The effect of mango aroma in low-sugar beverage: A sensory study on odor induced sweetness enhancement.

Excessive intake of sugar has become a public concern. However, it is challenging for food industries to decrease sugar level without sacrificing safety and sensory profile. Odor-induced sweetness enhancement (OISE) is believed to be a novel and promising strategy for sugar reduction. In order to investigate the OISE effect of mango aroma and evaluate its degree of sugar reduction in low-sugar beverages, a mathematical model was constructed through sensory evaluation in this study. The results showed that the maximum liking of low-sugar model beverages was 4.28 % sucrose and 0.57 % mango flavor. The most synergistic of OISE was at the concentration level of 2.24 % sucrose + 0.25 % mango flavor, which was equivalent to 2.96 % pure sucrose solution. With 32.14 % sugar reduction, the mango aroma was suggested to generate the OISE effect. However, the same level of garlic aroma was not able to enhance sweetness perception, suggesting that the congruency of aroma and taste is a prerequisite for the OISE effect to occur. This study demonstrated that the cross-modal interaction of mango aroma on sweetness enhancement in low-sugar model beverages could provide practical guidance for developing sugar-reduced beverages without applying sweeteners.

DUSP8-attenuated ERK1/2 signaling mediates lipogenesis and steroidogenesis in chicken granulosa cells.

The lipogenesis and steroidogenesis of granulosa cells are crucial during follicular development, yet it remains unclear whether dual-specificity phosphatase 8 (DUSP8) is involved. In this study, the specific role of DUSP8 in lipogenesis and steroidogenesis was investigated through culturing chicken granulosa cells in vitro. The results revealed that the expression levels of adipogenic genes were elevated after DUSP8 overexpression and reduced after knockdown. The same was observed for lipid deposition in granulosa cells. Meanwhile, the steroidogenic gene expression and progesterone synthesis were promoted after DUSP8 overexpression and inhibited after knockdown. In addition, we also found that DUSP8 blocked the phosphorylation of extracellular regulatory kinase 1/2 (ERK1/2). Based on the previous results that activated ERK1/2 signaling inhibited lipid deposition and progesterone synthesis in chicken granulosa cells, we demonstrated that DUSP8 promoted lipid deposition and progesterone synthesis through mediating the ERK1/2 signaling pathway. The results will improve our understanding of the molecular regulatory mechanisms regarding lipid metabolism and progesterone synthesis in chicken granulosa cells.

Profiling flavor characteristics of cold brew coffee with GC-MS, electronic nose and tongue: effect of roasting degrees and freeze-drying.

BACKGROUND: Roasting is an important process in the formation of coffee flavor characteristics, which determines the quality of coffee and consumer acceptance. However, the influence of roasting degree on the flavor characteristics of cold brew coffee has not been fully described. RESULTS: In the present study, the flavor characteristics of cold brew coffee with different roasting degrees were compared in detail by using chromatographic and electronic sensory approaches, and the flavor changes induced by freeze-drying were investigated. Pyrazine and heterocyclic compounds were the main aroma compounds in coffee, and gradually dominated with the increase of roasting. Pyridine was consistently present in cold brew coffees of different roasting degrees and showed significant gradient of quantity accumulation. Aroma compounds such as pyrazine, linalool and furfuryl acetate were the main contributors to coffee roasting, floral and fruity flavor. Freeze-drying preserved the fruity and floral aromas of medium-roasted cold brew coffee, whereas reducing the bitterness, astringency and acidity properties that are off-putting to consumers. CONCLUSION: The higher consumer acceptance and enjoyment in medium roast cold brew coffee may be related to its stronger floral and fruity aroma. The aroma profile qualities of freeze-drying processed medium roasted cold brewed coffee were more dominant and more suitable for freeze-drying processing than medium dark roasting. Application of freeze-drying for cold brew coffee will promote the convenience of drinking. The present study provides valuable technical guidance in improving the flavor and quality of cold brew coffee, and also promotes its commercialization process. © 2024 Society of Chemical Industry.

Insights into the mechanisms driving microbial community succession during pepper fermentation: Roles of microbial interactions and endogenous environmental changes.

Elucidating the driving mechanism of microbial community succession during pepper fermentation contributes to establishing efficient fermentation regulation strategies. This study utilized three-generation high-throughput sequencing technology, microbial co-occurrence network analysis, and random forest analysis to reveal microbial community succession processes and driving mechanisms during pepper fermentation. The results showed that more positive correlations than negative correlations were observed among microorganisms, with positive correlation proportions of 60 %, 51.03 %, and 71.43 % between bacteria and bacteria, fungi and fungi, and bacteria and fungi in sipingtou peppers, and 69.23 %, 54.93 %, and 79.44 % in zhudachang peppers, respectively. Microbial interactions, mainly among Weissella hellenica, Lactobacillus plantarum, Hanseniaspora opuntiae, and Kazachstania humillis, could drive bacterial and fungal community succession. Notably, the bacterial community successions during the fermentation of two peppers were similar, showing the transition from Leuconostoc pseudomesenteroides, Lactococcus lactis, Weissella ghanensis to Weissella hellenica and Lactobacillus plantarum. However, the fungal community successions in the two fermented peppers differed significantly, and the differential biomarkers were Dipodascus geotrichum and Kazachstania humillis. Differences in autochthonous microbial composition and inherent constituents brought by pepper varieties resulted in different endogenous environmental changes, mainly in fructose, malic acid, and citric acid. Furthermore, endogenous environmental factors could also drive microbial community succession, with succinic acid, lactic acid, and malic acid being the main potential drivers of bacterial community succession, whereas fructose, glucose, and succinic acid were the main drivers of fungal community succession. These results will provide insights into controlling fermentation processes by raw material combinations, optimization of environmental parameters, and microbial interactions.

Physiological role of dietary energy in the sexual maturity: clues of body size, gonad development, and serum biochemical parameters of Chinese indigenous chicken.

Sexual maturity is a crucial factor in the formation and development of poultry reproductive capacity. The nutritional status has been confirmed to play an important role in the regulation of sexual maturity. To investigate the effect of dietary energy levels on sexual maturity in chicken, diets with 3 energy levels (group L: 2,573 kcal/kg, group C: 2,836 kcal/kg, group H: 3,122 kcal/kg) were implemented to feed Guangyuan Gray chickens. During this trial, body weight, body size, organ development, sexual maturity, reproductive performance and blood biochemical parameters were monitored. The earlier sexual maturity was observed in group H, as well as a heavier first egg weight, larger interpubic distance and higher total cholesterol (T-CHO) content at sexual maturity. The dietary energy levels had no significant effect on body weight at first egg and egg production at 300 d of age. Although dietary energy levels had a significant effect on body weight, comb length, tibia length and girth, abdominal fat weight, oviduct weight and length, T-CHO, triglyceride (TG) content and estradiol (E2) level during the rearing period. No significant difference of gonadotropin releasing hormone (GnRH) and luteinizing hormone (LH) level among 3 groups was observed during the trial. The dietary energy levels had effects on mRNA expression of GnRH, estrogen receptor 1 (ESR1), estrogen receptor 2 (ESR2) in hypothalamus, gonadotropin inhibitory hormone receptor (GnIHR) in pituitary and luteinizing hormone receptor (LHR), ESR2 in ovary. The GnIHR/GnRHR ratio in pituitary was higher before sexual maturity and decreased at sexual maturity. The results of correlations analysis found that all the body size, carcass traits, serum biochemical parameters negatively correlated with age at first egg except for interpubic distance and serum blood glucose content. Collectively, dietary energy levels had effects on sexual maturity of chicken, which may be achieved by affecting body weight, gonad development, endocrine and the mRNA expression of genes related to hypothalamus-pituitary-gonad axis. These results further set our understanding of how dietary energy regulates sexual maturity.

A Comparison of the Meat Quality, Nutritional Composition, Carcass Traits, and Fiber Characteristics of Different Muscular Tissues between Aged Indigenous Chickens and Commercial Laying Hens.

The aim of this study is to assess the differences in the meat quality, nutritional composition, carcass traits, and myofiber characteristics between Hy-Line grey chickens (HLG, commercial breed) and Guangyuan grey chickens (GYG, indigenous breed). A total of 20 55-week-old chickens were selected for slaughter. The HLG exhibited a larger carcass weight, breast muscle weight, and abdominal fat weight (p < 0.05). The GYG exhibited a higher crude protein content, lower shear force, and smaller fiber size in the thigh muscles, whereas the HLG presented higher pH values and lower inosine-5'-monophosphate content in the breast muscles (p < 0.05). Darker meat based on higher redness and yellowness values was observed in the GYG instead of the HLG (p < 0.05). The research results also revealed parameter differences between different muscle types. Simultaneously, a correlation analysis showed significant correlations between the meat quality traits and myofiber characteristics (p < 0.05). In conclusion, aged indigenous chickens perform better in terms of tenderness and nutritional value in the thigh muscles, and may exhibit a better flavor in the breast muscles, but have a smaller breast muscle weight. Therefore, the current investigation provides a theoretical basis for the different needs of consumers and the processing of meat from old laying hens.

Medical visual question answering: A survey.

Medical Visual Question Answering (VQA) is a combination of medical artificial intelligence and popular VQA challenges. Given a medical image and a clinically relevant question in natural language, the medical VQA system is expected to predict a plausible and convincing answer. Although the general-domain VQA has been extensively studied, the medical VQA still needs specific investigation and exploration due to its task features. In the first part of this survey, we collect and discuss the publicly available medical VQA datasets up-to-date about the data source, data quantity, and task feature. In the second part, we review the approaches used in medical VQA tasks. We summarize and discuss their techniques, innovations, and potential improvements. In the last part, we analyze some medical-specific challenges for the field and discuss future research directions. Our goal is to provide comprehensive and helpful information for researchers interested in the medical visual question answering field and encourage them to conduct further research in this field.

Comparative Analysis of Different Proteins and Metabolites in the Liver and Ovary of Local Breeds of Chicken and Commercial Chickens in the Later Laying Period.

The liver and ovary perform a vital role in egg production in hens. In the later laying period, the egg-laying capacity of female hens, particularly that of local breeds, declines significantly. Hence, it is essential to study the features and conditions of the ovary and liver during this period. In this research, we characterized the proteins and metabolites in the liver and ovary of 55-week-old Guangyuan gray chickens (Group G) and Hy-Line gray chickens (Group H) by using liquid chromatography chip/electrospray ionization quadruple time-of-flight/mass spectroscopy (LC-MS/MS). In total, 139 differentially expressed proteins (DEPs) and 186 differential metabolites (DMs) were identified in the liver, and 139 DEPs and 36 DMs were identified in the ovary. The upregulated DEPs and DMs in both the liver and ovary of Group G were primarily enriched in pathways involved in amino acid and carbohydrate metabolism. This suggests that energy metabolism was highly active in the Guangyuan gray chickens. In contrast, the upregulated DEPs and DMs in Group H were mainly enriched in pathways associated with lipid metabolism, which may explain the higher egg production and the higher fatty liver rate in Hy-Line gray hens in the later laying period. Additionally, it was found that the unique protein s-(hydroxymethyl) glutathione dehydrogenase (ADH4) in Group G was implicated in functions such as fatty acid degradation, glycolysis, and pyruvate metabolism, whereas the unique proteins, steroid sulfatase (STS), glucosylceramidase (LOC107050229), and phospholipase A2 Group XV (PLA2G15), in Group H were involved in the metabolism of steroid hormones and glycerol phosphate. In conclusion, variations in how carbohydrates, lipids, and amino acids are processed in the liver and ovary of local breeds of chicken and commercial hens towards the end of their laying period could explain the disparities in their egg production abilities.

Enhancement effect of odor and multi-sensory superposition on sweetness.

The impact of sugary foods on public health has contributed to the development of low-sugar and sugar-substituted products, and sugar reduction has become a major challenge for the food industry. There is growing empirical evidence that odor can enhance the perception of sweetness without increasing the caloric load. This current review summarizes the researches on odor-induced sweetness enhancement published in recent years and discusses the mechanisms and influencing factors of odor-sweetness interactions. In addition, by combing existing studies, this paper also summarizes the research methods and strategies to investigate odor-induced sweetness enhancement. Finally, the feasibility of synergistic enhancement of sweetness through the superposition of odor with other senses (texture, visual, etc.) is also discussed and analyzed. In conclusion, odor-induced sweetness enhancement may present an alternative or complementary approach for developing foods with less sugar.

Lipid Deposition and Progesterone Synthesis Are Increased by miR-181b-5p through RAP1B/ERK1/2 Pathway in Chicken Granulosa Cells.

Steroid hormones secreted by granulosa cells are essential for maintaining normal development of chicken follicles. Our previous sequencing data indicated that miR-181b-5p and RAS-related protein 1B (RAP1B) appeared to function in chicken granulosa cells, which was further explored in this study. The results suggested that miR-181b-5p facilitated the aggregation of lipid droplets and the synthesis of progesterone. In contrast, RAP1B astricted lipid deposition and progesterone secretion. Cotransfection of the RAP1B overexpression vector with miR-181b-5p mimic eliminated the promoting effect of miR-181b-5p. Dual-luciferase reporter assay confirmed that miR-181b-5p bound directly to the 3' untranslated region (3' UTR) of RAP1B. We also found that miR-181b-5p and RAP1B reduced and enhanced the phosphorylation levels of extracellular signal-regulated kinases 1 and 2 (ERK1/2), respectively. The application of ERK1/2 activators and inhibitors demonstrated that ERK1/2 is a negative regulator of lipid deposition and progesterone synthesis. In conclusion, we revealed that miR-181b-5p accelerated lipid deposition and progesterone synthesis through the RAP1B/ERK1/2 pathway in chicken granulosa cells. miR-181b-5p and RAP1B may serve as new biomarkers in breeding to improve chicken reproductive performance and prevent ovary-related diseases.

The Implications in Meat Quality and Nutrition by Comparing the Metabolites of Pectoral Muscle between Adult Indigenous Chickens and Commercial Laying Hens.

Aged chickens are often a secondary dietary choice, owing to the poor organoleptic qualities of their meat. With regard to the meat quality of chickens, the metabolic profiles of pectoral muscle in Guangyuan grey chickens (group G) and Hy-Line grey hens (group H) aged 55 weeks were compared via ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). A total of 74 metabolites were identified with differential changes in the ion model. Lipids and lipid-like molecules comprised the largest proportion among the different metabolites. The content of myristic acid and palmitic acid were found to be higher in the pectoral muscle of group G, while group H showed significantly higher levels of glycerophospholipid molecules, such as LPC(18:2/0:0), Pi(38:5), Pc(16:0/16:0), and Pe(16:1e/14-hdohe). KEGG pathway analysis indicated that the abundant metabolites in group G were mainly involved in energy metabolism and fatty acid biosynthesis and metabolism, whereas those of group H were mainly attributed to the metabolism of unsaturated fatty acids and amino acids. Overall, the differences in lipid and amino acid metabolism in pectoral muscle appear to be responsible for the difference in meat quality between indigenous chickens and commercial laying hens.

Improving Cycle Life of Zinc-Air Batteries with Calcium Ion Additive in Electrolyte or Separator.

The electrolyte carbonation and the resulting air electrode plugging are the primary factors limiting the cycle life of aqueous alkaline zinc-air batteries (ZABs). In this work, calcium ion (Ca2+) additives were introduced into the electrolyte and the separator to resolve the above issues. Galvanostatic charge-discharge cycle tests were carried out to verify the effect of Ca2+ on electrolyte carbonation. With the modified electrolyte and separator, the cycle life of ZABs was improved by 22.2% and 24.7%, respectively. Ca2+ was introduced into the ZAB system to preferentially react with CO32- rather than K+ and then precipitated granular CaCO3 prior to K2CO3, which was deposited on the surface of the Zn anode and air cathode to form a flower-like CaCO3 layer, thereby prolonging its cycle life.

Correction to: Evaluation of salinity resistance and combining ability analysis in the seedlings of mulberry hybrids (Morus alba L.).

[This corrects the article DOI: 10.1007/s12298-023-01304-w.].

miR-136-5p/FZD4 axis is critical for Wnt signaling-mediated myogenesis and skeletal muscle regeneration.

Skeletal muscle can undergo a regenerative process in response to injury or disease to maintain muscle quality and function. Myogenesis depends on the proliferation and differentiation of myoblasts, and miRNAs can maintain the balance between them by precisely regulating many key factors in the myogenic network. Here, we found that miR-136-5p was significantly upregulated during the proliferation and differentiation of C2C12 cells. We demonstrate that miR-136-5p acts as a myogenic negative regulator during the development of mouse C2C12 myoblasts. In terms of mechanism, miR-136-5p inhibits the formation of ß-catenin/LEF/TCF DNA-binding factor transcriptional regulatory complex by targeting FZD4, a gating protein in the Wnt signaling pathway, thereby enhancing downstream myogenic factors and finally promoting myoblast proliferation and differentiation. In addition, in BaCl2 -induced muscle injury mouse model, miR-136-5p knockdown accelerated the regeneration of skeletal muscle after injury, and further led to the improvement of gastrocnemius muscle mass and muscle fiber diameter, while being suppressed by shFZD4 lentivirus infection. In summary, these results demonstrate the essential role of miR-136-5p/FZD4 axis in skeletal muscle regeneration. Given the conservation of miR-136-5p among species, miR-136-5p may be a new target for treating human skeletal muscle injury and improving the production of animal meat products.

Fixed-Time Control for a Flexible Smart Structure With Actuator Failure: A Broad Learning System Approach.

This article proposes an adaptive fault-tolerant control (AFTC) approach based on a fixed-time sliding mode for suppressing vibrations of an uncertain, stand-alone tall building-like structure (STABLS). The method incorporates adaptive improved radial basis function neural networks (RBFNNs) within the broad learning system (BLS) to estimate model uncertainty and uses an adaptive fixed-time sliding mode approach to mitigate the impact of actuator effectiveness failures. The key contribution of this article is its demonstration of theoretically and practically guaranteed fixed-time performance of the flexible structure against uncertainty and actuator effectiveness failures. Additionally, the method estimates the lower bound of actuator health when it is unknown. Simulation and experimental results confirm the efficacy of the proposed vibration suppression method.

Evaluation of salinity resistance and combining ability analysis in the seedlings of mulberry hybrids (Morus alba L.).

Soil salinization has become one of the major abiotic stresses influencing food security and maintenance of sustainable eco-environment. Highly salt-tolerant germplasm in mulberry, an important perennial woody plant, could restore the ecology and increase the agricultural income. Studies on the salt tolerance of mulberry are limited. Therefore, the aim of this study was to estimate the genetic variation and develop a reliable and effective evaluation of salt tolerance in 14 F1 mulberry hybrids that were directionally constructed using nine genotypes, including two females and seven males. A salt stress test was performed using 0.3%, 0.6%, and 0.9% (w/v) NaCl to investigate four morphological indexes of the growth rate: the shoot height (SHR), leaf number (LNR), leaf area (LAR), and the total weight of the whole plant after defoliation (BI) in the seedlings of the 14 combinations. The most suitable concentration for evaluating salt tolerance was identified as 0.9% NaCl based on the changes in the salt tolerance coefficient (STC). Comprehensive evaluation (D) values were obtained using principal components and membership functions based on four morphological indexes and their STCs, grouped into three principal component indexes cumulatively contributing to approximately 88.90% of the total variance. Two highly salt-tolerant, three moderately salt-tolerant, five salt-sensitive, and four highly salt-sensitive genotypes were screened. Anshen × Xinghainei and Anshen × Xinghaiwai had the highest D values. The analyses of combining ability further showed that the variances for LNR, LAR, and BI were elevated significantly with the increasing NaCl concentrations. Anshen × Xinghainei from two superior parents (female: Anshen, male: Xinghainei) with relatively higher general combing abilities for SHR, LAR, and BI was the best hybrid combination under high salinity stress, and presented the best specific combining ability for BI. Of all the traits tested, LAR and BI were greatly affected by additive effects and might be the two most reliable indexes. These traits show higher correlation with the salt tolerance of mulberry germplasm at the seedling stage. These results may enrich the mulberry resources by breeding and screening for elite germplasms with high salt tolerance. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01304-w.

Nutrition and Flavor Evaluation of Amino Acids in Guangyuan Grey Chicken of Different Ages, Genders and Meat Cuts.

The composition and content of amino acids in foodstuffs have a vital impact on the nutritional value and taste. With the aim of understanding the nutrition and flavor of Guangyuan grey chicken, the composition and content of amino acids in the pectoralis and thigh muscle of chickens at the age of 90 d, 120 d and 150 d were determine using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and an amino acid analyzer. A total of 17 amino acids were detected both in pectoralis and thigh muscle via the amino acid analyzer, of which the content of glutamate was the highest. Additionally, 21 deproteinized free amino acids were detected via LC-MS/MS. Among all samples, the content of glutamine in thigh muscle was the highest. The content of histidine in the pectoralis was the highest. In terms of the flavor amino acids (FAAs), the umami-taste and sweet-taste amino acids were higher in the thigh muscle of 120 d male chicken. From the perspective of protein nutrition, the essential amino acid was higher in pectoral muscle, and the composition was better. The results of the amino acid score showed that the content of leucine and valine were inadequate in Guangyuan grey chicken. Collectively, the content of amino acid in Guangyuan grey chicken was affected by age, gender and meat cut. This study confirms that meat of chicken in different ages, genders, and cuts presents different nutritional values and flavors owing to the variation of amino acids content.

Medical matting: Medical image segmentation with uncertainty from the matting perspective.

High-quality manual labeling of ambiguous and complex-shaped targets with binary masks can be challenging. The weakness of insufficient expression of binary masks is prominent in segmentation, especially in medical scenarios where blurring is prevalent. Thus, reaching a consensus among clinicians through binary masks is more difficult in multi-person labeling cases. These inconsistent or uncertain areas are related to the lesions' structure and may contain anatomical information conducive to providing an accurate diagnosis. However, recent research focuses on uncertainties of model training and data labeling. None of them has investigated the influence of the ambiguous nature of the lesion itself. Inspired by image matting, this paper introduces a soft mask called alpha matte to medical scenes. It can describe the lesions with more details better than a binary mask. Moreover, it can also be used as a new uncertainty quantification method to represent uncertain areas, filling the gap in research on the uncertainty of lesion structure. In this work, we introduce a multi-task framework to generate binary masks and alpha mattes, which outperforms all state-of-the-art matting algorithms compared. The uncertainty map is proposed to imitate the trimap in matting methods, which can highlight fuzzy areas and improve matting performance. We have created three medical datasets with alpha mattes to address the lack of available matting datasets in medical fields and evaluated the effectiveness of our proposed method on them comprehensively. Furthermore, experiments demonstrate that the alpha matte is a more effective labeling method than the binary mask from both qualitative and quantitative aspects.

Improving the Encapsulation Efficiency of Lipase in Molecular Cages and Its Application.

Here, lipase encapsulation is constructed by locking enzyme molecules in nanomolecular cages on the surface of SH-PEI@PVAC magnetic microspheres. To improve the encapsulation efficiency in enzyme loading, the thiol group is efficiently modified on the grafted polyethyleneimine (PEI) using 3-mercaptopropionic acid. N2 adsorption-desorption isotherms reveal the existence of mesoporous molecular cages on the microsphere surface. The robust immobilizing strength of carriers to lipase demonstrates the successful encapsulation of enzymes in nanomolecular cages. The encapsulated lipase shows high enzyme loading (52.9 mg/g) and high activity (51.4 U/mg). Different sizes of molecular cages are established, and the cage size showed important effects on lipase encapsulation. It shows that enzyme loading is low at a small size of molecular cages, which is attributed to that the nanomolecular cage is too small to house lipase. The investigation in lipase conformation suggests that the encapsulated lipase retains its active conformation. Compared with the adsorbed lipase, the encapsulated lipase shows higher thermal stability (4.9 times) and higher resistance to denaturants (5.0 times). Encouragingly, the encapsulated lipase shows high activity and reusability in lipase-catalyzed synthesis of propyl laurate, suggesting the potential application value of encapsulated lipase.

Co-Immobilization of GOD & HRP on Y-Shaped DNA Scaffold and the Regulation of Inter-Enzyme Distance.

In multienzymes cascade reaction, the inter-enzyme spacing is supposed to be a factor affecting the cascade activity. Here, a simple and efficient Y-shaped DNA scaffold is assembled using two partially complementary DNA single strands on magnetic microspheres, which is used to coimmobilize glucose oxidase (GOD) and horseradish peroxidase (HRP). As a result, on poly(vinyl acetate) magnetic microspheres (PVAC), GOD/HRP-DNA@PVAC multienzyme system is obtained, which can locate GOD and HRP accurately and control the inter-enzyme distance precisely. The distance between GOD and HRP is regulated by changing the length of DNA strand. It showed that the cascade activity is significantly distance-dependent. Moreover, the inter-enzyme spacing is not the closer the better, and too short distance would generate steric hindrance between enzymes. The cascade activity reached the maximum value of 967 U mg-1 at 13.6 nm, which is 3.5 times higher than that of free enzymes. This is ascribed to the formation of substrate channeling.