, 85
1.09 -0.56 0.0031 0.58
13 LysoPC(14:0/0:0) C
22
H
46
NO
7
P 9.68 M
+FA-H
512.2989 512.2994 -0.98 512, 452, 227 M+H 468.3082 468.3085 -0.58 468, 450, 285,
184, 104
2.23 -0.71 0.0021 0.78
14 Deoxycholic acid C
24
H
40
O
4
9.77 M
+FA-H
437.2899 437.2909 -2.20 391, 355, 347,
345, 329, 327
- - - - - 1.94 -0.59 0.0305 0.45
15 LysoPC(20:5/0:0) C
28
H
48
NO
7
P 9.81 M
+FA-H
586.3157 586.3150 1.13 586, 526, 301,
257, 224
M+H 542.3240 542.3241 -0.18 542, 524, 258,
184, 104
4.74 -0.65 0.0092 0.70
16 LysoPC(16:1/0:0) C
24
H
48
NO
7
P 10.06 M
+FA-H
538.3133 538.3150 -3.23 538, 478, 253,
224
M+H 494.3242 494.3241 0.16 494, 476, 311,
258, 184
6.38 -0.65 0.0243 0.62
17 Tetradecanoylcarnitine C
21
H
41
NO
4
10.24 - - - - - M+H 372.3105 372.3108 -0.91 372, 313, 211,
144, 85
2.37 -0.71 0.0014 0.62
18 Hydroxyhexadecanoic
acid
C
16
H
32
O
3
10.41 M-H 271.2279 271.2279 0.11 271, 211 - - - - - 1.13 -0.68 0.0071 0.63
19 LysoPE(22:6/0:0) C
27
H
44
NO
7
P 10.53 M-H 524.2778 524.2783 -0.95 524, 327, 283,
229, 214, 196,
140
M+H 526.2928 526.2928 -0.04 526, 508, 385,
354, 311
3.59 0.46 0.0343 1.17
20 LysoPE(20:4/0:0) C
25
H
44
NO
7
P 10.62 M-H 500.2775 500.2783 -1.60 500, 303, 259,
196, 140
M+H 502.2923 502.2928 -1.04 502, 484, 361,
330, 287, 269
5.20 0.84 0.0009 1.47
21 Hexadecenoylcarnitine C
23
H
43
NO
4
10.62 - - - - - M+H 398.3265 398.3265 0.03 398, 339, 237,
144, 85
1.72 -0.63 0.0009 0.41
22 11-HEPE C
20
H
30
O
3
10.76 M-H 317.2121 317.2122 -0.38 317, 299, 255,
179, 135, 107
- - - - - 6.32 -0.75 0.0092 0.42
23 LysoPI(18:2/0:0) C
27
H
49
O
12
P 10.76 M-H 595.2860 595.2889 -4.85 595, 415, 315,
279, 241, 152
- - - - - 1.84 -0.67 0.0092 0.67
(Continued )
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 7 / 17
Table 1. (Continued)
No. Name Formula Rt ESI- ESI+ VIP p
corr
p
value
Fold
Adduct m/z calm/z ppm Fragments Adduct m/z calm/z ppm Fragments
24 LysoPI(20:4/0:0) C
29
H
49
O
12
P 10.80 M-H 619.2872 619.2889 -2.73 619, 439, 315,
303, 241, 152
- - - - - 5.92 -0.67 0.0071 0.73
25 LysoPC(16:0/0:0) C
24
H
50
NO
7
P 11.48 M
+FA-H
540.3302 540.3307 -0.93 540, 480, 255,
242, 224
2M+H 991.6721 991.6723 -0.16 599, 478, 184,
86
8.73 -0.73 0.0044 0.84
26 10-HDoHE C
22
H
32
O
3
11.55 M-H 343.2272 343.2279 -1.95 343, 325, 281,
234, 205, 161,
133, 107
- - - - - 5.99 -0.69 0.0305 0.54
27 LysoPI(20:3/0:0) C
29
H
51
O
12
P 11.64 M-H 621.3053 621.3045 1.22 621, 441, 315,
305, 241, 152
- - - - - 1.04 -0.66 0.0152 0.55
28 11-HETE C
20
H
32
O
3
11.73 M-H 319.2281 319.2279 0.72 319, 301, 257,
179, 163, 135
- - - - - 8.78 -0.67 0.0469 0.66
29 Palmitoylcarnitine C
23
H
45
NO
4
11.87 - - - - - M+H 400.3411 400.3421 -2.60 400, 341, 239,
144, 85
2.87 -0.59 0.0044 0.61
30 LysoPC(18:1/0:0) C
26
H
52
NO
7
P 11.93 M
+FA-H
566.346 566.3463 -0.53 566, 506, 281,
242, 224
M+H 522.3550 522.3554 -0.80 504, 339, 258,
184, 166, 124,
104, 86
20.04 -0.88 0.0205 0.62
31 LysoPE(18:1)/0:0) C
23
H
46
NO
7
P 11.93 M-H 478.293 478.2939 -1.88 478, 281, 196 M+H 480.3088 480.3085 0.69 480, 462, 419,
339, 308
2.61 0.68 0.0021 2.35
32 LysoPG(18:2/0:0) C
24
H
45
O
9
P 11.99 M-H 507.2704 507.2728 -4.81 507, 282, 279,
224, 152
- - - - - 1.24 -0.62 0.0118 0.68
33 Oleoylcarnitine C
25
H
47
NO
4
12.18 - - - - - M+H 426.3574 426.3578 -0.91 426, 367, 144,
85
3.00 -0.52 0.0343 0.68
34 Stearidonic acid C
18
H
28
O
2
13.43 M-H 275.2014 275.2017 -0.91 275, 231 M+H 277.2167 277.2162 1.80 277, 235, 185,
163, 149, 121,
93, 79
1.49 -0.66 0.0193 0.60
35 LysoPC(20:1/0:0) C
28
H
56
NO
7
P 14.06 M
+FA-H
594.3772 594.3776 -0.67 594, 534, 309,
224
M+H 550.3864 550.3867 -0.58 550, 532, 184,
104
2.32 0.46 0.0021 1.51
36 PC 25:1; O C
33
H
64
NO
9
P 14.66 - - - - - M+H 650.4385 650.4392 -1.00 650, 184 1.48 -0.54 0.0266 0.59
37 Neuroprotectin D1/
Resolvin D5
C
22
H
32
O
4
15.19 M-H 359.2222 359.2228 -1.61 359, 341, 327,
315, 297
- - - - - 1.17 0.70 0.0041 1.53
38 Arachidonic acid C
20
H
32
O
2
15.60 M-H 303.2337 303.2330 2.47 303, 259 - - - - - 5.75 0.63 0.0193 1.46
39 Oleic acid C
18
H
34
O
2
17.49 M-H 281.2488 281.2486 0.71 281, 219 - - - - - 9.59 0.60 0.0071 1.53
https://doi.org/10.1371/journal.pone.0310689.t001
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 8 / 17
The intragroup comparisons of individual metabolites are presented in histograms (Fig 4).
Phospholipids, fatty acids (FAs), lipoylcarnitines and bile acids (BAs) were disrupted, with
decreased FA metabolites and lipoylcarnitines after modeling. The changes in the abovemen-
tioned metabolites were reversed after treatment with YYD and RER. Further, RER effectively
reversed more metabolites than YYD.
Bile acid metabolic analysis
BAs are important host-derived and microbially modified signaling molecules that are synthe-
sized in the liver. The concentrations of BAs are important indicators of either the pathological
or physiological status of certain organs, particularly the liver and intestine. Because disorders
of BA homeostasis are implicated in a variety of diseases including pancreatitis, the present
study further analyzed the BAs. A total of 16 BAs were extracted from the serum (Table 2,
S4 Fig), and their intensities were compared via a histogram (Fig 5). Almost all of the BAs
showed a downward trend after modeling, with only five BAs (DCA, UDCA, 3-oxo-CA, UCA,
and DU1) exhibiting statistical significance. Although none of the BAs were significantly
reversed in the dosed group, a tendency toward improvement was observed, especially for
THDCA, T2, T4, and TDCA in the YYD group and for TCDCA, DCA, TU2, TU1, UDCA,
3-oxo-CA, DU2, UCA, and DU1 in the RER group.
Pathway analysis
Pathway analysis revealed that the underlying regulatory effects of YYD and RER on AP in
mice were mainly attributed to lipid metabolism, energy metabolism and amino acid metabo-
lism. The important metabolic pathways included glycerophospholipid metabolism, BA bio-
synthesis, AA metabolism, UFA biosynthesis, FA oxidation, FA degradation, and
phenylalanine metabolism (Fig 6).
Fig 3. Partial least squares discriminate analysis (PLS-DA) analysis and permutation plot between the control,
model, YYD and RER groups. (A) Negative ion mode (i) Score plot, (ii) Permutation plot (R2X 0.634, R2Y 0.829, Q2
0.500); (B) Positive ion mode (i) Score plot, (ii) Permutation plot (R2X 0.603, R2Y 0.887, Q2 0.683).
https://doi.org/10.1371/journal.pone.0310689.g003
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 9 / 17
Discussion
Our previous study revealed the potential efficacious materials and candidate quality markers
for YYD treatment. In this study, we further investigated the effect of YYD and its effective
components, namely, RER, on AP using a metabolomic approach that integrated global and
unique metabolic characteristics.
AP is an inflammatory disease. During AP, a waterfall-like cascade release of proinflamma-
tory cytokines, such as TNF-αand IL-6, leads to pancreatic inflammation and necrosis. The
constantly activated inflammatory response evolves into MODS and systemic inflammatory
response syndrome (SIRS). Both YYD and RER exerted anti-inflammatory effects by signifi-
cantly reversing the abnormal increase in the serum TNF-αand IL-6 levels in AP mice. The
therapeutic effects of these compounds on AP were further confirmed by H&E staining of pan-
creatic and ileal sections from the respective groups. As expected, pancreatic edema,
Fig 4. Intensity comparation of significantly changed metabolites of four groups of control,model, YYD and RER
groups.
#
P<0.05 compared with Control;
##
P<0.01 compared with Control; *P<0.05 compared with Model; **
P<0.01 compared with Model.
https://doi.org/10.1371/journal.pone.0310689.g004
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 10 / 17
Table 2. Identification results of significantly changed bile acids in the serum of AP mice.
Name Formula Rt min Adduct calm/z m/z ppm Fragments pvalue
Control vs Model Model vs YYD Model vs RER
TUDCA C
26
H
45
NO
6
S 5.95 M-H 498.2895 498.2910 3.01 498, 80 0.6058 0.8665 0.6058
THDCA C
26
H
45
NO
6
S 6.37 M-H 498.2895 498.2900 1.00 498, 80 0.743 0.0059 0.2359
TDCA C
26
H
45
NO
6
S 7.22 M-H 498.2895 498.2879 -3.21 498, 80 0.673 0.0289 0.1672
TCDCA C
26
H
45
NO
6
S 7.53 M-H 498.2895 498.2889 -1.20 498, 80 0.673 0.1893 0.1672
T01 C
26
H
45
NO
7
S 5.51 M-H 514.2844 514.2855 2.14 514, 80 1 0.6943 0.3704
T02 C
26
H
45
NO
7
S 5.66 M-H 514.2844 514.2853 1.75 514, 80 0.4234 0.0059 0.3704
T03 C
26
H
45
NO
7
S 6.3 3 M-H 514.2844 514.2835 -1.75 514, 80 0.4807 0.1893 0.4234
T04 C
26
H
45
NO
7
S 6.4 M-H 514.2844 514.2859 2.92 514, 80 0.3213 0.0006 0.0464
UDCA C
24
H
40
O
4
8.26 M+FA-H 437.2909 437.29 -2.06 437, 391 0.036 0.0037 0.9626
DCA C
24
H
40
O
4
9.79 M+FA-H 437.2909 437.2913 0.91 437, 391 0.0464 0.0037 0.4807
TU1 C
24
H
40
O
5
7.05 M+FA-H 453.2858 453.2867 1.99 453, 407 0.1388 0.0003 0.4234
TU2 C
24
H
40
O
5
7.4 M+FA-H 453.2858 453.2867 1.99 453, 407 0.1388 0.0289 0.8884
UCA C
24
H
40
O
5
8.05 M+FA-H 453.2858 453.2875 3.75 453, 407 0.0274 0.2319 0.743
DU1 C
24
H
38
O
5
7.36 M+FA-H 451.2701 451.2684 -3.77 451, 405 0.0464 0.014 0.4807
DU2 C
24
H
38
O
5
7.65 M+FA-H 451.2701 451.2699 -0.44 451, 405 0.3213 0.0037 0.3704
3-oxo-CA C
24
H
38
O
5
7.93 M+FA-H 451.2701 451.2688 -2.88 451, 405 0.036 0.0012 0.8884
https://doi.org/10.1371/journal.pone.0310689.t002
Fig 5. Intensity comparation of significantly changed bile acids of four groups.
#
P<0.05 compared with Control;
##
P<0.01 compared with Control; *P<0.05 compared with Model; ** P<0.01 compared with group Model.
https://doi.org/10.1371/journal.pone.0310689.g005
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 11 / 17
hemorrhage, necrosis, and inflammation were alleviated by treatment with YYD and RER.
Previous studies have confirmed the therapeutic effect of resveratrol, emodin and rhein against
AP due to their anti-inflammatory and antioxidant properties in experimental AP [11,12,16].
The present study demonstrated that the RER combination at levels similar to those found in
YYD provided an effective treatment for AP, partly via the inhibition of proinflammatory
cytokine release.
Accumulating clinical and experimental evidence has shown that homeostatic metabolism
of lipids and fatty acids (FAs) is commonly disrupted during AP [23]. The significantly altered
metabolites in the serum of the L-arginine murine model included elevated levels of unsatu-
rated fatty acids (UFAs) and O-phosphocholine, as well as reduced levels of lysophospholipids,
lipoylcarnitines and bile acids (BAs). Alterations in the levels of phospholipids, which are cru-
cial components of cell membranes, might reflect the decomposition of tissues during AP pro-
gression [24], which is consistent with the histopathological observations. Moreover,
glycerophospholipids have been reported to play important roles in membrane-dependent
processes involved in the TLR response [25]. Thus, the dysregulation of glycerophospholipid
metabolism influences the secretion of cytokines, such as TNF-αand IL-6, thereby promoting
inflammation.
In general, the degradation of phospholipids generates one or two FAs. FAs have an impact
on cell and tissue functions as energy sources and membrane components [19]. Since Because
FAs are reportedly involved in inflammatory pathology and disturbed FA metabolism easily
leads to hyperlipidemia, FAs are inevitably implicated in AP progression [26]. For example,
oleic acid, linoleic acid, and arachidonic acid (AA) induce acinar cell injury at high concentra-
tions, thereby resulting in pancreatitis [27]. Substantially elevated serum FA levels have been
observed in the AP population, and abnormal fluctuations in FA levels (such as those of AA
and linoleic acid) have been reported to be correlated with the deterioration of AP [28]. A
meta-analysis including 20 reports from 11 countries has revealed that UFAs derived from the
lipolysis of unsaturated visceral triglycerides increased systemic injury and organ failure dur-
ing pancreatitis [29]. In the present study, an increase in AA was observed. As the precursor of
Fig 6. Summary of metabolic pathways significantly enriched in YYD/RER treatment.(A) Enrichment overview of
the top 15 metabolic pathways associated with YYD/RER treatment-attributable metabolites. (B) Bubble plot of
metabolic pathways identified by MESA analysis of YYD/RER treatment-attributable metabolites identified metabolic
pathways.
https://doi.org/10.1371/journal.pone.0310689.g006
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 12 / 17
prostaglandins, an increase in AA might induce inflammation [30,31]. Moreover, changes in
lysophospholipid and UFA levels were reversed after administration of YYD and RER, indicat-
ing that YYD and RER inhibit the inflammatory response to alleviate AP symptoms.
Carnitine plays an important role in FA β-oxidation (FAO), which transfers long-chain
fatty acids (LCFAs) across the mitochondrial membrane in the form of lipoylcarnitine.
Decreased lipoylcarnitine levels usually indicate impaired FAO, which can result in lipid accu-
mulation, lipid peroxidation, oxidative stress, irreversible mitochondrial damage, and even
immune dysfunction [32,33]. Moreover, lipid accumulation induced ROS promote the secre-
tion of cytokines via NF-κB. In the present study, lipoylcarnitines were expressed at lower lev-
els in the AP group, but both YYD and RER treatment reversed these changes, especially for
oleoylcarnitine and palmitoylcarnitine. These results suggested that YYD and RER may
improve AP through FA oxidation regulation.
BAs are involved in the pathogenesis of pancreatitis due to their role in the induction of
cytosolic calcium overload and subsequent cell necrosis [34–36]. Disordered BA metabo-
lism may cause cholestasis or lead to the reflux of BAs into the pancreas, which is an inducer
of AP [37,38]. In the murine model, almost 16 detected BAs showed a decreasing trend,
and an increasing trend in these BAs was observed after treatment with YYD and RER.
Among them, UDCA and TCDCA have protective effects in acute biliary pancreatitis, and
UDCA has been approved for use in gallstone dissolution and in treating primary biliary
cholangitis [39,40].
BAs undergo enterohepatic circulation [41]. The liver and pancreas communicate with the
intestine through bile ducts and pancreatic ducts. BA metabolism is an important part of lipid
metabolism. BAs can form chylomicrons (CMs) together with phospholipids in the intestine,
which is an important metabolic pathway for cholesterol and triglycerides. Disordered BA
metabolism always implies liver damage, and dyshomeostasis of the gut-liver-pancreas axis
may influence host-microbe interactions [42]. The decreased villus length and necrotic epithe-
lial cells observed in histologic sections of the distal ileum implied a damaged intestinal barrier.
Subsequently, toxins, such as lipopolysaccharides (LPSs), produced by microbes can penetrate
into systemic circulation more easily. LPS activates Toll-like receptor 4 (TLR4), a key trans-
membrane recognition receptor, to release IL-6, TNF-αand other proinflammatory factors,
thus promoting inflammation [43]. The 16 BAs detected in the present study included both
primary and microbially modified secondary BAs, implying a defective intestinal barrier.
Rhein, emodin, and resveratrol have been reported to ameliorate SAP-induced intestinal bar-
rier injury and ameliorate AP-associated liver damage [11–16]. In YYD, rhein, emodin, resver-
atrol and their derivatives have been found to be widely distributed in the bile, intestine and
liver. In addition, microbe-derived aromatic substances, such as hippuric aid, equol 4’-sulfate,
and phenol sulfate, showed obvious fluctuations in the model group compared to the control
group, further confirming the disruption of intestinal homeostasis in AP [44]. After treatment,
the aberrant shift was recovered, especially after RER treatment. Taken together, these results
indicated that YYD and RER might achieve anti-AP effects by regulating disordered lipid and
BA metabolism (Fig 7). However, further experiments are essential to reveal the underlying
molecular mechanism involved.
Conclusion
The present study utilized strategy that integrates global and unique metabolic characteristics
to reveal the efficacy and mechanism of YYD and RER on murine AP. Because RER can be
easily extracted, prepared, and subjected to strict quality control, this combination holds
potential for further development as a lead drug in the treatment of AP.
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 13 / 17
Supporting information
S1 Fig. The overlapping total ion chromatograms (TICs) of QC samples in positive and
negative modes.
(DOCX)
S2 Fig. PCA-Class score plots in positive and negative modes.
(DOCX)
S3 Fig. Volcano plots of control vs model.
(DOCX)
S4 Fig. Structures of 16 bile acids extracted in the serum of AP mice.
(DOCX)
Author Contributions
Conceptualization: Guanwen Gong, Yongping Wu, Yuan Cao.
Data curation: Guanwen Gong, Yongping Wu.
Funding acquisition: Guanwen Gong.
Investigation: Guanwen Gong, Yongping Wu.
Methodology: Guanwen Gong, Yuan Cao.
Project administration: Yuan Cao.
Resources: Yuan Cao.
Fig 7. Altered metabolites and pathways implicated in AP that could be ameliorated by both treatment of YYD
and RER in murine plasma. Metabolites colored in green were decreased in AP model while red were increased.
Reversed metabolites were illustrated with histogram. LCFAO: long chain fatty acid oxidation; CPT1: carnitine-
palmitoyltransferase-1; CPT1: carnitine-palmitoyltransferase-2; TCA cycle: tricarboxylic acid cycle; COX:
cyclooxyganese; PGG2: Prostaglandin G2; PGH2: Prostaglandin H2; PGs: Prostaglandins; CM: chylomicron; LPS:
lipopolysaccharide; NF-κB: nuclear factor kappa-B; TLR4: Toll-like receptor 4.
https://doi.org/10.1371/journal.pone.0310689.g007
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 14 / 17
Supervision: Yuan Cao.
Validation: Yanwen Jiang.
Writing – original draft: Guanwen Gong, Yongping Wu.
Writing – review & editing: Yuan Cao.
References
1. Boxhoorn L, Voermans RP, Bouwense SA, Bruno MJ, Verdonk RC, Boermeester MA, et al. Acute pan-
creatitis. Lancet. 2020; 396:726–734. https://doi.org/10.1016/S0140-6736(20)31310-6 PMID:
32891214
2. Lee PJ, Papachristou GI. New insights into acute pancreatitis. Nat Rev Gastroenterol Hepatol. 2019;
16:479–496. https://doi.org/10.1038/s41575-019-0158-2 PMID: 31138897
3. Chinese Pancreatic Surgery Association, Chinese Society of Surgery, Chinese Medical Association.
Guidelines for diagnosis and treatment of acute pancreatitis in China (2021). ChinJ Dig Surg. 2021;
20:730–739.
4. Garg PK, Singh VP. Organ failure due to systemic injury in acute pancreatitis. Gastroenterology. 2019;
156:2008–2023. https://doi.org/10.1053/j.gastro.2018.12.041 PMID: 30768987
5. Whitcomb DC. Clinical practice. Acute pancreatitis. N Engl J Med. 2006; 354:2142–2150. https://doi.
org/10.1056/NEJMcp054958 PMID: 16707751
6. Gong GW, Tang WH, Zhou Z, Jiang YW, Wang CZ, Cheng H, et al. Potential efficacious materials
investigation of Yi-Yi Mixture based on metabolome-oriented network pharmacology strategy. J Chro-
matogr B. 2022; 1197:123199. https://doi.org/10.1016/j.jchromb.2022.123199 PMID: 35305386
7. Bhat KPL, Kosmeder JW II, Pezzuto JM. Biological effects of resveratrol. Antioxid Redox Sign. 2001;
3:1041–1064.
8. Yin C, Han X, Lu Q, Qi X, Guo C, Wu X. Rhein incorporated silk fibroin hydrogels with antibacterial and
anti-inflammatory efficacy to promote healing of bacteria-infected burn wounds. Int J Biol Macromol.
2022; 201:14–19. https://doi.org/10.1016/j.ijbiomac.2021.12.156 PMID: 34995653
9. Monisha BA, Kumar N, Tiku AB. Emodin and its role in chronic diseases. Adv Exp Med Biol. 2016;
928:47–73. https://doi.org/10.1007/978-3-319-41334-1_3 PMID: 27671812
10. Xiang H, Zhang Q, Qi B, Tao X, Xia S, Song H, et al. Chinese herbal medicines attenuate acute pancre-
atitis: pharmacological activities and mechanisms. Front Pharmacol. 2017; 8:216. https://doi.org/10.
3389/fphar.2017.00216 PMID: 28487653
11. Jha RK, Ma Q, Lei Z, Sha H. Resveratrol ameliorates the deleterious effect of severe acute pancreatitis.
Cell Biochem Biophys. 2012; 62:397–402. https://doi.org/10.1007/s12013-011-9313-2 PMID:
22042732
12. Ning JW, Zhang Y, Yu MS, Gu ML, Xu J, Usman A, et al. Emodin alleviates intestinal mucosal injury in
rats with severe acute pancreatitis via the caspase-1 inhibition. Hepatobiliary Pancreat Dis Int. 2017;
16:431–436. https://doi.org/10.1016/S1499-3872(17)60041-9 PMID: 28823375
13. Zhuang S, Zhong J, Zhou Q, Zhong Y, Liu P, Liu Z. Rhein protects against barrier disruption and inhibits
inflammation in intestinal epithelial cells. Int Immunopharmacol. 2019; 71: 321–327. https://doi.org/10.
1016/j.intimp.2019.03.030 PMID: 30952096
14. Sha H, Ma Q, Jha RK, Xu F, Wang L, Wang Z, et al. Resveratrol ameliorates hepatic injury via the mito-
chondrial pathway in rats with severe acute pancreatitis. Eur J Pharmacol. 2008; 601:136–142. https://
doi.org/10.1016/j.ejphar.2008.10.017 PMID: 18977215
15. Wang G, Sun B, Gao Y, Meng QH, Jiang HC. The effect of emodin-assisted early enteral nutrition on
severe acute pancreatitis and secondary hepatic injury. Mediators Inflamm. 2007; 2007:29638. https://
doi.org/10.1155/2007/29638 PMID: 18288270
16. Yang X, Geng H, You L, Yuan L, Meng J, Ma Y, et al. Rhein protects against severe acute pancreatitis
in vitro and in vivo by regulating the JAK2/STAT3 pathway. Front Pharmacol. 2022; 13:778221.
17. Sanchez-Lopez E, Zhong Z, Stubelius A, Sweeney SR, Booshehri LM, Antonucci L, et al. Choline
uptake and metabolism modulate macrophage IL-1βand IL-18 production. Cell Metab. 2019; 29:1350–
1362.
18. Guo J, Li X, Wang D, Guo Y, Cao T. Exploring metabolic biomarkers and regulation pathways of acute
pancreatitis using ultra-performance liquid chromatography combined with a mass spectrometry-based
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 15 / 17
metabolomics strategy. RSC Adv. 2019; 9:12162–12173. https://doi.org/10.1039/c9ra02186h PMID:
35517037
19. Huang JH, He D, Chen L, Dong CY, Zhang SH, Qin YH, et al. GC-MS based metabolomics strategy to
distinguish three types of acute pancreatitis. Pancreatol. 2019; 19:630–637. https://doi.org/10.1016/j.
pan.2019.05.456 PMID: 31262499
20. Takis P, Taddei A, Pini R, Grifoni S, Tarantini F, Bechi P, et al. Fingerprinting acute digestive diseases
by untargeted NMR based metabolomics. Int J Mol Sci. 2018; 19:3288. https://doi.org/10.3390/
ijms19113288 PMID: 30360494
21. Schmidt M, Mandavilli U, Lewandrowski K, Rattner D, Compton C, Warshaw GA, et al. A better model
of acute pancreatitis for ev aluating therapy. Ann Surg. 1992; 215:44–56. https://doi.org/10.1097/
00000658-199201000-00007 PMID: 1731649
22. Chen YF, Sun SS, Zhang Y, Zhou CH, Zou DW. Changes of intestinal barrier function in mice with
acute necrotizing pancreatitis induced by sodium taurocholate. Chin J Pancreatol. 2023; 23:431–436.
23. Peng Y, Hong J, Raftery D, Xia Q, Du D. Metabolomic-based clinical studies and murine models for
acute pancreatitis disease: A review. BBA–Mol Basis Dis. 2021; 1867:166123. https://doi.org/10.1016/
j.bbadis.2021.166123 PMID: 33713791
24. Yang Q, Sun J, Chen YQ. Multi-dimensional, comprehensive sample extraction combined with LC-GC/
MS analysis for complex biological samples: application in the metabolomics study of acute pancreati-
tis. RSC Adv. 2016; 6:25837–25849.
25. Ko
¨berlin MS, Heinz LX, Superti-Furga G. Functional crosstalk between membrane lipids and TLR biol-
ogy. Curr Opin Cell Biol. 2016; 39:28–36. https://doi.org/10.1016/j.ceb.2016.01.010 PMID: 26895312
26. Wong SW, Kwon MJ, Choi AM, Kim HP, Nakahira K, Hwang DH. Fatty acids modulate Toll-like receptor
4 activation through regulation of receptor dimerization and recruitment into lipid rafts in a reactive oxy-
gen species-dependent manner. J Biol Chem. 2009; 284:27384–27392. https://doi.org/10.1074/jbc.
M109.044065 PMID: 19648648
27. Chang YT, Chang MC, Tung CC, Wei SC, Wong JM. Distinctive roles of unsaturated and saturated
fatty acids in hyperlipidemic pancreatitis. World J Gastroenterol. 2015; 21: 9534–9543. https://doi.org/
10.3748/wjg.v21.i32.9534 PMID: 26327761
28. Sztefko K, Panek J. Serum free fatty acid concentration in patients with acute pancreatitis. Pancreatol-
ogy. 2001; 1:230–236. https://doi.org/10.1159/000055816 PMID: 12120200
29. Khatua B, El-Kurdi B, Patel K, Rood C, Noel P, Crowell M, et al. Adipose saturation reduces lipotzxc-
voxic systemic inflammation and explains the obesity paradox. Sci Adv. 2021; 7:eabd6449.
30. Wu Y, Li K, Zeng M, Qiao B, Zhou B. Serum metabolomics analysis of the anti-inflammatory effects of
gallic acid on rats with acute inflammation. Front Pharmacol. 2022; 13:830439. https://doi.org/10.3389/
fphar.2022.830439 PMID: 35392557
31. Harris RC, Zhang MZ. Cyclooxygenase metabolites in the Kidney. Compr Physiol. 2011; 1:1729–1758.
https://doi.org/10.1002/cphy.c100077 PMID: 23733687
32. Sun N, Shen C, Zhang L, Wu X, Gao Y. Hepatic Kru¨ppel-like factor 16 (KLF16) targets PPARαto
improve steatohepatitis and insulin resistance. Gut. 2020; 70:2183–2195.
33. Xu S, Chaudhary O, Rodrı
´guez-Morales P, Sun X, Chen D, Zappasodi R, et al. Uptake of oxidized lipids
by the scavenger receptor CD36 promotes lipid peroxidation and dysfunction in CD8+ T cells in tumors.
Immunity. 2021; 54:1561–1577.
34. Tran QT, Tran VH, Sendler M, Doller J, Wiese M, Bolsmann R, et al. Role of bile acids and bile salts in
acute pancreatitis: from the experimental to clinical studies. Pancreas. 2021; 50:3–11.https://doi.org/
10.1097/MPA.0000000000001706 PMID: 33370017
35. Muili KA, Wang D, Orabi AI, Sarwar S, Luo Y, Javed TA, et al. Bile acids induce pancreatic acinar cell
injury and pancreatitis by activating calcineurin. J Biol Chem. 2013; 288:570–580. https://doi.org/10.
1074/jbc.M112.428896 PMID: 23148215
36. Ferdek PE, Jakubowska MA, Gerasimenko JV, Gerasimenko OV, Petersen OH. Bile acids induce
necrosis in pancreatic stellate cells dependent on calcium entry and sodium-driven bile uptake. J Phy-
siol. 2016; 594:6147–6164. https://doi.org/10.1113/JP272774 PMID: 27406326
37. Santos JS, Kemp R, Ardengh JC, Jr JE. Conservative management of cholestasis with and without
fever in acute biliary pancreatitis. World J Gastrointest Surg. 2012; 4:55–61. https://doi.org/10.4240/
wjgs.v4.i3.55 PMID: 22530079
38. Pallagi P, Go¨ro¨g M, Papp N, Mada
´csy T, Varga A
´, Crul T, et al. Bile acid- and ethanol-mediated activa-
tion of Orai1 damages pancreatic ductal secretion in acute pancreatitis. J Physiol. 2022; 600:1631–
1650. https://doi.org/10.1113/JP282203 PMID: 35081662
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 16 / 17
39. Ward JBJ, Lajczak NK, Kelly OB, O’Dwyer AM, Giddam AK, Gabhann JN, et al. Ursodeoxycholic acid
and lithocholic acid exert anti-inflammatory actions in the colon. Am J Physiol—Gastr L. 2017; 312:
G550–558. https://doi.org/10.1152/ajpgi.00256.2016 PMID: 28360029
40. Goossens JF, Bailly C. Ursodeoxycholic acid and cancer: From chemoprevention to chemotherapy.
Pharmacol Ther. 2019; 203:107396. https://doi.org/10.1016/j.pharmthera.2019.107396 PMID:
31356908
41. Guzior DV, Quinn RA. Review: microbial transformations of human bile acids. Microbiome. 2021;
9:140. https://doi.org/10.1186/s40168-021-01101-1 PMID: 34127070
42. Silveira MAD, Bilodeau S, Greten TF, Wang XW, Trinchieri G. The gut-liver axis: host microbiota inter-
actions shape hepatocarcinogenesis. Trends Cancer. 2022; 8:583–597. https://doi.org/10.1016/j.
trecan.2022.02.009 PMID: 35331674
43. Huang SQ, Wen Y, Sun HY, Deng J, Zhang YL, Huang QL, et al. Abdominal paracentesis drainage
attenuates intestinal inflammation in rats with severe acute pancreatitis by inhibiting the HMGB1-medi-
ated TLR4 signaling pathway. World J Gastroenterol. 2021; 27:815–834. https://doi.org/10.3748/wjg.
v27.i9.815 PMID: 33727772
44. Zheng X, Xie G, Zhao A, Zhao L, Yao C, Chiu NH, et al. The footprints of gut microbial-mammalian co-
metabolism. J Proteome Res. 2011; 10:5512–5522. https://doi.org/10.1021/pr2007945 PMID:
21970572
PLOS ONE
Integrated metabolic characteristics to reveal the intervention effect of Yiyi decoction
PLOS ONE | https://doi.org/10.1371/journal.pone.0310689 November 21, 2024 17 / 17
