Gut Microbiome-Serum Metabolism Revealed the Allergenicity of Ferulic Acid Combined with Glucose-Modified ß-Lactoglobulin.

A novel strategy combining ferulic acid and glucose was proposed to reduce ß-lactoglobulin (BLG) allergenicity and investigate whether the reduction in allergenicity was associated with gut microbiome and serum metabolism. As a result, the multistructure of BLG changed, and the modified BLG decreased significantly the contents of IgE, IgG, IgG1, and mMCP-1 in serum, improved the diversity and structural composition of gut microbiota, and increased the content of short-chain fatty acids (SCFAs) in allergic mice. Meanwhile, allergic mice induced by BLG affected arachidonic acid, tryptophan, and other metabolic pathways in serum, the modified BLG inhibited the production of metabolites in arachidonic acid metabolism pathway and significantly increased tryptophan metabolites, and this contribution helps in reducing BLG allergenicity. Overall, reduced allergenicity of BLG after ferulic acid was combined with glucose modification by regulating gut microbiota, the metabolic pathways of arachidonic acid and tryptophan. The results may offer new thoughts alleviating the allergy risk of allergenic proteins.

Ferulic acid nanoemulsion as a promising anti-ulcer tool: in vitro and in vivo assessment.

OBJECTIVE: Ferulic acid (FA) is a promising nutraceutical molecule which exhibits antioxidant and anti-inflammatory properties, but it suffers from poor solubility and bioavailability. In the presented study, FA nanoemulsions were prepared to potentiate the therapeutic efficacy of FA in prevention of gastric ulcer. METHODS: FA nanoemulsions were prepared, pharmaceutically characterized, and the selected nanoemusion was tested for its ulcer-ameliorative properties in rats after induction of gastric ulcer using ethanol, by examination of stomach tissues, assessment of serum IL-1ß and TNF-α, assessment of nitric oxide, prostaglandin E2, glutathione, catalase and thiobarbituric acid reactive substance in stomach homogenates, as well as histological and immunohistochemical evaluation. RESULTS: Results revealed that the selected FA nanoemulsion showed a particle size of 90.43 nm, sustained release of FA for 8 h, and better in vitro anti-inflammatory properties than FA. Moreover, FA nanoemulsion exhibited significantly better anti-inflammatory and antioxidant properties in vivo, and the gastric tissue treated with FA nanoemulsion was comparable to the normal control upon histological and immunohistochemical evaluation. CONCLUSION: Findings suggest that the prepared ferulic acid nanoemulsion is an ideal anti-ulcer system, which is worthy of further investigations.

Effect of ferulic acid incorporation on structural, rheological, and digestive properties of hot-extrusion 3D-printed rice starch.

The influence of ferulic acid (FA) on rice starch was investigated by incorporating it at various concentrations (0, 2.5, 5, 7.5, and 10 %, w/w, on dry starch basis) and subjecting the resulting composites to hot-extrusion 3D printing (HE-3DP) process. This study examined the effects of FA addition and HE-3DP on the structural, rheological, and physicochemical properties as well as the printability and digestibility of rice starch. The results indicated that adding 0-5 % FA had no significant effect; however, as the amount of FA increased, the printed product edges became less defined, the product's overall stability decreased, and it collapsed. The addition of FA reduced the elasticity and viscosity, making it easier to extrude the composite gel from the nozzle. Moreover, the crystallinity and short-range ordered structure of the HE-3D printed rice starch gel decreased with the addition of FA, resulting in a decrease in the yield stress and an increase in fluidity. Furthermore, the addition of FA reduced the digestibility of the HE-3D-printed rice starch. The findings of this study may be useful for the development of healthier modified starch products by adding bioactive substances and employing the 3D printing technology.

Enhancing the bioavailability and antioxidant activity of natamycin E235-ferulic acid loaded polyethylene glycol/carboxy methyl cellulose films as anti-microbial packaging for food application.

This study investigated the development of biodegradable films made from a combination of polyethylene glycol (PEG), carboxymethyl cellulose (CMC) and mixtures from natamycin and ferulic acid. The films were characterized for their surface microstructure, antioxidant activity, thermal stability, mechanical properties, permeability and antifungal/bacterial activity. The addition of natamycin and ferulic acid to the film matrix enhanced antioxidant activity, thermal stability, antimicrobial activity, reduced the water vapor permeability (WVP) to 1.083 × 10-10 g × m-1s-1Pa-1, imparted opaque color and increased opacity up to 3.131 A mm-1. The attendance of natamycin and ferulic acid inside films created a clear roughness shape with agglomerates on the surface of films and caused a clear inhibition zone for Aspergillus niger, E. coli and C. botulinum. The utilization of PG/CMC/N-F packaging material on Ras cheese had a noticeable effect, resulting in a slight decrease in moisture content from 34.23 to 29.17 %. Additionally, it helped maintain the titrable acidity within the range of 0.99 % to 1.11 % and the force required for puncture from 0.035 to 0.052 N with non-significant differences. Importantly, these changes did not significantly affect the sensory qualities of Ras cheese during the storage period.

Browning reactions of hydroxycinnamic acids and heterocyclic Maillard reaction intermediates - Formation of phenol-containing colorants.

Non-enzymatic conversion of phenolic compounds plays an important role during thermal processing of plant-based food such as coffee, cocoa, and peanuts. However, the more prominent Maillard reaction is mainly studied at a mechanistic level for carbohydrates and amino compounds to clarify reactions that contribute to ('classic') melanoidin formation, but the role of phenolic compounds in such reactions is rarely discussed yet. To understand their contribution to non-enzymatic browning, reactions between ubiquitous phenolic acids, such as caffeic acid and ferulic acid, and prominent heterocyclic Maillard intermediates, namely furfural, hydroxymethylfurfural, and pyrrole-2-carbaldehyde were investigated. Following incubation under roasting conditions (220 °C, 0-30 min), heterogenous products were characterized by high-resolution mass spectrometry, and, after isolation, by nuclear magnetic resonance spectroscopy. By this, color precursors were identified, and it was shown that in addition to aromatic electrophilic substitution, nucleophilic and condensation reactions are key mechanisms contributing to the formation of phenol-containing melanoidins.

Repurposing of Antifungal Drug Flucytosine/Flucytosine Cocrystals for Anticancer Activity against Prostate Cancer Targeting Apoptosis and Inflammatory Signaling Pathways.

This study aimed to repurpose the antifungal drug flucytosine (FCN) for anticancer activity together with cocrystals of nutraceutical coformers sinapic acid (SNP) and syringic acid (SYA). The cocrystal screening experiments with SNP resulted in three cocrystal hydrate forms in which two are polymorphs, namely, FCN-SNP F-I and FCN-SNP F-II, and the third one with different stoichiometry in the asymmetric unit (1:2:1 ratio of FCN:SNP:H2O, FCN-SNP F-III). Cocrystallization with SYA resulted in two hydrated cocrystal polymorphs, namely, FCN-SYA F-I and FCN-SYA F-II. All the cocrystal polymorphs were obtained concomitantly during the slow evaporation method, and one of the polymorphs of each system was produced in bulk by the slurry method. The interaction energy and lattice energies of all cocrystal polymorphs were established using solid-state DFT calculations, and the outcomes correlated with the experimental results. Further, the in vitro cytotoxic activity of the cocrystals was determined against DU145 prostate cancer and the results showed that the FCN-based cocrystals (FCN-SNP F-III and FCN-SYA F-I) have excellent growth inhibitory activity at lower concentrations compared with parent FCN molecules. The prepared cocrystals induce apoptosis by generating oxidative stress and causing nuclear damage in prostate cancer cells. The Western blot analysis also depicted that the cocrystals downregulate the inflammatory markers such as NLRP3 and caspase-1 and upregulate the intrinsic apoptosis signaling pathway marker proteins, such as Bax, p53, and caspase-3. These findings suggest that the antifungal drug FCN can be repurposed for anticancer activity.

Improved antioxidant activity of pretreated lignin nanoparticles: Evaluation and self-assembly.

Lignin nanoparticles (LNPs) have gained significant attention for their potential as natural antioxidants. This study investigated the effect of various pretreatment methods on the lignin structure and subsequent antioxidant activity of LNPs. Among four pretreated LNPs, hydrothermal LNPs exhibited the highest antioxidant activity, surpassing unpretreated, acid-pretreated and kraft LNPs, with an impressive efficacy of 91.6%. The relationship between LNPs' structure and antioxidant activity was revealed by 2D heteronuclear singular quantum correlation (1H13C HSQC) and 31P nuclear magnetic resonance (NMR). 1H13C HSQC suggested the cleavage of ß-O-4 ether bonds, as well as a decrease in ferulic acid and p-coumaric acid, which directly influenced the antioxidant activity of LNPs. 31P NMR demonstrated a positive correlation between the total hydroxyl group content and the antioxidant activity. Besides, an isothermal kinetic model for scavenging free radicals was established based on Langmuir kinetic model instead of Freundlich model. Moreover, multilayer LNPs, based on layer-by-layer self-assembly, were prepared and exhibited remarkable antioxidant activity of 95.8%. More importantly, when blended with pure cosmetic cream, the multilayer LNPs maintained antioxidant activity of 86.7%. These finding may promote the practical applications of biomolecules, e.g. lignin additives in cosmetics and pharmaceuticals.

Affinity gel synthesis from the p-aminobenzoic acid derivative 4-amino-2-methylbenzoic acid and purification of polyphenol oxidase from various plant sources.

The polyphenol oxidase (PPO) enzyme, which causes enzymatic browning, has been repeatedly purified from fruit and vegetables by affinity chromatography. In the present research, Sepharose 4B-l-tyrosine-4-amino-2-methylbenzoic acid, a novel affinity gel for the purification of the PPO enzyme with high efficiency, was synthesized. Additionally, Sepharose 4B-l-tyrosine-p-aminobenzoic acid affinity gel, known in the literature, was also synthesized, and 9.02, 16.57, and 28.13 purification folds were obtained for the PPO enzymes of potato, mushroom, and eggplant by the reference gel. The PPO enzymes of potato, mushroom, and eggplant were purified 41.17, 64.47, and 56.78-fold from the new 4-amino-2-methylbenzoic acid gel. Following their isolation from the new affinity column, the assessment of PPO enzyme purity involved the utilization of SDS-PAGE. According to the results from SDS-PAGE and native PAGE, the molecular weight of each enzyme was 50 kDa. Then, the inhibition effects of naringin, morin hydrate, esculin hydrate, homovanillic acid, vanillic acid, phloridzin dihydrate, and p-coumaric acid phenolic compounds on purified potato, mushroom, and eggplant PPO enzyme were investigated. Among the tested phenolic compounds, morin hydrate was determined to be the most potent inhibitor on the potato (Ki: 0.07 ± 0.03 µM), mushroom (Ki: 0.7 ± 0.3 µM), and eggplant (Ki: 4.8 ± 1.2 µM) PPO enzymes. The studies found that the weakest inhibitor was homovanillic acid for the potato (Ki: 1112 ± 324 µM), mushroom (Ki: 567 ± 81 µM), and eggplant (Ki: 2016.7 ± 805.6 µM) PPO enzymes. Kinetic assays indicated that morin hydrate was a remarkable inhibitor on PPO.

Ferulic acid's therapeutic odyssey: nano formulations, pre-clinical investigations, and patent perspective.

INTRODUCTION: Ferulic acid (FA) is a phenolic phytochemical that has garnered the attention of the research community due to its abundant availability in nature. It is a compound that has been explored for its multifaceted therapeutic potential and benefits in modern and contemporary healthcare. AREAS COVERED: This review furnishes a compilation of the molecular mechanisms underlying the anti-diabetic, anticancer, antioxidant, and anti-inflammatory effects of FA. We also aim to excavate an in-depth analysis of the role of nanoformulations to achieve release control, reduce toxicity, and deliver FA at specified target sites. To corroborate the safety and efficacy of FA, a multitude of pre-clinical studies have also been conducted by researchers and have been discussed comprehensively in this review. The various patented innovations and newer paradigms pertaining to FA have also been presented. EXPERT OPINION: Enormous research has been conducted and should still be continued to find the best possible novel drug delivery system for FA delivery. The utilization of nanocarriers and nanoformulations has intrigued the scientists for delivery of FA, but before that, it is necessary to shed light upon toxicity, safety, and regulatory concerns of FA.

In vitro Evaluation of the Antileishmanial and Antischistosomal Activities of p-Coumaric Acid Prenylated Derivatives.

We have evaluated eight p-coumaric acid prenylated derivatives in vitro for their antileishmanial activity against Leishmania amazonensis promastigotes and their antischistosomal activity against Schistosoma mansoni adult worms. Compound 7 ((E)-3,4-diprenyl-4-isoprenyloxycinnamic alcohol) was the most active against L. amazonensis (IC50=45.92 µM) and S. mansoni (IC50=64.25 µM). Data indicated that the number of prenyl groups, the presence of hydroxyl at C9, and a single bond between C7 and C8 are important structural features for the antileishmanial activity of p-coumaric acid prenylated derivatives.

Dynamic changes in ferulic acid and diferulic acids in wheat flour doughs during the breadmaking process.

Ferulic acid (FA), a phytochemical concentrated in wheat bran, influences structural characteristics of arabinoxylan (AX) and rheological properties of wheat dough. This study investigates the dynamic changes in FA and diferulic acids, closely associated with AX molecular weight, during the breadmaking process. FA predominantly exists in a tightly bound state within the arabinoxylan matrix, with a substantial increase in free FA content observed during the initial fermentation phase. Furthermore, this research identified four specific wheat-derived diferulic acids: 8-5'-DFA, 5-5'-DFA, 8-O-4'-DFA, and 8-5'-DFA (benzofuran form), tracking their variations throughout breadmaking. The notable upsurge in diferulic acid levels in the early fermentation stages suggests that the cleavage of ferulic acid moieties may not be the primary factor contributing to the reduction in AX molecular weight. Future investigations into the effects of FA and diferulic acids on arabinoxylan and wheat dough properties promise to enhance understanding of the intricacies of the breadmaking process.

P-coumaric Acid: Advances in Pharmacological Research Based on Oxidative Stress.

P-coumaric acid is an important phenolic compound that is mainly found in fruits, vegetables, grains, and fungi and is also abundant in Chinese herbal medicines. In this review, the pharmacological research progress of p-coumaric acid in recent years was reviewed, with emphasis on its role and mechanism in oxidative stress-related diseases, such as inflammation, cardiovascular diseases, diabetes, and nervous system diseases. Studies have shown that p-coumaric acid has a positive effect on the prevention and treatment of these diseases by inhibiting oxidative stress. In addition, p-coumaric acid also has anti-tumor, antibacterial, anti-aging skin and other pharmacological effects. This review will provide reference and inspiration for further research on the pharmacological effects of p-coumaric acid.

In silico Evaluation of Ferulic Acid Based Multifunctional Conjugates as Potential Drug Candidates.

BACKGROUND: Recent research has shown that ferulic acid (FA, trans-4-hydroxy-3- methoxycinnamic acid) has remarkable antioxidant properties and a wide range of biological activities. Conjugation of two or more biologically active compounds to produce a novel molecular scaffold is justified by the need to enhance biological activity against a single target or obtain a conjugate that behaves as a multi-target-directed ligand. In addition, the conjugation strategy decreases dose-dependent side effects by promoting the use of smaller doses of conjugated components to treat the disease. Moreover, the patient's compliance is positively affected when conjugating two active compounds into a single more active compound as this reduces the number of pills to be taken daily. OBJECTIVE: This study aims to shed light on studies that design and synthesize FA-based hybrid compounds with enhanced biological activities and to in silico assess these compounds as potential drug candidates. METHODS: The conjugate compounds were found by searching the literature using the keywords (ferulic acid-based hybrid or ferulic acid-based conjugate). To study conjugate pharmacokinetic parameters and toxicity (ADMET), software suites from Biovia Inc. (San Diego, California) were integrated into Discovery Studio 4.5. The structures were created using ChemDraw Ultra 7.0. RESULTS: 14 conjugates exhibiting variable biological activities were collected and three of them (compounds 3,5, and 6) in addition to the cis FA (compound 12) are the best-predicted compounds with low Daphnia toxicity and hepatotoxicity with acceptable pharmacokinetic properties. CONCLUSION: Cis FA, FA conjugates 3,5, and 6 act as good drug candidates that can be used to modify new hits.

Insight into structure-activity relationships of hydroxycinnamic acids modified porous starch: The effect of phenolic hydroxy groups.

Antioxidant capacity of hydroxycinnamic acids-modified starch mainly depends on their chemical structure. Herein, cinnamic acid as well as meta-substituted and para-substituted cinnamic acid were selected for esterification with porous starch (labelled as CA@PS, m-CA@PS and p-CA@PS), with the successful formation of porous starch (labelled as PS) esters then confirmed by 1H NMR, 13C solid-state NMR and FT-IR spectroscopy. Three PS esters with almost same degrees of substitution (DS) were obtained, and antioxidant assays, including DPPH radical scavenging, reducing power and hydroxyl radical scavenging tests, were subsequently used to evaluate the antioxidant activity of the esterified PS. Overall, CA@PS showed weak antioxidant activity because of the absence of phenolic hydroxy, while p-CA@PS displayed better antioxidant capacity. Because its conjugated structure offered the stronger electron-donating effect, that could enhance antioxidant capacity. Therefore, antioxidant capacity depended significantly on overall chemical structure, including numbers and substitution positions of phenolic hydroxy groups.

Spectrophotometric and Fluorimetric High-Throughput Assays for Phenolic Acid Decarboxylase.

Biocatalytic decarboxylation of hydroxycinnamic acids yields phenolic styrenes, which are important precursors for antioxidants, epoxy coatings, adhesives and other polymeric materials. Bacillus subtilis decarboxylase (BsPAD) is a cofactor-independent enzyme that catalyzes the cleavage of carbon dioxide from p-coumaric-, caffeic-, and ferulic acid with high catalytic efficiency. Real-time spectroscopic assays for decarboxylase reactions remove the necessity of extensive sample workup, which is required for HPLC, mass spectrometry, gas chromatography, or NMR methods. This work presents two robust and sensitive assays based on photometry and fluorimetry that allow decarboxylation reactions to be followed with high sensitivity while avoiding product extraction and long analysis times. Optimized assay procedures were used to measure BsPAD activity in cell lysates and to determine the kinetic constants (KM and Vmax ) of the purified enzyme for p-coumaric-, caffeic- and ferulic acid. Substrate inhibition was shown for caffeic acid.

Ultrasonication significantly enhances grafting efficiency of chitosan-ferulic acid conjugate and improves its film properties under Fenton system.

Ultrasonication (US)-assisted Fenton-system (US-Fenton) with different US time was developed for synthesizing chitosan (CS)-ferulic acid (FA) conjugates. The optimal US-Fenton for a suitable time was selected for preparing a film with CS-FA conjugate and its structural, functional, rheological, and physical properties were also investigated. Compared with Fenton-system, US-Fenton enhanced the grafting ratio of the conjugates, which increased firstly and then decreased as US time. The conjugate obtained by US-Fenton for 1 min (FUS1) possessed the highest grafting ratio (121.28 mg FA/g) and its grafting time was also shortened from 12 h to 1 min contrasted with Fenton grafted method. Structural characterization results showed that FA was conjugated on CS via ester and amide bonds with decreased crystallinity. Scanning electron microscopy and molecular weight analysis indicated that the degradation degree of CS-FA conjugates increased with US time. The DPPH and ABTS radical-scavenging activities of FUS1 were the closest to ascorbic acid, and it also showed the best antibacterial effect among the test conjugates. Accordingly, FUS1 was selected to obtain the film for contrasting with CS film. FUS1 film solution exhibited a decreased viscosity. In comparison to CS film, UV transmittance of FUS1 film approached zero, and its moisture, oxygen, and carbon dioxide permeabilities significantly decreased (P < 0.05). Moreover, its water solubility and tensile strength increased by 58.09% and 25.72% than those of CS film, respectively. Therefore, US-Fenton for 1 min could be a promising method for efficiently preparing active food package materials and FUS1 film possessed broad application prospects.

The impact of processing on the release and antioxidant capacity of ferulic acid from wheat: A systematic review.

The antioxidant capacity and bioaccessibility of ferulic acid (FA)1 in wheat are highly limited by the lack of free ferulic acid (FFA).2 However, many studies claim that wheat processing can efficiently increase FFA content and ultimately influence the overall antioxidant capacity. Hence, this systematic review investigated changes in FFA content, antioxidant capacity and bioaccessibility of wheat after different processing treatments. A literature search of two databases (PubMed and Web of Science) was undertaken covering the last 20 years, yielding 1148 articles. Studies which employed bioprocessing, thermal processing and milling of wheat were considered. After exclusion criteria were applied, 36 articles were included. These covered single processing methods (n = 25, bioprocessing: n = 9, thermal processing: n = 9, milling n = 7) and combined processing methods (n = 11, bioprocessing & thermal processing = 7, bioprocessing, thermal processing & milling = 2, thermal processing & milling = 2). The total ferulic acid (TFA)3 content, degree of covalent bond hydrolysis and the percentage of FFA degraded or transformed to other compounds dominated the final changes in FFA content, antioxidant capacity and bioaccessibility. This systematic review is the first to comprehensively summarize the best efficient processing method for releasing FA and increasing antioxidant capacity and or bioaccessibility in wheat. The combination of particle size reduction, pre-hydrolysis thermal processing (except at high temperature and extended duration) and enzymatic hydrolysis (ferulic acid esterase (FAE)4 or fermentation) has the highest potential of releasing FA. However, the literature on the bioaccessibility of FA in wheat is limited and more work is required to demonstrate the link between the release of FA by processing and the consequent health benefits.

Fungal feruloyl esterases can catalyze release of diferulic acids from complex arabinoxylan.

Feruloyl esterases (FAEs, EC 3.1.1.73) catalyze the hydrolytic cleavage of ester bonds between feruloyl and arabinosyl moieties in arabinoxylans. Recently, we discovered that two bacterial FAEs could catalyze release of diferulic acids (diFAs) from highly substituted, cross-linked corn bran arabinoxylan. Here, we show that several fungal FAEs, notably AnFae1 (Aspergillus niger), AoFae1 (A. oryzae), and MgFae1 (Magnaporthe oryzae (also known as M. grisae)) also catalyze liberation of diFAs from complex arabinoxylan. By comparing the enzyme kinetics of diFA release to feruloyl esterase activity of the enzymes on methyl- and arabinosyl-ferulate substrates we demonstrate that the diFA release activity cannot be predicted from the activity of the enzymes on these synthetic substrates. A detailed structure-function analysis, based on AlphaFold2 modeled enzyme structures and docking with the relevant di-feruloyl ligands, reveal how distinct differences in the active site topology and surroundings may explain the diFA releasing action of the enzymes. Interestingly, the analysis also unveils that the carbohydrate binding module of the MgFae1 may play a key role in the diFA releasing ability of this enzyme. The findings contribute further understanding of the function of FAEs in the deconstruction of complex arabinoxylans and provide new opportunities for enzyme assisted upgrading of complex bran arabinoxylans.

Thermal-assisted synthesis of ferulic acid-chitosan complex in water and its application as safe antioxidant.

Safe antioxidants are highly demanded in food preservation, yet existing preparation methods of typical bio-based antioxidants all suffer from either toxic catalysts or poor water solubility of the products. Herein, a water-soluble safe antioxidant, ferulic acid-chitosan complex, was facilely prepared in water with the assistance of mild-temperature heating. The chemical structure of ferulic acid-chitosan complex was determined by spectroscopy, and its thermal stability and rheological properties were studied in detail. Different from its precursors, the ferulic acid-chitosan complex exhibits much improved water solubility, thanks to its ionic structure. The as-prepared chitosan-ferulic acid complex displays higher antioxidative property than free ferulic acid, which was illustrated by the good preservation of freshly prepared apple juice. Such thermal-assisted synthesis strategy is demonstrated as an effective approach to prepare hydrophilic chitosan complex bearing hydrophobic organic acid, which enables great feasibility to the development of chitosan-based functional biomaterials.

Developing CHCA/PPD as a novel matrix for enhanced matrix-assisted laser desorption/ionization-mass spectrometry imaging for analysis of antibiotics in grass carp tissues.

Antibiotics have important medical value, but they need to be monitored when used as veterinary drugs. We report α-cyano-4-hydroxycinnamic acid/p-phenylenediamine (CHCA/PPD) hybrid as a novel matrix for enhanced matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) in situ the spatial distribution of antibiotic drugs in grass carp tissues. METHOD: We have used MALDI-TOF-MSI in positive ion mode for the analysis of quinolones and sulfonamides in grass carps. A novel CHCA/PPD matrix was prepared and applied using a simple method to improve the analysis. RESULTS: Compared with the traditional matrix, CHCA/PPD significantly improved the detection intensity of quinolones and sulfonamides with better sensitivity (17.20%-94.30%) and reproducibility. For demonstration, this novel matrix was successfully applied to visualize enrofloxacin (ENR) in grass carp tissues, with the entire abundance differences clearly observed based on MALDI-MSI. The concentration levels in different tissues were determined, with a calibration curve of 10-2000 µg/ml (R2 > 0.993). CONCLUSION: This study was the first to introduce CHCA/PPD as a novel matrix, and the classical acid-base mixing was used to improve the ionization effect of the traditional matrix CHCA in MALDI. Based on CHCA/PPD, MALDI-MSI detected ENR in different grass carp tissues for the first time and realized the spatial distribution and concentration detection.