Immobilized enzyme-based enzyme electrode biosensors, characterized by high sensitivity and efficiency, strong specificity, and compact size, demonstrate broad application prospects in life science research, disease diagnosis and monitoring, etc. Immobilization of enzyme is a critical step in determining the performance (stability, sensitivity, and reproducibility) of the biosensors. Random immobilization (physical adsorption, covalent cross-linking, etc.) can easily bring about problems, such as decreased enzyme activity and relatively unstable immobilization. Whereas, directional immobilization utilizing amino acid residue mutation, affinity peptide fusion, or nucleotide-specific binding to restrict the orientation of the enzymes provides new possibilities to solve the problems caused by random immobilization. In this paper, the principles, advantages and disadvantages and the application progress of enzyme electrode biosensors of different directional immobilization strategies for enzyme molecular sensing elements by specific amino acids (lysine, histidine, cysteine, unnatural amino acid) with functional groups introduced based on site-specific mutation, affinity peptides (gold binding peptides, carbon binding peptides, carbohydrate binding domains) fused through genetic engineering, and specific binding between nucleotides and target enzymes (proteins) were reviewed, and the application fields, advantages and limitations of various immobilized enzyme interface characterization techniques were discussed, hoping to provide theoretical and technical guidance for the creation of high-performance enzyme sensing elements and the manufacture of enzyme electrode sensors.

This article systematically analyzes the historical evolution of the name， origin， medicinal parts， harvesting and processing， and other aspects of Manjiao and Zanthoxyli Radix by referring to the herbal medicine， medical books， prescription books and other documents of the past dynasties， combined with the relevant modern research materials， in order to provide a basis for the development of famous classical formulas containing the two medicinal materials. According to the herbal textual research， Manjiao was first recorded in Shennong Bencaojing of the Han dynasty with aliases such as Zhujiao， Goujiao and Zhijiao. Throughout history， Manjiao was sourced from the stems and roots of Zanthoxylum armatum in the Rutaceae family， and its leaves and fruits can also be used in medicine. The traditional recorded production area was mainly in Yunzhong（now Tuoketuo region in Inner Mongolia）， with mentions in Zhejiang， Hunan， Fujian， Guangdong， Guangxi， Yunnan， Taiwan， and other provinces. Presently， this species is distributed from the south of Shandong， to Hainan， Taiwan， Tibet and other regions. The roots can be harvested year-round， while the fruits are harvested in autumn after maturity. In ancient times， the roots and stems were mostly used for brewing or soaking in wine， whereas nowadays， the roots are often sliced and then used as a raw material in traditional Chinese medicine， and the fruits should be stir-fried before use. Manjiao has a bitter taste and warm property， and was historically used to treat wind-cold dampness， joint pain， limb numbness， and knee pain. Modern researches have summarized its effects as dispelling wind， dispersing cold， promoting circulation， and relieving pain， and it is used for treating rheumatoid arthritis， toothache， bruises， as well as an anthelmintic. Zanthoxyli Radix initially known as Rudi Jinniugen， recorded in Bencao Qiuyuan of the Qing dynasty， with the alternate name of Liangbianzhen. In recent times， it is more commonly referred to as Liangmianzhen， sourced from the dried roots of Z. nitidum of the Rutaceae family， mainly produced in Guangxi and Guangdong. It can be harvested throughout the year， cleaned， sliced， and dried after harvesting. Zanthoxyli Radix is pungent， bitter， warm and slightly toxic， with the functions of promoting blood circulation， removing stasis， relieving pain， dispelling wind， and resolving swelling. Based on the results of herbal textual research， it is clarified that the ancient Manjiao and the modern Zanthoxyli Radix are not the same species. This article corrects the mistaken belief of by previous scholars that Zanthoxyli Radix is the same as ancient Manjiao， and suggests that formulas described as Manjiao should use Z. armatum as the medicinal herb， while those described as Liangmianzhen or Rudi Jinniu should use Z. nitidum. The processing was performed according to the processing requirements prescribed in the formulas， otherwise， the raw products are recommended for use.

This article systematically analyzes the historical evolution of the name， origin， medicinal parts， harvesting and processing， and other aspects of Manjiao and Zanthoxyli Radix by referring to the herbal medicine， medical books， prescription books and other documents of the past dynasties， combined with the relevant modern research materials， in order to provide a basis for the development of famous classical formulas containing the two medicinal materials. According to the herbal textual research， Manjiao was first recorded in Shennong Bencaojing of the Han dynasty with aliases such as Zhujiao， Goujiao and Zhijiao. Throughout history， Manjiao was sourced from the stems and roots of Zanthoxylum armatum in the Rutaceae family， and its leaves and fruits can also be used in medicine. The traditional recorded production area was mainly in Yunzhong（now Tuoketuo region in Inner Mongolia）， with mentions in Zhejiang， Hunan， Fujian， Guangdong， Guangxi， Yunnan， Taiwan， and other provinces. Presently， this species is distributed from the south of Shandong， to Hainan， Taiwan， Tibet and other regions. The roots can be harvested year-round， while the fruits are harvested in autumn after maturity. In ancient times， the roots and stems were mostly used for brewing or soaking in wine， whereas nowadays， the roots are often sliced and then used as a raw material in traditional Chinese medicine， and the fruits should be stir-fried before use. Manjiao has a bitter taste and warm property， and was historically used to treat wind-cold dampness， joint pain， limb numbness， and knee pain. Modern researches have summarized its effects as dispelling wind， dispersing cold， promoting circulation， and relieving pain， and it is used for treating rheumatoid arthritis， toothache， bruises， as well as an anthelmintic. Zanthoxyli Radix initially known as Rudi Jinniugen， recorded in Bencao Qiuyuan of the Qing dynasty， with the alternate name of Liangbianzhen. In recent times， it is more commonly referred to as Liangmianzhen， sourced from the dried roots of Z. nitidum of the Rutaceae family， mainly produced in Guangxi and Guangdong. It can be harvested throughout the year， cleaned， sliced， and dried after harvesting. Zanthoxyli Radix is pungent， bitter， warm and slightly toxic， with the functions of promoting blood circulation， removing stasis， relieving pain， dispelling wind， and resolving swelling. Based on the results of herbal textual research， it is clarified that the ancient Manjiao and the modern Zanthoxyli Radix are not the same species. This article corrects the mistaken belief of by previous scholars that Zanthoxyli Radix is the same as ancient Manjiao， and suggests that formulas described as Manjiao should use Z. armatum as the medicinal herb， while those described as Liangmianzhen or Rudi Jinniu should use Z. nitidum. The processing was performed according to the processing requirements prescribed in the formulas， otherwise， the raw products are recommended for use.

This article systematically analyzes the historical evolution of the name， origin， academic name， medicinal parts， origin， harvesting， processing and other aspects of Abri Herba and Abri Mollis Herba by referring to the herbal medicine， medical books， prescription books and other documents of the past dynasties， combined with the modern literature， so as to provide a basis for the development of famous classical formulas containing this type of medicinal materials. According to the herbal textual research， Abri Herba was first recorded in Lingnan Caiyaolu， with other aliases such as Huangtoucao and Xiye Longlincao. It originates from the dried whole plant of Abrus cantoniensis， a Fabaceae plant， which can be used medicinally except for its fruits. Currently， this species is mainly distributed in Guangdong and Guangxi， and also found in Hunan and Thailand， it can be harvested throughout the year， mainly in spring and autumn. The roots， stems， and leaves can be used for medicinal purposes， but the pods are toxic and need to be removed. After harvesting， impurities and pods are removed， and it is dried and processed for medicinal use. Abri Herba has a sweet and slightly bitter taste， is cool in nature， and is associated with the liver and stomach meridians， it is used for clearing heat and relieving dampness， dispersing blood stasis and relieving pain， and is mainly used to treat jaundice-type hepatitis， stomach pain， rheumatic bone pain， contusion and ecchymosis pain， and mastitis. Abri Mollis Herba was first recorded in the 1982 edition of Zhongyaozhi as another origin for Abri Herba， and was singled out in some monographs such as Xinhua Bencao Gangyao in 1988 for use， while some other monographs use it as a local habitual products or confused products of Abri Herba with aliases such as Daye Jigucao， Qingtingteng， and Maoxiangsi. It comes from the dried whole herb of A. mollis without pods， and is mainly produced in Guangxi and Guangdong， and occasionally found in Hong Kong， Hainan and Fujian. The collection and processing are similar to Abri Herba， after harvesting， impurities and pods are removed， and it is dried and cut for medicinal use. Abri Mollis Herba has a sweet and light taste， is cool in nature， and is associated with the liver and stomach meridians， with the efficacy of clearing heat and detoxifying， and promoting dampness， it is mainly used to treat infectious hepatitis， mastitis， furuncles， burns and scalds， and pediatric malnutrition. Based on the research， A. mollis was first recorded to be used as a medicine in the same origin as A. cantoniensis， and as plants of the same genus， have similar morphological characteristics， and their medicinal parts， collection and processing， properties and flavors， and meridian affiliations are consistent. And in the folk， Abri Mollis Herba is often used as Abri Herba， which has been used for a long time and is now dominated by the cultivation of A. mollis. So it is recommended that the subsequent version of Chinese Pharmacopoeia should include A. mollis in the origin of Abri Herba， and it is also recommended that in famous classical formulas refered to Jiguccao can use A. cantoniensis and A. mollis as the sources of the herb， refered to Mao Jiguccao can use A. mollis as the sources of the herb. Processing is carried out according to the requirements specified in the original formulas， and raw products are recommended to be included in the medicine if there are no requirements.

This article systematically analyzes the historical evolution of the name， origin， academic name， medicinal parts， origin， harvesting， processing and other aspects of Abri Herba and Abri Mollis Herba by referring to the herbal medicine， medical books， prescription books and other documents of the past dynasties， combined with the modern literature， so as to provide a basis for the development of famous classical formulas containing this type of medicinal materials. According to the herbal textual research， Abri Herba was first recorded in Lingnan Caiyaolu， with other aliases such as Huangtoucao and Xiye Longlincao. It originates from the dried whole plant of Abrus cantoniensis， a Fabaceae plant， which can be used medicinally except for its fruits. Currently， this species is mainly distributed in Guangdong and Guangxi， and also found in Hunan and Thailand， it can be harvested throughout the year， mainly in spring and autumn. The roots， stems， and leaves can be used for medicinal purposes， but the pods are toxic and need to be removed. After harvesting， impurities and pods are removed， and it is dried and processed for medicinal use. Abri Herba has a sweet and slightly bitter taste， is cool in nature， and is associated with the liver and stomach meridians， it is used for clearing heat and relieving dampness， dispersing blood stasis and relieving pain， and is mainly used to treat jaundice-type hepatitis， stomach pain， rheumatic bone pain， contusion and ecchymosis pain， and mastitis. Abri Mollis Herba was first recorded in the 1982 edition of Zhongyaozhi as another origin for Abri Herba， and was singled out in some monographs such as Xinhua Bencao Gangyao in 1988 for use， while some other monographs use it as a local habitual products or confused products of Abri Herba with aliases such as Daye Jigucao， Qingtingteng， and Maoxiangsi. It comes from the dried whole herb of A. mollis without pods， and is mainly produced in Guangxi and Guangdong， and occasionally found in Hong Kong， Hainan and Fujian. The collection and processing are similar to Abri Herba， after harvesting， impurities and pods are removed， and it is dried and cut for medicinal use. Abri Mollis Herba has a sweet and light taste， is cool in nature， and is associated with the liver and stomach meridians， with the efficacy of clearing heat and detoxifying， and promoting dampness， it is mainly used to treat infectious hepatitis， mastitis， furuncles， burns and scalds， and pediatric malnutrition. Based on the research， A. mollis was first recorded to be used as a medicine in the same origin as A. cantoniensis， and as plants of the same genus， have similar morphological characteristics， and their medicinal parts， collection and processing， properties and flavors， and meridian affiliations are consistent. And in the folk， Abri Mollis Herba is often used as Abri Herba， which has been used for a long time and is now dominated by the cultivation of A. mollis. So it is recommended that the subsequent version of Chinese Pharmacopoeia should include A. mollis in the origin of Abri Herba， and it is also recommended that in famous classical formulas refered to Jiguccao can use A. cantoniensis and A. mollis as the sources of the herb， refered to Mao Jiguccao can use A. mollis as the sources of the herb. Processing is carried out according to the requirements specified in the original formulas， and raw products are recommended to be included in the medicine if there are no requirements.

T7 RNA polymerase (T7 RNAP) is one of the simplest known RNA polymerases. Its unique structural features make it a critical model for studying the mechanisms of RNA synthesis. This review systematically examines the static crystal structure of T7 RNAP, beginning with an in-depth examination of its characteristic “thumb”, “palm”, and “finger” domains, which form the classic “right-hand-like” architecture. By detailing these structural elements, this review establishes a foundation for understanding the overall organization of T7 RNAP. This review systematically maps the functional roles of secondary structural elements and their subdomains in transcriptional catalysis, progressively elucidating the fundamental relationships between structure and function. Further, the intrinsic flexibility of T7 RNAP and its applications in research are also discussed. Additionally, the review presents the structural diagrams of the enzyme at different stages of the transcription process, and through these diagrams, it provides a detailed description of the complete transcription process of T7 RNAP. By integrating structural dynamics and kinetics analyses, the review constructs a comprehensive framework that bridges static structure to dynamic processes. Despite its advantages, T7 RNAP has a notable limitation: it generates double-stranded RNA (dsRNA) as a byproduct. The presence of dsRNA not only compromises the purity of mRNA products but also elicits nonspecific immune responses, which pose significant challenges for biotechnological and therapeutic applications. The review provides a detailed exploration of the mechanisms underlying dsRNA formation during T7 RNAP catalysis, reviews current strategies to mitigate this issue, and highlights recent progress in the field. A key focus is the semi-rational design of T7 RNAP mutants engineered to minimize dsRNA generation and enhance catalytic performance. Beyond its role in transcription, T7 RNAP exhibits rapid development and extensive application in fields, including gene editing, biosensing, and mRNA vaccines. This review systematically examines the structure-function relationships of T7 RNAP, elucidates the mechanisms of dsRNA formation, and discusses engineering strategies to optimize its performance. It further explores the engineering optimization and functional expansion of T7 RNAP. Furthermore, this review also addresses the pressing issues that currently need resolution, discusses the major challenges in the practical application of T7 RNAP, and provides an outlook on potential future research directions. In summary, this review provides a comprehensive analysis of T7 RNAP, ranging from its structural architecture to cutting-edge applications. We systematically examine: (1) the characteristic right-hand domains (thumb, palm, fingers) that define its minimalistic structure; (2) the structure-function relationships underlying transcriptional catalysis; and (3) the dynamic transitions during the complete transcription cycle. While highlighting T7 RNAP’s versatility in gene editing, biosensing, and mRNA vaccine production, we critically address its major limitation—dsRNA byproduct formation—and evaluate engineering solutions including semi-rationally designed mutants. By synthesizing current knowledge and identifying key challenges, this work aims to provide novel insights for the development and application of T7 RNAP and to foster further thought and progress in related fields.

This study explores the basic characteristics and potential functions of the terpene synthase(TPS) gene family members in Lonicera japonica. The L. japonica TPS(LjTPS) gene family was identified and functionally analyzed using bioinformatics methods. The results showed that a total of 70 members of the LjTPS gene family were identified in L. japonica, with protein lengths ranging from 130 to 1 437 amino acids. Most of these proteins were hydrophilic, and they were unevenly distributed across nine chromosomes. Phylogenetic analysis showed that the LjTPS gene family members were divided into six subfamilies, mainly consisting of members from the TPS-a, TPS-b, and TPS-e subfamilies. Promoter cis-acting element analysis showed that LjTPS members contained a large number of stress-responsive cis-acting elements. Aphid inoculation experiments showed that key enzyme genes in the MVA pathway for terpenoid backbone synthesis in L. japonica, such as HMGS, HMGR, MK, MPD, and the key enzyme gene in the DXP pathway, DXS, exhibited an initial increase followed by a decrease under aphid stress. The qRT-PCR analysis showed that the expression levels of the α-farnesene synthase genes LjTPS34 and LjTPS39 were down-regulated, while the expression levels of(E)-β-caryophyllene synthase genes LjTPS15 and LjTPS17 were up-regulated 12 h before aphid feeding, then began to decline. Farnesyl pyrophosphate synthase(FPS), which interacted with these genes, also displayed a pattern of increasing followed by decreasing expression. The expression of linalool synthase genes LjTPS12 and LjTPS33 was significantly up-regulated after 72 h of aphid feeding(P<0.000 1), reaching 24.39 and 22.64 times the initial expression, respectively. This pattern was in close alignment with the trend of linalool content in L. japonica. This study provides a theoretical foundation for future research on the interaction between L. japonica and pests, as well as on the functional roles of the LjTPS gene family.
Animals ; Aphids/physiology* ; Alkyl and Aryl Transferases/chemistry* ; Lonicera/parasitology* ; Phylogeny ; Plant Proteins/chemistry* ; Gene Expression Regulation, Plant ; Multigene Family ; Terpenes/metabolism*

This study aims to explore the scientific connotation of sinking Rehmanniae Radix has the best quality and compare the quality between floating and sinking fresh Rehmanniae Radix samples. Ultra-performance liquid chromatography tandem quadrupole electrostatic field Orbitrap high-resolution mass spectrometry(UHPLC-Q-Orbitrap HRMS) was employed to detect the chemical components in floating and sinking fresh Rehmanniae Radix samples. The fingerprint of fresh Rehmanniae Radix was established by high performance liquid chromatography(HPLC), and four index components were determined simultaneously. The cluster analysis, principal component analysis(PCA), and orthogonal partial least squares-discriminant analysis(OPLS-DA) were conducted to compare the quality of floating and sinking fresh Rehmanniae Radix samples. An evaporative light-scattering detector was used to compare the content of five sugars. The extract yield and drying rate were determined, and the quality connotation of sinking Rehmanniae Radix has the best quality was explained by multiple indicators. A total of 41 components were preliminarily identified from fresh Rehmanniae Radix by UHPLC-Q-Orbitrap HRMS, including 7 iridoid glycosides, 9 phenylethanol glycosides, 6 amino acids, 4 sugars, 3 phenolic acids, 5 nucleosides, 3 organic acids, 1 ionone, 1 furan, 1 coumarin, and 1 phenylpropanoid. The results showed that the main chemical components were consistent between floating and sinking fresh Rehmanniae Radix. Nine common peaks were identified in the fingerprints of 15 batches of floating and sinking fresh Rehmanniae Radix samples, and the similarity of fingerprints was greater than 0.9. The cluster analysis, PCA, and OPLS-DA classified floating and sinking fresh Rehmanniae Radix sasmples into two categories, indicating differences in the quality between them. The total content of catalpol, rehmannioside D, ajugol, and verbascoside in sinking fresh Rehmanniae Radix samples was higher than that in floating samples of the same batch and specification, and the main differential component was catalpol. The total content of fructose, glucose, sucrose, raffinose, and stachyose in sinking fresh Rehmanniae Radix samples was higher than that in floating samples of the same batch and specification, and the main differential component was stachyose. The extract yield and drying rate of the sinking samples were higher than those of floating samples. This study preliminarily showed that floating and sinking fresh Rehmanniae Radix samples had the same components but great differences in the content of medicinal substance basis. The total content of four glycosides and five sugars, extract yield, and drying rate of sinking fresh Rehmanniae Radix samples is higher than that of floating samples of the same batch and specification. These findings, to a certain extent, explains the scientificity of sinking Rehmanniae Radix has the best quality recorded in ancient books and provide a reference for the quality control and clinical application of fresh Rehmanniae Radix.
Chromatography, High Pressure Liquid/methods* ; Drugs, Chinese Herbal/chemistry* ; Rehmannia/chemistry* ; Chemometrics ; Mass Spectrometry/methods* ; Quality Control ; Principal Component Analysis ; Plant Extracts

Diabetic foot (DF) is one of the most serious chronic complications of diabetes. The incidence rate among global diabetes patients is as high as 15% to 25%, and about 50% of patients will develop contralateral foot ulcers within 5 years after the first unilateral ulcer. As a non-invasive prevention and control solution, the application progress of functional insoles is mainly reflected in the following aspects:(1) Material innovation. The application of new composite materials and smart materials has significantly enhanced the pressure reduction effect and comfort. (2) Structural optimization. The development of multi-layer design and local pressure reduction structure has achieved more precise pressure distribution regulation. (3) Manufacturing process. 3D printing and parametric design have enabled the personalized customization of functional insoles. (4) Intelligent monitoring. It integrates functions such as pressure sensing and temperature monitoring, achieving real-time monitoring and early warning of foot conditions. Clinical research has confirmed that personalized functional insoles could reduce the incidence of foot ulcers and shorten the healing time of ulcers. At present, the research hotspots mainly focus on the development of smart materials, the construction of multi-functional integration and remote monitoring systems. However, in-depth research is still needed in the aspects of biomechanical mechanisms, standardized evaluation systems and long-term efficacy assessment. The development of future functional insoles should focus on the coordinated advancement of "personalization-intelligence-standardization", with the aim of providing more effective solutions for the prevention and treatment of DF.
Humans ; Diabetic Foot/therapy* ; Foot Orthoses

OBJECTIVES: To determine the pharmacological impact of hesperidin, the main component of Citri Reticulatae Pericarpium, on depressive behavior and elucidate the mechanism by which hesperidin treats depression, focusing on the gut-brain axis. METHODS: Fifty-four Sprague Dawley male rats were randomly allocated to 6 groups using a random number table, including control, model, hesperidin, probiotics, fluoxetine, and Citri Reticulatae Pericarpium groups. Except for the control group, rats in the remaining 5 groups were challenged with chronic unpredictable mild stress (CUMS) for 21 days and housed in single cages. The sucrose preference test (SPT), immobility time in the forced swim test (FST), and number in the open field test (OFT) were performed to measure the behavioral changes in the rats. Enzyme-linked immunosorbent assay was used to determine the levels of 5-hydroxytryptamine (5-HT) and brain-derived neurotrophic factor (BDNF) in brain tissue, and the histopathology was performed to evaluate the changes of colon tissue, together with sequencing of the V3-V4 regions of 16S rRNA gene on feces to explore the changes of intestinal flora in the rats. RESULTS: Compared to the control group, the rats in the model group showed notable reductions in body weight, SPF, and number in OFT (P<0.01). Hesperidin was found to ameliorate depression induced by CUMS, as seen by improvements in body weight, SPT, immobility time in FST, and number in OFT (P<0.05 or P<0.01). Regarding neurotransmitters, it was found that at a dose of 50 mg/kg hesperidin treatment upregulated the levels of 5-HT and BDNF in depressed rats (P<0.05). Compared to the control group, the colon tissue of the model group exhibited greater inflammatory cell infiltration, with markedly reduced numbers of goblet cells and crypts and were significantly improved following treatment with hesperidin. Simultaneously, the administration of hesperidin demonstrated a positive impact on the gut microbiome of rats treated with CUMS, such as Shannon index increased and Simpson index decreased (P<0.01), while the abundance of Pseudomonadota and Bacteroidota increased in the hesperidin-treated group (P<0.05). CONCLUSION The mechanism responsible for the beneficial effects of hesperidin on depressive behavior in rats may be related to inhibition of the expressions of BDNF and 5-HT and preservation of the gut microbiota.
Animals ; Hesperidin/therapeutic use* ; Rats, Sprague-Dawley ; Depression/drug therapy* ; Male ; Stress, Psychological/drug therapy* ; Brain/metabolism* ; Brain-Derived Neurotrophic Factor/metabolism* ; Serotonin/metabolism* ; Gastrointestinal Microbiome/drug effects* ; Behavior, Animal/drug effects* ; Rats ; Brain-Gut Axis/drug effects* ; Chronic Disease ; Colon/drug effects*