Moutan Cortex: A Review of Origins, Phytochemical Characterization Strategies, and Anti-fibrosis-related Pharmacological Mechanisms and Applications

Abstract

Traditional medicine has long acknowledged the therapeutic effects of Moutan cortex (MC), derived from the dried root bark of the tree peony. In recent times, scientific investigations have shed light on its bioactive components and the mechanisms underlying its health-promoting effects. Here, we review the origin of MC, encompassing its worldwide resource distribution, plant morphological characteristics, and medicinal values. Additionally, a multi-dimensional analysis is carried out on the present research strategies concerning the components of MC, aiming to provide insights into the identification of the active components in MC. Simultaneously, this article focuses on the anti-fibrotic pharmacological mechanisms of the two crucial active components, paeonol and paeoniflorin, derived from MC. We comprehensively summarize the multiple mechanisms and pathways through which these components exhibit anti-fibrotic actions within specific pathological sites. Moreover, it reviews the advancements in patents and clinical research associated with paeonol and paeoniflorin, emphasizing their substantial potential for translational applications. Elucidating the key active components derived from MC and their pharmacological mechanisms holds critical scientific and practical value across multiple fields.

Keywords

Moutan cortex - Paeoniaceae - Paeonia x suffruticosa Andrews - paeonol - paeoniflorin - anti-fibrotic - application

Abbreviations

α-SMA: Alpha-Smooth Muscle Actin

αvβ3: Integrin αVβ3

γ-GT: Gamma-Glutamyl Transferase

ANP: Atrial Natriuretic Peptide

Ang II: Angiotensin II

ARE: Antioxidant Response Element

ASC: Apoptosis-Associated Speck-Like Protein Containing a CARD

Bad: BCL2-Associated Agonist of Cell Death

Bax: BCL2-Associated X Protein

Bel-2: B-cell Lymphoma 2

BDNF: Brain-Derived Neurotrophic Factor

BMP: Bone Morphogenetic Protein

BNP: Brain Natriuretic Peptide

CD34: Cluster of Differentiation 34

COL1: Collagen Type I

CTGF: Connective Tissue Growth Factor

cTnI: Cardiac Troponin I

DBIL: Direct Bilirubin

DLAT: Dihydrolipoamide S-Acetyltransferase

FDX1: Ferredoxin 1

FN: Fibronectin

FSP-1: Fibroblast-Specific Protein-1

HA: Hyaluronic Acid

HMBG1: High Mobility Group Box 1

ICAM-1: Intercellular Adhesion Molecule 1

IFN-γ: Interferon-gamma

iNOS: Inducible Nitric Oxide Synthase

Jagged1: Jagged Canonical Notch Ligand 1

JAK1: Jagged Canonical Notch Ligand 1

MCP-1: Monocyte Chemoattractant Protein-1

MEK: Mitogen-Activated Protein Kinase Kinase

Ngfr: Nerve Growth Factor Receptor

NOX1: NADPH Oxidase 1

Notch1: Notch Receptor 1

PAI-1: Plasminogen Activator Inhibitor-1

PARP-1: Poly [ADP-ribose] Polymerase-1

PCIII: Procollagen Type III

PCNA: Proliferating Cell Nuclear Antigen

PDGF: Platelet-Derived Growth Factor

p-AKT: Phosphorylated Protein Kinase B

p-ERK1/2: Phosphorylated Extracellular Signal-Regulated Kinase 1/2

pIκBα: Phosphorylated Inhibitor of Kappa B Alpha

p-JNK: Phosphorylated c-Jun N-terminal Kinase

p-p38: Phosphorylated p38 Mitogen-Activated Protein Kinase

p-Smad2: Phosphorylated Mothers Against Decapentaplegic Homolog 2

PPARγ: Peroxisome Proliferator-Activated Receptor Gamma

P21: Cyclin-Dependent Kinase Inhibitor 1

RAGE: Receptor for Advanced Glycation Endproducts

SEM: Scanning Electron Microscope

Smad2/3: drosophila mothers against decapentaplegic protein 2/3

STAT3: Signal Transducer and Activator of Transcription 3

TCM: Traditional Chinese medicine

TBA: Total Bile Acids

TEM: Transmission Electron Microscope

TGF-β: Transforming Growth Factor-beta

TLR4: Toll-Like Receptor 4

VEGF: Vascular Endothelial Growth Factor
 

Introduction

Paeonia x suffruticosa Andrews, commonly known as the tree peony, is renowned as the ‘King of Flowers’ and has an abundant historical legacy in China [1]. It was gradually introduced to foreign countries through trade or cultural exchanges [2]. It is highly valued not only for its striking aesthetic beauty but also for its medicinal properties, with its root bark, known as Moutan cortex (MC), serving as a vital herbal medicine in TCM for centuries. Throughout history, in traditional Chinese medical classics such as the ‘Shennongʼs Herbal Classic’ and the ‘Compendium of Materia Medicaʼ, MC has been documented for its therapeutic effects, particularly for its abilities to ‘eliminate heatʼ, ‘cool the bloodʼ, ‘promote blood circulationʼ, and ‘relieve blood stasisʼ. It is typically applied either alone or as an important part in numerous prescriptions to treat a variety of diseases, including febrile diseases, blood stasis-related disorders like amenorrhea, and traumatic injuries [3], [4], [5].

From a modern pharmacological perspective, MC has attracted increasing attention due to its abundant bioactive components. These components, including paeonol, paeoniflorin, and other phenolic compounds, have demonstrated a broad spectrum of biological activities. For instance, they exhibit anti-inflammatory properties by regulating the expression of inflammatory cytokines and anti-oxidant effects by scavenging free radicals, which contribute to potential health benefits in preventing and treating fibrosis-related diseases [6], [7]. Moreover, with the growing global interest in natural products and traditional medicine, the study of MC has expanded beyond traditional medicine research. It has become a subject of investigation in multiple disciplines, such as phytochemistry, pharmacology, and drug development [8].

Currently, while some studies have explored the pharmacological activities of MC, research targeting specific pathological locations and elucidating clear mechanisms remains fragmented. It is essential to propose suitable strategies for the chemical analysis of MC. Moreover, gaining insights into the current application status and development directions related to MC can provide valuable references for further research and rational utilization.

This review focuses on the strategies for the analysis of chemical components in MC, the pharmacological mechanisms of its active components against fibrosis within specific pathological sites, and the progress of their applications in patents and clinical research. By integrating the latest research findings, we hope to provide a more in-depth understanding of MC and offer insights for further research and development in the field.

Search Strategy

To identify relevant articles on pharmacological mechanisms, we systematically searched the following electronic databases from January to February 2025: PubMed, SpringerLink, Web of Science, ScienceDirect, and China National Knowledge Infrastructure. A comprehensive review of international literature was conducted using critical and representative search terms such as ‘Moutan Cortexʼ, ‘paeonolʼ, ‘paeoniflorinʼ, ‘fibrosisʼ, ‘organ fibrosisʼ, ‘cardiovascular fibrosisʼ, ‘pulmonary fibrosisʼ, ‘hepatic fibrosisʼ, ‘renal fibrosisʼ, etc. For application-related information, the keywords ‘paeonol’ and ‘paeoniflorin’ were used to search the Patentstar database [9] and Pharmacodia database [10], respectively, to retrieve relevant patent and clinical study data.

Origins

The tree peony, which is native to China, has a cultivation history spanning more than 1400 years [1]. Based on the data from the GBIF database [11], the distribution of the tree peony ([Fig. 1 a]) is not only concentrated in China but also extends to widespread cultivation around the globe, especially in Europe, North America, Korea, and Japan.

Fig. 1 Resource distribution and morphology of the tree peony and MC. a Global resource distribution of Paeonia x suffruticosa Andrews (image data sourced from the GBIF database); b Production regions of MC in China (the plant morphology and medicinal characteristics of the MC were sourced from [8]).

The tree peony is a woody shrub belonging to the Paeonia subgenus Moutan in the Paeoniaceae family, which typically attains a growth height of approximately 2 meters. It is characterized by its woody, multi-stemmed structure, which distinguishes it from herbaceous peonies that die back each year. The bark of mature stems is gray-brown to purplish-brown, and the root cortex is thick, often used in traditional medicine. The leaves are large, dark green, with ovate to lanceolate leaflets that are often deeply lobed or dissected. The flowers are solitary and terminal and range in color from white and pink to purple and red, often with prominent yellow stamens. The calyx consists of five persistent sepals, and the corolla may have five petals or be double-flowered. The fruit consists of follicles that are ovoid and densely covered with yellowish-brown hairs, dehiscing along the ventral suture when mature [12], [13].

The root system of the tree peony is fibrous and deep. The medicinal properties of the species are found primarily in the root bark [14]. The dried root bark of the tree peony is known as MC, which is widely referred to by different names in various regions. In Chinese, it is called ‘Mudanpi’, in Japan it is known as ‘Botanpi’, and in Korea, it is referred to as ‘Mok-Dan-Pi’ [15]. Despite slight variations in its naming, the herb is most commonly recognized internationally by its Chinese name, ‘Mudanpi’. In appearance, MC is typically brown or reddish-brown on the outside, with a pale, light-colored inner surface. It is thin, slightly flexible, and may have a rough texture. MC is often in strips or fragmented. The resources of MC ([Fig. 1 b]) are widely distributed across China, with the largest production in east and southwest China, particularly in Anhui, Sichuan, Chongqing, Shandong, and other production regions. Among these, Fenghuang Mountain in Tongling, Anhui, is renowned for producing the highest quality MC, often referred to as ‘genuine medicinal herbs’ [16].

MC has a medicinal history spanning thousands of years in China. It was first recorded in ‘Shennongʼs Herbal Classic’ [17] and has been included in the Pharmacopoeia of the Peopleʼs Republic of China [18]. In addition, there are subtle differences in some aspects of the use of MC in different national pharmacopoeia, such as the Japanese Pharmacopoeia, Korean Pharmacopoeia, and Vietnamese Pharmacopoeia [8].

In TCM, MC is historically described as having functions such as to ‘cool the blood’, ‘invigorate circulation’, and ‘clear heat’. It has been conventionally applied in contexts associated with cardiovascular discomfort, extravasated blood, blood stagnation, and certain gynecological conditions [18]. Modern pharmacological studies have demonstrated that both crude extracts derived from MC and isolated compounds exhibit a broad spectrum of bioactivities [19]. MC is gaining popularity in numerous countries for its anti-inflammatory, anti-oxidant, and blood-circulation-promoting properties [3], [4], [20]. As an important botanical in the integrative medicine movement, MC is gaining increased attention in the scientific literature and commercial supplement formulations, underscoring its considerable potential for research and development in both pharmacological applications and natural product innovation [21].

Phytochemical Characterization Strategies

To date, 164 compounds have been isolated and charactered from MC, which can be classified into seven major classes: terpenoids (including monoterpenes, monoterpenes glycosides, and triterpenes), phenols, flavonoids, volatile oils, alkaloids, tannins, and others [15], [22]. In addition, monoterpene glycosides and phenols are predominant components in MC. The pharmacological mechanisms and applications of monomeric compounds from MC, such as paeonol, paeoniflorin, paeonolysaccharide, and gallic acid, have become key areas of research, particularly in the fields of anti-fibrosis, anti-inflammation, and anti-oxidation [8].

In this section, we provide an updated review of research from various technical and analytical perspectives to offer insights and guidance for identifying key bioactive components in MC.

Comprehensive analytical methods

In contrast to traditional and single identification methods such as HPLC, GC, and NMR, the comprehensive application of multi-dimensional analytical techniques can enhance sensitivity and accuracy, offering a more systematic and accurate way to characterize the chemical components of phytomedicine.

UPLC-MS offers significant advantages for low-abundance compounds, including high sensitivity, rapid analysis, and the ability to resolve complex mixtures. Furthermore, a time segment scanning-based quasi-multiple reaction monitoring mode was proposed as an improvement over the traditional full-mass scan mode. This new approach demonstrated superior specificity, sensitivity, and linearity, significantly enhancing the quantitative performance of MC [23].

GC-MS is one of the most important and widely used techniques for analyzing volatile compounds. It enhances the reliability of compound identification by leveraging analytical parameters. This method was applied to analyze the content differences of the chemical components in MC ethanol extracts under ultrasonic processing conditions [24].

Spectroscopic and mass spectrometric techniques offer complementary advantages for compound identification, enhancing accuracy and reliability through multi-dimensional analysis. A new isoflavone, 6′-dihydroxy-2-methyl-2′-methoxy-3′-acetylisoflavone, was isolated from the MC, and its structure was determined using UV, IR, NMR, and high-resolution electrospray ionization mass spectrometry [22].

TEM and SEM techniques provide high-resolution imaging and detailed structural analysis, offering a comprehensive understanding of both physical morphology and chemical composition. A previous study identified a new polysaccharide from MC using GC-MS and NMR, revealing a flaky structure under SEM and an irregular spherical structure under TEM [25].

Additionally, by integrating network pharmacology, a comprehensive understanding of the active components and pharmacological activities of MC can be obtained [26], [27].

Targeted screening

The strategy of screening bioactive compounds from phytomedicinal extracts based on disease targets or biomarkers allows for more precise identification of therapeutic agents. This approach enhances the accuracy of distinguishing effective compounds, ensuring more reliable and targeted therapeutic interventions. It also aids in optimizing the development of plant-based medicines with specific pharmacological effects.

The researchers assessed the anticoagulant activity of MC and captured its bioactive compounds using platelet-immobilized chromatography. Through HPLC-DAD and LC-ESI-MS/MS analysis, they identified four active compounds: oxypaeoniflorin, tetragalloylglucose, pentagalloylglucose, and benzoylpaeoniflorin. These compounds were shown to upregulate the expression of hsp-70 and coronin-1B, while reducing platelet adhesion. Therefore, it is possible to discover new lead compounds to treat cardiovascular diseases [28].

Few systematic studies have explored the integral calcium antagonistic components of MC for treating hypertension. In this study, a dual-luciferase reporter assay combined with UHPLC-QTOF-MS was used to separate, screen, and identify potential calcium channel blockers from MC extracts. As a result, three monoterpenoids (paeoniflorin, benzoylpaeoniflorin, and mudanpioside C), one phenolic acid (paeonol), and one gallotannin (1,2,3,4,6-O-pentagalloylglucose) were identified as potential calcium antagonists in MC [29].

An innovative strategy combining solid-phase extraction with an enzyme activity switch and MS analysis was proposed for the screening of NA (neuraminidase) inhibitors from MC. Using this approach, two benzoic acid derivatives, paeoniflorin and oxypaeoniflorin, were successfully identified as a new class of natural NA inhibitors [30].

COX-2 (cyclooxygenase-2) plays a pivotal role in inflammation. In this study, bio-affinity ultrafiltration coupled with UPLC-MS was employed to screen and identify potential COX-2 inhibitors within the MC extract. A total of 11 potential COX-2 inhibitors, including gallic acid, suffruticoside A/B/C/D, and 6′-O-Galloyl paeoniflorin, were eventually identified from the MC extract [31].

Quality evaluation

The medicinal components of phytomedicine are significantly influenced by factors such as production region, collection period, plant part used, and processing methods. These variations can affect both the potency and consistency of the therapeutic effects, highlighting the need for standardized quality control in herbal medicine.

Studies have shown that the bioactive compounds in MC vary depending on the collection period and production region. Additionally, by analyzing the relationship between these bioactive components and their anti-oxidant and anti-aging activities, six key compounds–gallic acid, p-hydroxybenzoic acid, quercetin-3-O-glucoside, mudanpioside C, benzoyloxypaeoniflorin, and benzoyl paeoniflorin–were identified as pharmacodynamic markers that merit significant attention [32].

The chemical components contained in MC exhibit unique distribution and accumulation in different parts. Through metabolomic analysis, Xiao et al. characterized the metabolomic variations in different root parts of Paeonia x suffruticosa Andrews and suggested that the quality of axial roots was better than that of lateral roots [33]. Additionally, Li et al. systematically characterized the spatial distribution of major metabolites in the root of Paeonia x suffruticosa Andrews using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) [34]. Cao et al. conducted a laser microdissection combined with the UPLC-MS technique to separate and identify the cork layer, cortex, phloem, and xylem of Paeonia x suffruticosa Andrews [35]. It was found that the xylem contained relatively fewer types and lower levels of chemical constituents, so the processing method of removing wood core while retaining cork bark for Paeonia x suffruticosa Andrews was recommended.

The quality of MC is highly susceptible to sulfuring and desulfuring. Through metabolomic analysis, 28 compounds were screened and identified as potential markers for distinguishing normal, sulfured, and desulfured MC, providing a basis for evaluating its quality [36]. The phenol extraction process from MC was optimized using single-factor experiments and response surface methodology (RSM). Under optimal conditions (liquid-to-solid ratio of 21 : 1 mL/g, ethanol concentration of 62%, ultrasonic time of 31 minutes, temperature of 36 °C, and power of 420 W), the phenol yield reached 14.01 mg/g. Following optimization, the phenolic purity of the HPD-300 macroporous resin increased to 41.40%, 5.97 times higher than that of the crude extract. Qualitative analysis identified seven compounds, including gallic acid, oxypaeoniflorin, paeonolide, paeonol, paeoniflorin, ethyl gallate, and pentagalloylglucose [37]. Optimizing the preparation process of extracts enhances the concentration and purity of active ingredients, thereby improving their effectiveness for various applications.

Traditional methods of evaluating phytomedicinal quality often lack reproducibility and the capability to objectively quantify components, resulting in subjective assessments. Advances in artificial intelligence, large models, and machine learning have enabled objective quantification of phytomedicinal evaluation, improving standardization and consistency of its quality assessment. By integrating electronic nose technology with chemometrics, reliable quantitative models for gallic acid, 5-HMF, paeoniflorin, and paeonol were developed to digitize the aroma of MC [38]. This will likely enhance the integration of traditional medicine with modern healthcare systems.

The standardization of MC extracts remains a critical yet challenging prerequisite for ensuring pharmacological reproducibility, safety, and regulatory acceptance [39]. One major challenge lies in the natural variability in the content of key bioactive compounds due to factors including geographical origin, cultivation conditions, harvest time, and post-harvest processing methods. Furthermore, the lack of universally accepted analytical protocols hinders the establishment of definitive chemical profiles for MC extracts. Building upon the AI-driven strategies, the integration of deep learning with non-destructive techniques offers a promising solution to standardize MC quality control. AI models can effectively analyze spectral data to accurately predict key bioactive compound levels, even amidst natural variability. They also standardize analysis by learning consistent patterns, reducing reliance on inconsistent protocols and aiding definitive extract chemical profiles for quality standards [40], [41].

Pharmacokinetics

Pharmacokinetic studies are essential for analyzing the absorption, distribution, metabolism, and elimination profiles of drug components, enabling the identification of key factors influencing their bioavailability and therapeutic effects.

An organism operates as an integrated system, and when it enters a pathological state, its endogenous metabolic components are collectively disturbed. Previous studies have shown that in the BHH (blood-heat and hemorrhage) model, 24 endogenous metabolic components underwent significant alterations, affecting key metabolic pathways such as glycerophospholipid metabolism and arachidonic acid metabolism [42], [43].

A rapid and accurate UHPLC-DAD method was developed for the simultaneous quantification of nine absorbed components in the serum of rats after administration of raw MC (RMC) and processed MC (PMC) extracts. Through pharmacokinetic analysis, the mean concentration-time profiles and relevant pharmacokinetic parameters were obtained. Notably, gallic acid, 5-HMF, and methyl paraben exhibited higher exposure, bioavailability, and slower elimination rates in the BHH-PMC group compared to the BHH-RMC group. These findings also indicated that frying charcoal enhanced the absorption of these active compounds. However, oxypaeoniflorin was not detected in the BHH-PMC group, which may be due to the degradation or structural changes of MC during the processing [44].

Systematically characterizing the in vivo metabolites is essential for gaining a deeper understanding of the mechanism of the active compound. To profile the metabolites of paeonol, a reliable strategy combining UPLC-Q/TOF-MS analysis was proposed to analyze the metabolites in the urine, feces, bile, and plasma of rats following paeonol administration. As a result, 25 metabolites were identified across these samples, while 14 of these metabolites have been previously unreported. In a word, the dominant metabolic pathways included oxidation, demethylation, hydrogenation, glucuronic acid, and sulfate conjugations. Additionally, the hydrogenation of paeonol was reported for the first time [45].

Anti-Fibrosis-Related Pharmacological Mechanisms

Paeonol and paeoniflorin are present in relatively high concentrations in the MC. Scientific research has revealed that both paeonol and paeoniflorin possess a multitude of biological activities, including anti-inflammatory, anti-oxidant, and immunomodulatory effects [46], [47], [48]. These findings suggest that they may serve as the key substance basis for exerting anti-fibrotic effects. For fibrosis occurring at distinct pathological sites, a comprehensive summary ([Fig. 2 ]) has been compiled on the pharmacological mechanisms through which paeonol ([Table 1]) and paeoniflorin ([Table 2]) exert their therapeutic effects against fibrosis.

Fig. 2 Multi-organ pharmacological mechanisms of the anti-fibrotic-related effects of paeonol and paeoniflorin. The diagram summarizes the key molecular targets and signaling pathways through which paeonol and paeoniflorin ameliorate fibrosis across different target organs.

Table 1 The therapeutic mechanisms of paeonol or its derivatives in anti-fibrosis.

Models	Animals and administered doses	Cell line and administered doses	Site of action	Therapeutic mechanisms	References
ApoE mice were fed with a high fat diet to establish an atherosclerosis model.	ApoE-/- mice. Paeonol was at doses of 200 and 400 mg/kg/d.	/	Aorta	Increasing intestinal microbiota-derived SCFA and restoring Treg/Th17 balance. Anti-inflammatory effect	[54]
Myocardial pathological remodeling (including apoptosis, hypertrophy, and fibrosis) was induced by intraperitoneal injection of adriamycin.	Danio rerio. Paeonol was at a dose of 28.84 µg/g/d.	Rat embryonic cardiomyoblast cell line H9c2. Paeonol was at concentrations ranging from 1 to 2.5 mM.	Heart	Regulation of Notch1 signaling pathway	[55]
TAC-induced HF in mice.	Male C57BL/6 mice. Paeonol was at doses of 10, 20, and 50 mg/kg/d.	/	Heart	Anti-inflammatory and anti-cardiomyocyte apoptosis effects; regulating the ERK1/2/JNK signaling pathway	[56]
A mouse model of BLM was established by intratracheal instillation.	Male C57BL/6 mice. Paeonol was at a dose of 10 mg/kg.	Human fetal lung fibroblast. Paeonol was at concentrations ranging from 0 to 0.4 mM.	Lung	Anti-inflammatory, anti-oxidative stress effects enhance the proportion of Treg cells in CD4+T cells; regulating MAPKs/Smad3 signaling	[65], [66]
Pulmonary fibrosis was induced by intratracheal instillation of TNF-α.	Male C57BL/6 mice. Paeonol was at doses of 10 and 30 mg/kg/d.	/	Lung	Anti-inflammatory and anti-oxidative stress effects	[66]
A model of hepatitis and fibrosis induced by thioacetamide injection.	Male C57BL/6 J mice. Paeonol was at doses of 2.5 and 40 mg/kg/d.	Human HSCs line (LX-2). Paeonol was at concentrations ranging from 5 to 100 µM.	Liver	Regulation of serum liver injury markers, TGF-β/Smad2/3 and STAT3 phosphorylation signaling pathways	[78]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice	Male C57BL/6 J mice. Paeonol was at a dose of 100 mg/kg/d.	/	Liver	Anti-oxidative stress and regulating TGF-β/Smad3 signaling pathway	[79]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice.	Male SD rat. Paeonol was at doses of 50, 100, and 200 mg/kg/d	HSCs. Paeonol was at concentrations of 10, 20, 40, and 60 µmol/L	Liver	Regulation of NF-kB signaling pathway and mitochondrial apoptosis in HSCs	[80]
The hepatotoxicity model was induced by intraperitoneal injection of MTX.	Male Wistar rat. Paeonol was at a dose of 100 mg/kg/d.	/	Liver	Anti-oxidative stress. Regulation of serum liver injury indicators, HSCs apoptosis, and efflux protein secretion	[81]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice.	Male SD rat. Paeonol was at a dose of 100 mg/kg/d.	Hepatic stellate cells line (HSC-T6). Paeonol was at a concentration of 25 µM/L.	Liver	Anti-inflammatory and regulating serum liver injury markers, hepatic stellate cell apoptosis, and TGF-β/Smad2/3 signaling pathway	[82]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice.	Male C57BL/6 mice. 4-methoxysulfonyl paeonol was at a dose of 5 mg/kg/d (intraperitoneal injection).	Human HSCs line (LX2) and rat HSCs line (HSC-T6). Paeonol was at a concentration of 5 µM.	Liver	Regulate the TGF-β1/Smad, PDGF-BB/MAPK and Akt signaling pathways and inhibit the proliferation of HSCs	[83]
Diabetic nephropathy induced by intraperitoneal STZ injection.	Male C57BL/6 mice. Paeonol was at a dose of 150 mg/kg/d.	GMCs. Paeonol was at concentrations of 5, 10, and 20 µg/mL.	Kidney	It has anti-inflammatory and anti-oxidative stress effects and regulates the Nrf2/ARE signaling pathway	[103]
Renal interstitial fibrosis rat model was established by UUO.	Male SD rat. Paeonol was at a dose of 50 mg/kg/d.	NRK-49F cell. Paeonol was at a concentration of 100 µM.	Kidney	Inhibition of inflammatory response and cell migration. Regulation of HOTAIR/miR-124/Notch1 axis, Notch1/Jagged1 signaling pathway	[104]
Arecoline-induced myofibroblast activation model.	/	Normal BMFs and fBMFs. Paeonol was at concentrations of 10 and 20 µM.	/	Regulates the TGF-β1/Smad2/3 signaling pathway	[108]
STZ induces diabetic erectile dysfunction in mice.	Male SD rat. Paeonol was at a dose of 100 mg/kg/d.	Primary rat CCSMCs. Paeonol was at a concentration of 100 µM.	Organs of reproduction	Regulates TGF-β1/Smad2/3 and HMGB1/RAGE/NF-kB signaling pathways	[109]

Table 2 The therapeutic mechanisms of paeoniflorin in anti-fibrosis.

Models	Animals and administered doses	Cell line and administered doses	Site of action	Therapeutic mechanisms	References
Aortic banding was used to establish a cardiac overloaded model.	Male C57BL/6 mice. Paeoniflorin was at a dose of 20 mg/kg/d.	/	Heart	Anti-inflammatory and anti-cardiomyocyte apoptosis; regulating TGF-β/Smads and NF-κB signaling pathways	[57]
Spontaneous hypertensive.	Male SHR and male Wistar-Kyoto rat. Paeoniflorin was at a dose of 9 mg/kg/d.	/	Heart	Anti-inflammatory effect; regulating MAPK and ERK1/2/JNK signaling pathways	[58]
The mice model was established by subcutaneous injection of Ang II.	Male C57BL/6 mice. Paeoniflorin was at a dose of 100 mg/kg/d.	/	Heart	Anti-apoptosis and anti-oxidative stress effects; regulating the PI3K-Akt signaling pathway	[60]
MI models were established in 60 rats by ligation at the origin of the left anterior descending coronary artery.	Male SD rats. at Paeoniflorin was at doses of 2.25, 4.5, and 9 mg/kg/d.	/	Heart and blood vessels	Anti-apoptotic, anti-inflammatory and anti-oxidative stress effects	[59]
Chronic heart failure model.	Male Wistar rat. Paeoniflorin was at a dose of 20 mg/kg/d.	Cardiac fibroblasts from neonatal Wistar rats. Paeoniflorin was at a concentration of 100 µmol/L.	Heart	Regulating the p38/MAPK signaling pathway	[61]
A mouse model of BLM was established by intratracheal instillation.	Male ICR mice. Paeoniflorin was at doses of 25, 50, and 100 mg/kg/d.	Human alveolar epithelial cells (A549 cells). Paeoniflorin was at concentrations of 1, 3, 10, and 30 µmol/L.	Lung	Regulating the TGF-β1/Smad signaling pathway	[68]
The fibrosis model of acute lung injury induced by influenza A/FM/1/47 (H1N1).	Female BALB/c mice. Paeoniflorin was at doses of 50 and100 mg/kg/d.	/	Lung	Anti-inflammatory effect. Regulation of NF-κB, p38MAPK and αvβ3/TGF-β1 signaling pathways	[69]
Model of silicosis induced by silica.	Male C57BL/6 J mice. Paeoniflorin was at doses of 50 and100 mg/kg/d.	RAW264.7 cells and mouse lung fibroblasts.	Lung	Anti-inflammatory effect and inhibition of fibroblast differentiation	[70]
ANIT-induced cholestatic liver fibrosis model.	Male SD rats. Paeoniflorin was at doses of 50, 100, and 200 mg/kg/d.	/	Liver	Regulation of serum markers of liver injury and activation of Nrf2 via PI3K/AKT-dependent pathway	[84], [85]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice.	Male SD rats. Paeoniflorin was at doses of 80 and 200 mg/kg/d.	Rat HSCs line (HSC-T6). Paeoniflorin was at a concentration of 0.2 µg/mL	Liver	Regulating serum liver injury indicators, promoting the apoptosis of HSCs, and inhibiting the activation of HSCs	[86]
Mouse model of primary biliary cholangitis.	Female mice. Paeoniflorin was at a dose of 50 mg/kg/d.	/	Liver	Anti-oxidative stress and regulation of serum liver injury indicators	[87]
Dimethylnitrosamine-induced liver fibrosis.	Male Wistar rat. Paeoniflorin was at a dose of 20 mg/kg/d.	/	Liver	Anti-inflammatory effect; regulation of serum markers of liver injury and macrophage viability	[88]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice.	Male C57BL/6 mice. Paeoniflorin was at doses of 50 and 100 mg/kg/day.	Human hepatic stellate cell line LX-2	Liver	Inhibition of HSCs activation and H3K27 trimethylation	[89]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice.	Male C57BL/6 mice. Paeoniflorin was at a dose of 100 mg/kg/d.	/	Liver	Inhibit the activation of HSCs, anti-inflammatory and anti-oxidative stress effects	[90]
Intraperitoneal injection of CCl4 was used to induce liver fibrosis in mice.	Male C57BL/6 mice. Paeoniflorin was at a dose of 50 mg/kg/d.	/	Liver	Regulating the TGF-β/Smad signaling pathway	[91]
X-ray radiation-induced liver fibrosis.	Male SD rats. Paeoniflorin was at doses of 20, 40, and 80 mg/kg/d.	/	Liver	Regulating Smad-related pathways	[92]
Hepatic fibrosis induced by Schistosoma mansoni.	Male BALB/c Swiss albino mice. Paeoniflorin was at a dose of 50 mg/kg/d.	/	Liver	Anti-angiogenic effect	[93]
Hepatic fibrosis caused by Schistosoma japonicum infection.	BALB/c mice. Paeoniflorin was at doses of 30 and 60 mg/kg/d.	Kupffer cell. Paeoniflorin was at a concentration of 100 µg/mL; HSCs. Paeoniflorin was at concentrations of 0, 30, 60, and 120 mg/L.	Liver	Anti-inflammatory effect, inhibition of macrophage activation and regulation of JAK2, STAT6, and TGF-β1/Smads signaling pathway	[94], [95], [96], [97], [98]
Renal interstitial fibrosis in mice with UUO.	Male ICR mice. Paeoniflorin was at doses of 25, 50, and 100 mg/kg.	/	Kidney	Regulating the TGF-β/Smad signaling pathway	[105]
Models of diabetic nephropathy.	/	Mouse podocyte (MPC-5). Paeoniflorin was at a concentration of 80 µM.	Kidney	Alleviating cell damage	[106]
Chronic renal failure rats and diabetic kidney disease db/db mice	Male SD rats. Paeoniflorin was at doses of 25 and 50 mg/kg. db/db mice Paeoniflorin was at doses of 50 and 100 mg/kg.	Human umbilical vein endothelial cells and rat kidney fibroblast cell line Paeoniflorin was at a concentration of 400 µM.	Kidney	Reducing ECM deposition; inhibiting Piezo1/Ca²⁺/HIF-1α signaling axis	[106]
The rat model for PI-IBS was established using a multistimulation paradigm composed of EPSD, TNBS, and CUMS.	Adult pregnant SD rats. Paeoniflorin was at doses of 10, 20, and 40 mg/kg/d.	/	Colonic submucosa and smooth muscle	PI3K/Akt and leptin/LepRb pathway	[110]

Cardiovascular fibrosis

Cardiovascular fibrosis is a pathological process characterized by excessive deposition of extracellular matrix (ECM), and its formation involves complex molecular and cellular interactions. Key driving factors include chronic inflammatory responses, oxidative stress, mechanical stress, and abnormal activation of various pro-fibrotic factors (such as TGF-β, CTGF, and Ang II). These factors promote the transformation of fibroblasts into myofibroblasts, enhance the synthesis and deposition of ECM proteins (such as collagen and fibronectin), and simultaneously inhibit the activity of matrix metalloproteinases (MMPs), leading to reduced ECM degradation. Additionally, endothelial cell dysfunction, immune cell infiltration, and dysregulation of epigenetic mechanisms play significant roles in the development and progression of cardiovascular fibrosis. Collectively, these processes contribute to the structural remodeling of the myocardium and vascular walls, ultimately resulting in impaired organ function [49], [50], [51], [52], [53].

Paeonol exerts multiple protective effects in cardiovascular fibrosis models. It suppresses inflammatory responses by lowering pro-inflammatory factors (IL-1β, IL-6, TNF-α, and IL-17A) and raising anti-inflammatory factors IL-10 [54]. It also reduces cardiomyocyte apoptosis by adjusting the Bcl-2/Bax ratio and decreasing cleaved caspase-3 in cardiac cells [55], [56]. Additionally, paeonol decreases myocardial Col1, Col3, CTGF, and FN-1 to inhibit collagen deposition in cardiac ECM [56]. Paeonol protects cardiomyocytes from doxorubicin-induced hypertrophy and fibrosis via activating the Notch1 pathway and its downstream genes [55]. Another study showed it inhibits aortic plaque formation, collagen deposition, and vascular fibrosis. This happens by increasing gut microbiota-derived SCFAs, restoring the regulatory T cells/T helper 17 cells (Treg/Th17) balance, and reducing lysyl oxidase and MMP-2/9 [54]. In transverse aortic constriction (TAC)-induced heart failure (HF) mice, paeonol can suppresses the ERK1/2 (extracellular signal-regulated kinase 1/2)/JNK (c-Jun N-terminal kinase) pathway [56].

Paeoniflorin exhibits multifaceted protective effects in cardiovascular fibrosis. It alleviates inflammatory responses by inhibiting myocardial NLRP3 (NLR family pyrin domain-containing 3) inflammasome, reducing the levels of NLRP3 and caspase-1, and modulating the expression of cytokines such as TNF-α, IL-6, IL-10, IL-18, and IL-1β [57], [58], [59]. It also attenuates cardiomyocyte apoptosis by upregulating anti-apoptotic Bcl-2 and downregulating pro-apoptotic Bax [59], [60]. Additionally, paeoniflorin inhibits collagen accumulation via reducing TGF-β1, CTGF, Col1, Col3, MMP-9, and α-SMA expression [57], [58], [60]. Simultaneously, paeoniflorin significantly improves oxidative stress by enhancing the levels of endogenous anti-oxidants such as CAT, SOD, and GSH, while reducing the levels of NOX2, NOX4, malondialdehyde (MDA), and ROS [59], [60].

In the myocardial fibrosis model induced by pressure overload, peony glycoside alleviates fibrosis by inhibiting the activation of TGF-β/Smads, MAPK (mitogen-activated protein kinase), and NF-κB (nuclear factor kappa B) pathways in cardiomyocytes. [57], [58]. Furthermore, paeoniflorin improves interstitial and perivascular fibrosis during left ventricular remodeling after acute myocardial infarction (MI) by reducing FDX1/DLAT expression and serum copper levels, while increasing pyruvate levels [59]. In a mouse model of atrial fibrosis induced by angiotensin II, peony glycoside improved cardiac function by reducing the levels of cTnI, cTnT, ANP, and BNP in the serum and exerted anti-fibrotic effects by blocking the activation of the PI3K-Akt pathway [60]. Additionally, paeoniflorin mitigates myocardial fibrosis and improves cardiac function in chronic HF rats by downregulating the p38/MAPK signaling pathway and reducing the perivascular collagen volume fraction [61].

Pulmonary fibrosis

Pulmonary fibrosis is a chronic and progressive pathological condition characterized by excessive accumulation of ECM in lung tissue and the re-epithelialization of the alveolar surface. This can be triggered by a variety of factors, including smoking, air pollution, medicines, radiation, or viral infections. The pathogenesis of pulmonary fibrosis involves complex molecular and cellular interactions. The key drivers are recurrent alveolar epithelial cell damage, dysregulated repair responses, and the persistent effects of oxidative stress and a chronic inflammatory microenvironment [62], [63], [64].

Studies have demonstrated that paeonol exhibits significant anti-inflammatory, anti-fibrotic, and anti-oxidant effects in pulmonary fibrosis models. It alleviates bleomycin (BLM)-induced pulmonary fibrosis in mice, as evidenced by the decreased total cell count, differential cell count, and total protein concentration in bronchoalveolar lavage fluid (BALF), as well as the reduction in collagen deposition and the levels of modified proteins such as malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) as oxidative stress markers. Furthermore, paeonol dose-dependently inhibits TGF-β1-induced expression of α-SMA and Col1A1 in human lung fibroblasts and effectively suppresses fibrotic responses by inhibiting the MAPKs/Smad3 signaling pathway. Paeonol also mitigates TNF-α-induced lung injury through multiple mechanisms. It improves smooth muscle cell hypertrophy and basement membrane thickening from TNF-α stimulation, reduces pulmonary edema, immune cell infiltration, and IL-1β levels in BALF, and decreases lung interstitial collagen deposition. Additionally, it exerts anti-oxidant effects by downregulating iNOS and COX-2 protein expression and upregulating catalase and the glutamate-cysteine ligase modifier subunit, thus protecting against acute lung injury (ALI)-induced pulmonary fibrosis [65], [66]. Furthermore, paeonol inhibits EP300 expression in CD4+T cells and primary mouse lung fibroblasts via the EP300/Foxp3 axis, increasing the proportion of Treg cells in CD4+T cells [67].

Paeoniflorin exerts notable anti-inflammatory and anti-fibrotic effects in pulmonary fibrosis treatment. Studies show it notably decreases MMP-12 and multiple inflammation-related factors (such as IFN-γ, MCP-1, TLR4, IL-6, IL-1β, IL-17A, and TNF-α) in fibroblasts, while decreasing the expression of fibrotic markers (including Col 1, Col 3, Fn1, α-SMA, PAI-1, FSP-1, and vimentin) and increasing E-cadherin expression in lung tissue, thereby effectively inhibiting the progression of fibrosis [68], [69], [70]. Furthermore, paeoniflorin significantly suppresses morphological changes and migration capabilities of A549 cells and attenuates TGF-β1-induced alveolar epithelial-mesenchymal transition (EMT) by inhibiting the expression of Snail, Smad2/3, and p-Smad2/3, downregulating activin receptor-like kinase 5, and upregulating Smad7 [70]. In the acute lung injury (ALI) model caused by the influenza A virus (IAV), peony glycoside exhibits a significant protective effect against pulmonary fibrosis induced by ALI by inhibiting the activation of the NF-κB, p38MAPK, and αvβ3/TGF-β1 signaling pathways [69].

Hepatic fibrosis

The main driver of hepatic fibrosis is the persistent inflammatory response and hepatocyte damage caused by chronic liver injury caused by alcohol abuse, viral infection, cholestasis, high-fat diets, etc. A key factor is the aberrant activation of profibrotic mediators such as TGF-β, PDGF, and CTGF, which promote the transformation of hepatic stellate cells (HSCs) into myofibroblasts, leading to excessive synthesis and deposition of ECM components including collagen, fibronectin, and laminin. Also, the imbalance between matrix metalloproteinases (MMPs) and their tissue inhibitors hinders ECM degradation, worsening fibrosis. In addition, oxidative stress, endoplasmic reticulum stress, abnormal activation of immune cells (like macrophages), and epigenetic modification dysregulation all play roles in liver fibrosis development. These processes disrupt hepatic lobular structure and impair liver function, potentially leading to cirrhosis and liver failure [71], [72], [73], [74], [75], [76], [77].

In vivo, paeonol shows hepatoprotective effects in hepatic fibrosis models. It lowers serum levels of ALT, AST, ALP, and LDH, reducing liver injury. It also mitigates fibrosis progression by decreasing hepatic laminin, α-SMA, Col1, FN, CTGF, HA, PCIII, and hydroxyproline. Furthermore, paeonol lessens inflammation and oxidative stress: it reduces inflammatory factors (e.g., TNF-α and IL-6), enhancing the activity of anti-oxidant enzymes (e.g., GSH-PX, SOD, and CAT), and lowering the expression of oxidative stress markers (e.g., NO, MDA, NOX1, NOX2, and GSH) [78], [79], [80], [81], [82]. In vitro, paeonol upregulates P21 and P27 in HSCs, while inhibiting the expression of Bcl-2 promoting the expression of Bax, caspase-3, caspase-9, and PARP-1, thereby inducing HSCs apoptosis [79], [80], [82].

In the CCl₄-induced hepatic fibrosis model, paeonol exerts anti-fibrotic effects through multiple mechanisms. First, paeonol inhibits the cell cycle progression of HSCs at the G2/M checkpoint, promotes the release of cytochrome c, modulates mitochondrial permeability, and reduces ATP production, thereby suppressing mitochondrial function in HSCs [79], [80], [82]. It also upregulates BMP-2, MMP-2, and MMP-9 in HSCs and downregulates PDGF and TIMP-1. This inhibits TGF-β/Smad2/3, NF-κB, and STAT3 phosphorylation pathways, suppressing HSC activation, reducing ECM collagen deposition, and accelerating its degradation [78], [79], [80], [82].

Further studies indicate that paeonol enhances the protein expression of P-glycoprotein and multidrug-resistance-associated protein 2 in HSCs, reducing methotrexate (MTX)-induced liver injury via efflux, protecting the liver [81]. Moreover, paeonol derivatives also demonstrate significant anti-fibrotic potential. They can inhibit the TGF-β1/Smad, PDGF-BB/MAPK, and Akt signaling pathways, thereby reducing collagen deposition and liver damage in the liver and effectively preventing the progression of liver fibrosis [83].

In hepatic fibrosis models, paeoniflorin demonstrates multifaceted protective effects. First, it significantly improves liver function by reducing serum levels of ALT, AST, ALP, and TBiL (total bilirubin) [84], [85], [86], [87], [88], [89]. Simultaneously, paeoniflorin decreases the production of inflammatory cytokines such as IL-1β, IL-6, and TNF-α, while enhancing the activities of HO-1, SOD, CAT, and GSH-PX and reducing MDA levels, thereby effectively alleviating inflammatory responses and oxidative stress [84], [85], [87], [90]. Furthermore, paeoniflorin inhibits the gene and protein expression of α-SMA, Col1A1, desmin, vimentin, FN, TIMP, and Hyp, reducing the deposition of ECM and further suppressing the progression of fibrosis [88], [89], [90], [91], [92].

Paeoniflorin in CCl₄-induced fibrosis inhibits HSC activation by suppressing the enhancer of the zeste homolog 2-mediated H3K27 trimethylation (H3K27me3) and upregulating PPARγ. Simultaneously, paeoniflorin promotes HSCs apoptosis by upregulating caspase-3 expression and downregulating the Bcl-2/Bax ratio. Additionally, it exerts anti-fibrotic effects by upregulating Smad7 and downregulating the expression of TGF-β1 and p-Smad2, thereby inhibiting the TGF-β/Smad signaling pathway [86], [89], [90], [91]. In the α-naphthylisothiocyanate (ANIT)-induced cholestatic model, paeoniflorin lowers TBIL, DBIL, TBA, and γ-GT, improving liver injury and bile duct proliferation in rats. This may involve activating the PI3K/Akt pathway, upregulating p-Akt and Nrf2 (nuclear factor erythroid 2-related factor 2), and enhancing GSH synthesis [84], [85].

In the schistosomiasis-induced hepatic fibrosis model, paeoniflorin inhibits fibrosis by reducing the levels of pro-angiogenic factors (such as VEGF, PCNA, α-SMA, and CD34) in liver tissue and increases TIMP-2, exerting anti-angiogenic effects [93]. Furthermore, paeoniflorin directly suppresses the alternative activation of macrophages (AAMs) by reducing the phosphorylation of JAK2 and STAT6, decreasing AAM-related markers and Arg-1 activity. It also indirectly inhibits AAMs by lowering IL-13, suppressing HSC proliferation. Furthermore, paeoniflorin exerts an anti-schistosomal liver fibrosis effect by inhibiting the TGF-β1/Smads signaling pathway [94], [95], [96], [97], [98].

Moreover, paeoniflorin demonstrates significant protective effects in other hepatic fibrosis models. For instance, it ameliorates dimethylnitrosamine-induced hepatic fibrosis by inhibiting macrophage activation and reducing CD68 expression [88], alleviates radiation-induced hepatic fibrosis by downregulating Smad3/4 expression and upregulating Smad7 expression [92], and mitigates liver injury and fibrosis in primary biliary cholangitis mice by inhibiting NLRP3 inflammasome formation and its cascade inflammatory responses [87].

Renal fibrosis

Renal fibrosis primarily arises from persistent inflammatory responses and renal tubular epithelial cell injury triggered by chronic kidney damage, such as diabetic nephropathy, hypertensive nephropathy, and chronic glomerulonephritis. A key factor is the aberrant activation of pro-fibrotic mediators, which promotes the transformation of fibroblasts and renal tubular epithelial cells into myofibroblasts [99], [100], [101], [102].

Paeonol exhibits multifaceted protective effects in renal fibrosis models. First, paeonol upregulates Sirt1 and activates the Nrf2/ARE signaling pathway in glomerular mesangial cells (GMCs), promoting the expression of Nrf2, HO-1, and SOD1 while reducing the levels of H2O2 and MDA. It also suppresses the expression of inflammatory and fibrotic factors (such as FN and ICAM-1), thereby ameliorating high glucose-induced oxidative stress and alleviating renal dysfunction and fibrosis in streptozotocin (STZ)-induced diabetic nephropathy mice [103]. Moreover, paeonol notably reduces HOTAIR (HOX transcript antisense RNA), α-SMA, vimentin, MMP-2, and MMP-9 in rat kidneys and increases E-cadherin, inhibiting HOTAIR-mediated EMT. This relieves severe inflammation, necrosis, and tubular swelling in HOTAIR rats and reduces renal interstitial collagen deposition. In addition, paeonol alleviated fibrosis in a rat model of renal interstitial fibrosis induced by unilateral ureteral obstruction (UUO) by reversing HOTAIR-induced activation of the Notch1/Jagged1 pathway [104].

Paeoniflorin demonstrates significant therapeutic efficacy in treating renal interstitial fibrosis in a UUO mouse model. First, paeoniflorin effectively inhibited the EMT process by reducing the expression of hydroxyproline, collagen, and plasminogen activator inhibitor-1 (PAI-1). Second, it could significantly improve renal interstitial fibrosis in UUO mice by inhibiting the TGF-β/Smad signaling pathway and restoring BMP-7 function [105]. On the other hand, paeoniflorin also reduces the expression of H2-Aa and inhibits T cell activation, particularly decreasing the proportion of Th1 cells, thereby alleviating fibrosis in glomerular podocytes in diabetic nephropathy mice [106]. It can also inhibit the Piezo1/Ca²⁺/HIF-1α signaling axis to promote renal microcirculation repair, reduce the expression of Col1 and FN, decrease ECM deposition, and downregulate the protein expression of vimentin and TGF-β1. This effectively improves renal function in chronic kidney disease and delays the progression of renal fibrosis [107].

Others

In the treatment of other fibrotic diseases, paeonol demonstrates broad therapeutic potential. In the management of oral submucosal fibrosis, paeonol reduces the expression of fibrotic markers such as α-SMA, Col1, and HOTAIR in fibrotic buccal mucosal fibroblasts (fBMFs) and inhibits the activation of the TGF-β/Smad2 signaling pathway, thereby attenuating myofibroblast activity and offering a novel strategy for treating oral submucosal fibrosis [108]. Additionally, in a mouse model of diabetic erectile dysfunction, paeonol significantly ameliorates inflammatory responses by reducing the expression of inflammatory factors, including NLRP3, ASC, caspase-1, IL-6, IL-1β, and TNF-α. Simultaneously, paeonol inhibits apoptosis of corpus cavernosum smooth muscle cells (CCSMCs) by decreasing the Bax/Bcl-2 ratio and suppressing the expression of Bad, caspase-3, and cleaved caspase-3. Furthermore, diosgenin can alleviate erectile dysfunction and penile fibrosis by inhibiting the HMGB1/RAGE/NF-κB signaling pathway and reducing the expression of TGF-β1, Smad2/3, p-Smad2/3, CTGF, and α-SMA in penile tissues [109].

Paeoniflorin also exhibits significant anti-fibrotic effects in a rat model of post-infectious irritable bowel syndrome (PI-IBS). Studies have shown that paeoniflorin inhibits the expression of leptin/LepRb in the colon and serum and modulates the leptin/PI3K/AKT pathway, thereby effectively reducing the excessive fibrosis of the colonic submucosa and smooth muscle [110].

Application and Development

For basic scientific research, both patent and clinical research are of great significance. Patents protect innovation and facilitate the transformation of technological achievements into applications. Clinical research focuses on drug/treatment safety and effectiveness in humans, which is related to improving medical standards and patient health.

Patent analysis

To comprehensively gather patent information related to paeonol and paeoniflorin, this study utilized the Patentstar database, which offers rich data sources and timely updates [111]. Using the ‘expert search’ approach, the invention names ‘paeonol’ and ‘paeoniflorin’ were respectively used as search keywords to search within the scope of ‘World Patents’.

As a result, there are a total of 246 patents related to paeonol and 265 patents related to paeoniflorin. The geographical analysis results ([Fig. 3 a] and [b]) indicate that the patents applied for in China account for the vast majority, demonstrating that China has a profound accumulation of the research and technological innovation of paeonol and paeoniflorin. In addition, countries such as the United States, Korea, Japan, and those in Europe have also applied for a certain number of patents. This also reflects the potential market value of paeonol and paeoniflorin in these countries. It is worth noting that the WIPO has also included relevant patents, which means that paeonol and paeoniflorin have a certain universality and recognition globally. Overall, this distribution pattern of paeonol and paeoniflorin patents not only highlights Chinaʼs leading edge in this field but also reflects the global popularity and development potential of research on paeonol and paeoniflorin.

Fig. 3 Patent and clinical research progress of paeonol and paeoniflorin. Paeonol: a Statistics of the national distribution; c Statistics of patent IPC classification; e Statistics of patents in different fields. Paeoniflorin: b Statistics of the national distribution; d Statistics of patent IPC classification; f Statistics of patents in different fields.

The IPC (International Patent Classification) classification analysis of patents related to paeonol ([Fig. 3 c]) indicates that these patents are mainly concentrated in A (human necessities) and C (chemistry; metallurgy), specifically in the fields of pharmaceutical preparations, extraction processes, and their medical applications [112]. From the perspective of IPC subclass, most of the patents involving paeonol fall into A61K (preparations for medical, dental, or toilet purposes) and A61P (medicinal preparations for specific therapeutic purposes). Patents in A61K are mainly related to the extraction methods, purification techniques, and the development of dosage forms of paeonol, while patents in A61P focus on the applications of paeonol in the treatment of specific diseases, such as for anti-inflammation, anti-oxidation, anti-fibrosis, etc.

Similar to paeonol, patents related to paeoniflorin also have a relatively high proportion in A61K and A61P ([Fig. 3 d]). Notably, the presence of paeoniflorin-related patents in A23L (foodstuffs or non-alcoholic beverages; their preparation or treatment) indicates it has potential uses in the food industry. This may be attributed to the health-promoting properties of paeoniflorin, such as its anti-oxidant and anti-inflammatory effects, which can improve the nutritional value or quality of food. Previous studies have shown that paeoniflorin has a significant antimicrobial effect against carbapenem-resistant Klebsiella pneumoniae [113]. It is expected to be applied to prevent food contamination, which is conducive to food freshness and preservation. In addition, some research has also provided a valuable theoretical basis for the application of paeoniflorin as a functional food for hyperlipidemia [114].

In parallel, patents were classified across various research fields ([Fig. 3 e] and [f]), revealing that paeonol and paeoniflorin are primarily focused on development and application in the medicine and cosmetic fields. The main therapeutic effects of paeonol and paeoniflorin have been discussed in the previous content. Notably, paeonol and paeoniflorin have scientifically proven cosmetic effects, such as anti-aging, skin-whitening, and the treatment of atopic dermatitis [115], [116], [117]. In the field of organic chemistry, the chemical structure modification and synthetic method optimization of paeonol and paeoniflorin are the research focuses. By modifying their molecular structures, derivatives with higher activity, stability, and bioavailability can be developed, thus expanding their applications in pharmaceutical and other fields [118], [119]. In the field of biotechnology, research on paeonol and paeoniflorin mainly focuses on the innovation of extraction processes and the optimization of content determination methods, which provides excellent technical support for subsequent in-depth research and applications [37], [120], [121].

In summary, this reflects that under the cross-disciplinary research, paeonol and paeoniflorin are constantly expanding their application boundaries, making more positive impacts on human health and life.

Clinical research progress

To obtain the latest research progress on the clinical research of paeonol and paeoniflorin, the Pharmacodia, PubMed, and CNKI databases were integrated for this retrieval. The Pharmacodia database encompasses the official websites of various countries, pharmaceutical company websites, literature, etc., providing detailed information on clinical drug studies. Searches were respectively conducted using ‘paeonol’ and ‘paeoniflorin’ as drug interventions or titles, and the relevant clinical research is summarized in [Table 3].

Table 3 Clinical research information on paeonol and paeoniflorin.

No.	Samples	Intervention	Disease	Trial registration ID	Trial status
Paeonol
1	n = 152	The synthetic combination of the isomers apocynin and paeonol was termed APPA	Knee Osteoarthritis (OA)	NCT04657926	Phase Ⅱ, Completed
	n = 150			EUC TR2020-000249-14-DK	Phase Ⅱ, Not recruiting
	n = 225			C TIS2023 – 508 791 – 11 – 00	Phase II, Recruiting
2	n = 90	Compound Paeonol Dripping Pill (CPDP)	Unstable Angina	/	/
3	n = 288	Safflower and Paeonol Ointment	Pain; Arthropathy; Soft Tissue Injury	CTR20131770	Phase Ⅱ, Completed
4	n = 92	Paeonol Ointment Combined with Tacrolimus Ointment	Skin Pruritus	/	/
5	n = 88	Toothpaste Containing Paeonol	Dentin Hypersensitivity	ChiCTR2000041417	N/A, Recruiting
6	n = 63	Toothpaste Containing Panax Notoginseng extract and Paeonol	Gingival Inflammation	/	/
7	n = 50	Sodium Paeonol Sulfate Combined with Nerve Block	Postherpetic Neuralgia	/	/
Paeoniflorin
1	n = 156	Paeoniflorin Combined with Photodynamic Therapy	Oral Lichen Planus (OLP)	NCT05973097	N/A, Recruiting
2	N/A	Paeoniflorin Combination of Hepatoprotective Drugs	Autoimmune Hepatitis	NCT02878863	Phase Ⅲ, Withdrawn
3	n = 42	Paeoniflorin Combined with Morphine	Pain of Advanced Cancer	/	/

Previous evidence has demonstrated that apocynin (AP) can inhibit the activation of NF-κB and upregulate the Nrf2 gene expression [122]. Paeonol (PA), by suppressing the expression of pro-inflammatory cytokines, metalloproteinases, and MCP-1, inhibits the activation of NF-κB [123]. Moreover, inhibiting NF-κB and upregulating Nrf2 are considered potential targets for the treatment of osteoarthritis (OA) [124]. In addition, there is also evidence suggesting the anti-inflammatory mechanism of the combined administration of apocynin and paeonol [125]. Inspired by these, Asger R. Bihlet et al. and Cao designed AP and paeonol PA into an oral fixed-combination product (APPA) with a ratio of 2 : 7 (AP : PA), to evaluate its effects on the knee OA. It is found that the oral administration of APPA at 800 mg daily for 28 consecutive days did not lead to statistically significant changes relative to placebo. Notably, the treatment was established to be safe and exhibited good tolerability among the subjects. Pre-planned subgroup analyses demonstrated a marked effect of APPA among subjects with nociplastic pain or severe OA, suggesting that further investigation into the effects of APPA in the relevant patient groups is justified [126].

Compound paeonol dripping pills (the content of paeonol is 0.3 mg per capsule) are more effective than Tongxinluo capsules and conventional Western medicine treatments in alleviating the symptoms of unstable angina and reducing the levels of inflammatory mediators [127]. Paeonol can also be utilized as a primary additive in the development of ointments. Clinically, the combination of paeonol ointment and tacrolimus ointment has been shown to effectively alleviate skin pruritus, reduce the levels of serum factors, and exhibit favorable safety profiles [128]. In clinical practice, toothpaste containing paeonol exhibits good anti-sensitivity and anti-inflammatory effects [129].

Paeoniflorin can effectively alleviate the oral inflammatory response and significantly improve the oral microenvironment and has a positive impact on the maintenance of oral health [130]. The study has revealed that paeoniflorin promotes the proliferation, migration, and multilineage differentiation of mesenchymal stem cells from oral lichen planus (OLP) lesions by regulating the Th1/Th2 balance, thereby prolonging skin graft survival and ameliorating inflammatory infiltration [131]. In clinical practice, paeoniflorin combined with morphine for the treatment of moderate-to-advanced cancer pain significantly reduces the morphine consumption by cancer patients and decreases the occurrence of morphine-related side effects [132].

In all, clinical research on paeonol and paeoniflorin has been steadily advancing in recent years. Clinical research findings have revealed that paeonol or paeoniflorin is almost always used in the form of combined interventions for treating specific diseases, in combination with other drugs or therapeutic techniques. In fact, they can enhance anti-inflammatory, anti-oxidant, and other therapeutic effects through synergistic actions, reduce side-effects by lowering the dosage of single drugs, and address complex disease mechanisms more comprehensively. Moreover, paeonol can be added as an active ingredient to products like ointments and toothpastes, endowing these products with anti-inflammatory, antibacterial, and analgesic properties. Its potential in cosmeceuticals also boosts commercial interest and expands its translational use beyond traditional pharmacotherapy.

Despite promising scientific research, patents, and clinical research results, the transition from bench to bedside remains fraught with challenges. First, key challenges include significant variability in administered doses across different animal and cellular models, complicating the direct comparison and translation of results. A lack of standardized dosing protocols and limited human pharmacokinetic data hinder the establishment of effective and reproducible dosing regimens for clinical applications. Second, methodological limitations–such as inadequate sample sizes, lack of randomization and blinding, and short trial durations–compromise the statistical power and reliability of many existing clinical studies. These shortcomings hinder definitive conclusions regarding the efficacy and safety of the interventions in human subjects. Third, there is a limited scope of indications–most clinical trials have focused on a narrow range of conditions, often prioritizing traditional applications over the novel disease targets suggested by preclinical data. Fourth, there is an absence of comprehensive safety profiling–structured long-term toxicological studies and systematic reporting of adverse effects in diverse populations are lacking. Additionally, potential compound-compound interactions continue to be underexplored.

To address these challenges, future studies should prioritize the following strategies: develop standardized dosing protocols for paeonol and paeoniflorin by establishing uniform dose ranges derived from pooled preclinical data and tailored to common animal and cellular models; conduct targeted human pharmacokinetic studies to characterize absorption, metabolism, and excretion to inform clinical dosing; implement rigorous, multicenter, randomized, double-blind, placebo-controlled trials with larger cohorts and extended follow-ups, incorporating adaptive designs for efficient dose optimization and patient stratification; leverage emerging technologies–such as artificial intelligence and big data analytics–to enhance target prediction, biomarker identification, and mechanistic insight; and enhance safety profiling through systematic toxicological studies, pharmacogenomic integration, and advanced modeling of compound–compound interactions [133], [134], [135].

Conclusions

MC, a traditional herbal medicine, contains bioactive compounds–particularly paeonol and paeoniflorin–that exhibit well-defined pharmacological effects and mechanisms, including anti-fibrotic, anti-inflammatory, and antioxidant activities. Bioactive components derived from MC epitomize the integration of phytotherapy and precision medicine, providing valuable insights for drug development and the global development of herbal products. Despite their promising potential as multi-target agents, the translation of paeonol and paeoniflorin faces several challenges. Key limitations include unstandardized experimental protocols, insufficient clinical evidence, and a lack of comprehensive safety assessments. To overcome these barriers, future research should adopt multidisciplinary strategies that incorporate the establishment of uniform experimental guidelines and data-sharing platforms, the implementation of rigorous large-scale clinical trials, and the integration of advanced technologies such as artificial intelligence and multi-omics. These approaches will help bridge the gap between traditional applications and evidence-based medicine, ultimately supporting the development of MC-derived compounds into widely accepted and therapeutically effective treatments.

Looking ahead, the successful translation of herbal-medicine-derived components will also depend on fostering collaboration between TCM researchers, pharmacologists, clinical investigators, and industry partners. This collaborative approach will not only drive the development of high-quality, evidence-based herbal products but also contribute to a more inclusive global healthcare landscape–one that values the contributions of herbal medicine while upholding the rigorous standards of modern science.

Contributorsʼ Statement

Wei Zheng conducted the literature collection and interpretation, drafted the initial manuscript, and led the manuscript revision. Yingting Li and Xingyi Wu assisted in literature collection and rechecked the revised manuscript. Luping Yuan conceived the research theme, reviewed and edited the final draft, and coordinated the overall revision process.

Conflict of Interest

The authors declare that they have no conflicts of interest.

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Publication History

Received: 25 February 2025

Accepted: 13 October 2025

Accepted Manuscript online:
13 October 2025

Article published online:
10 November 2025