The walnut cracking process is the most critical and delicate step for achieving high-quality kernels. The traditional method for cracking (manually) is labor-intensive, time-consuming, and tedious. The existing cracking approaches are low production efficiency and serious walnut kernel breakage. Increasing cracking efficiency with minimum kernel breakage has been a challenging issue in the preliminary processing of walnuts. Therefore, this study develops an innovative walnut cracker with self-grading and multi-station extrusion, combined with theoretical investigation and experiment verification. First, a statistical analysis of walnut physical properties was conducted, including dimensions, shell thickness as well as shape characteristics. The mechanical properties of walnut cracking were examined by a series of experiments. Based on mechanical theory, a grading mechanism was designed for preliminary processing before walnut cracking. Then a shaftless screw conveying mechanism and an extrusion cracking mechanism were developed. To evaluate the cracker\'s performance, a comprehensive examination was carried out. The experiments yielded impressive results, with a grading rate of 87.3%, a shell-breaking rate of 91.50%, and a kernel-exposed rate of 84.72%. These outcomes signify a substantial improvement in production efficiency while minimizing kernel breakage. The developed walnut cracker plays a crucial role in walnut processing and kernel extraction, thereby elevating economic value. PRACTICAL APPLICATION: A self-grading multi-station extrusion walnut cracker is developed, which includes a grading mechanism with a shaftless screw conveyor and a grid-type trommel screen for conveying and classifying walnuts. This cracker can adapt to different walnut varieties by changing the gap-adjusting guide to control the breaking gap. Compared to similar extrusion-type walnut crackers, the developed cracker not only incorporates preliminary classification but also exhibits superior performance. HIGHLIGHTS: A novel multi-station extrusion mechanism for walnuts cracking is developed. The cracker can accommodate various walnut sizes for self-grading and screening. The design with semi-arc plates converts extrusion force into alternating stress. The shell-breaking rate and kernel-exposed rate achieves 91.50% and 84.72%.

This study focuses on changes in the physiochemical properties of chitosan film when incorporated with a blend of essential oils of Tulsi and Ajwain. The essential oil blend-loaded films showed a decrement in transparency. Tulsi essential oil decreased the moisture content, swelling capacity, and water solubility. However, adding Ajwain along with Tulsi essential oil led to a significant increase in these properties. Meanwhile, the water vapor transmission rate didn\'t change significantly due to non-polar constituents in Tulsi essential oil, except when only Ajwain essential oil was present. The mechanical properties showed that the tensile strength of films increased with the addition of Tulsi essential oil (14.95 MPa to 31.27 MPa) but decreased further with increasing Ajwain oil concentration in films (32.13 MPa to 15.89 MPa). On the other hand, an increment in percent elongation at break (8.26 % to 24.02 %) was observed due to the excellent plasticization effect of Ajwain essential oil. Antioxidant activity was observed for the Tulsi essential oil-containing films and increased significantly with adding Ajwain essential oil. Finally, walnuts were packed in the active film. The active film showed better antioxidant activity against the oxidation of oil in walnuts, which the FTIR of the packed product confirmed.

Pollination is the first step in the plant\'s fruit development. Therefore, fruit setting does not occur without pollination. Some problems encountered in natural pollination cause pollination not to be achieved as desired and cause significant losses in yield and fruit quality. Artificial pollination applications with drones are the best way to solve these problems. In this study, the AirPoll artificial pollination machine, which performs artificial pollination through the air using drone technology, was developed and the operating success of the machine was tested in walnut gardens. In the experiment gardens, female flowers on 18 branches of 5 trees each in the artificially pollinated area with a drone and in the control area were marked with colored strings. Control trees were selected from a distance that would not be possible to transport pollen with a drone. As a result of the study carried out in 2020 and 2021, the average fruit setting rate in trees pollinated by drone was determined as 94.61%. In control trees, 32.33% fruit setting was achieved. Thus, it was determined that the productivity increase in artificial pollination with AirPoll was 62.28%. In addition, in the study, Computational Fluid Dynamics (CFD) simulation analysis was performed using ANSYS Fluent 2024 R1 software to predict the downward air flow and pollen distribution in the walnut tree crown. The analysis was carried out in 680 iterations using drone propellers at a rotation speed of 4500 rpm, 4 m/s airflow and a k-w viscous model. In the analysis, it was observed that the pollen was distributed homogeneously with the determined height and the created artificial pollination environment. Based on the results obtained from the simulations, a convergence criterion of 5e-3 for continuity and 1e-6 for speed, k, w was determined. Considering all the results, the ease of use of the developed AirPoll artificial pollination machine and the successful results obtained in field trials reveal the effectiveness of the AirPoll artificial pollination machine.

Polyphenol oxidase (PPO) activity drives walnut fruit browning, but the roles of its only two-family genes, JrPPO1 and JrPPO2, remain unclear. This study explores the spatiotemporal expression and enzymatic characteristics of JrPPO1 and JrPPO2 in walnut. Treatment with the PPO activator CuSO4 and H2O2 accelerated fruit browning and up-regulated JrPPO1/2 expression, whereas treatment with the PPO inhibitor ascorbic acid delayed browning, down-regulating JrPPO1 and up-regulating JrPPO2 expression. Compared to mJrPPO1, mJrPPO2 can exhibited better enzyme activity at higher temperatures (47 °C) and in more acidic environments (pH 4.25). mJrPPO2 exhibited a higher substrate specificity over mJrPPO1, and the preferred substrates are catechol, chlorogenic acid, and epicatechin. Additionally, mJrPPO2 adapted better to low concentration of oxygen (as low as 1.0% O2) and slightly elevated CO2 levels compared to mJrPPO1. Subcellular localization and spatiotemporal expression patterns showed that JrPPO1 is only expressed in green tissues and located in chloroplasts, while JrPPO2 is also located in chloroplasts, partly associated with membranes, and is expressed in both green and non-green tissues. Silencing JrPPO1/2 with virus-induced gene silencing (VIGS) reduced fruit browning, maintained higher total phenols, and decreased MDA production. Notably, silencing JrPPO1 had a greater impact on browning than JrPPO2, indicating JrPPO1\'s greater contribution to PPO activity and fruit browning in walnut fruits. Consequently, JrPPO1 can be effectively regulated both at the molecular level and by manipulating environmental conditions, to achieve the objective of controlling fruit browning.

Walnuts (Juglans regia L.), renowned for their nutritional potency, are a rich source of unsaturated fatty acids. Their regular intake plays a pivotal role in health maintenance and recuperation from a myriad of ailments. Fatty acyl-acyl carrier protein thioesterases, which orchestrate the hydrolysis of acyl-ACP thioester bonds, thereby yielding fatty acids of varying chain lengths, are instrumental in augmenting plant fatty acid content and modulating the balance between saturated and unsaturated fatty acids. Despite some investigative efforts into the synthesis and metabolic pathways of fatty acids in walnuts, our comprehension of Fat in walnuts remains rudimentary. This research undertook a comprehensive characterization of the JrFat family, predicated on the complete genome sequence of walnuts, leading to the identification of 8 JrFat genes and an exploration of their protein physicochemical properties. Utilizing Arabidopsis and soybean Fat genes as outgroups, JrFat genes can be categorized into 5 distinct subgroups, three of which encompass a pair of homologous gene pairs. These genes have demonstrated remarkable conservation throughout the evolutionary process, with highly analogous conserved base sequences. The promoter region of JrFats genes predominantly harbors light response and plant hormone response regulatory elements, with no discernible disparity in promoter elements among different JrFats. Predictive analyses indicate that JrFats proteins engage extensively with walnut fatty acid synthesis and metabolism-associated proteins. qRT-PCR analysis reveals an initial surge in the expression of JrFats during the development of walnut kernels, which either stabilizes or diminishes following the hard core period. Homologous gene pairs exhibit analogous expression patterns, and the expression trajectory of JrFats aligns with the dynamic accumulation of fatty acids in kernels. The expression of JrFatA2 exhibits a strong correlation with the content of Alpha-linolenic acid, while the expression of JrFatB2 is inversely correlated with the content of two saturated fatty acids. Collectively, these findings enrich our understanding of fatty acid synthesis and metabolism in walnuts and furnish gene resources for enhancing the content and ratio of fatty acids in walnuts.

There has been limited research on external browning (EB) of walnut. This work discovered 1888 metabolites and 34 anthocyanins in walnut pellicles (WPs) after three drying methods using widely-targeted and anthocyanin-targeted metabolomics. Based on OPLS-DA and correlation analysis, 64 temperature-responsive metabolites (TRMs; 13 anthocyanins and 51 flavonoids) were identified as critical components in relation to EB. Notably, 14 flavonoids exhibited a strong positive correlation (r > 0.9) with the browning index (BI), with upregulation of >60% after browning. Most of the identified anthocyanins were negatively linked with BI because of degradation (>45%), with correlation coefficients ranging from 0.75 to 0.97. Furthermore, anthocyanidin reductase and laccase were the two key enzymes involved in the EB of WPs, with their activities increasing by 10.57-fold and 1.32-fold, respectively, with increasing drying temperature. A metabolic pathway network of the TRM was built to provide insights into the potential mechanisms underlying EB in WPs.

Non-steroidal anti-inflammatory drugs (NSAIDs), the most highly prescribed drugs in the world for the treatment of pain, inflammation, and fever, cause gastric mucosal damage, including ulcers, directly or indirectly, by which the development of GI-safer (-sparing) NSAIDs relates to unmet medical needs. This study aimed to document the preventive effects of walnut polyphenol extracts (WPEs) against NSAID-induced gastric damage along with the molecular mechanisms. RGM-1 gastric mucosal cells were administered with indomethacin, and the expressions of the inflammatory mediators between indomethacin alone or a combination with WPEs were compared. The expressions of the inflammatory mediators, including COX-1 and COX-2, prostaglandin E2, 15-hydroxyprostaglandin dehydrogenase (15-PGDH), and antioxidant capacity, were analyzed by Western blot analysis, RT-PCR, and ELISA, respectively. HO-1, Nrf-2, and keap1 were investigated. The in vivo animal models were followed with in vitro investigations. The NSAIDs increased the expression of COX-2 and decreased COX-1 and 15-PGDH, but the WPEs significantly attenuated the NSAID-induced COX-2 expression. Interestingly, the WPEs induced the expression of 15-PGDH. By using the deletion constructs of the 15-PGDH promoter, we found that c-Jun is the most essential determinant of the WPE-induced up-regulation of 15-PGDH expression. We confirmed that the knockdown of c-Jun abolished the ability of the WPEs to up-regulate the 15-PGDH expression. In addition, the WPEs significantly increased the HO-1 expression. The WPEs increased the nuclear translocation of Nrf2 by Keap-1 degradation, and silencing Nrf2 markedly reduced the WPE-induced HO-1 expression. We found that the WPE-induced HO-1 up-regulation was attenuated in the cells harboring the mutant Keap1, in which the cysteine 151 residue was replaced by serine. These in vitro findings were exactly validated in indomethacin-induced gastric rat models. Daily walnut intake can be a promising nutritional supplement providing potent anti-inflammatory, antioxidative, and mucosa-protective effects against NSAID-induced GI damage.

BACKGROUND: Walnut anthracnose caused by Colletotrichum gloeosporioides seriously endangers the yield and quality of walnut, and has now become a catastrophic disease in the walnut industry. Therefore, understanding both pathogen invasion mechanisms and host response processes is crucial to defense against C. gloeosporioides infection.
RESULTS: Here, we investigated the mechanisms of interaction between walnut fruits (anthracnose-resistant F26 fruit bracts and anthracnose-susceptible F423 fruit bracts) and C. gloeosporioides at three infection time points (24hpi, 48hpi, and 72hpi) using a high-resolution time series dual transcriptomic analysis, characterizing the arms race between walnut and C. gloeosporioides. A total of 20,780 and 6670 differentially expressed genes (DEGs) were identified in walnut and C. gloeosporioides against 24hpi, respectively. Generous DEGs in walnut exhibited opposite expression patterns between F26 and F423, which indicated that different resistant materials exhibited different transcriptional responses to C. gloeosporioides during the infection process. KEGG functional enrichment analysis indicated that F26 displayed a broader response to C. gloeosporioides than F423. Meanwhile, the functional analysis of the C. gloeosporioides transcriptome was conducted and found that PHI, SignalP, CAZy, TCDB genes, the Fungal Zn (2)-Cys (6) binuclear cluster domain (PF00172.19) and the Cytochrome P450 (PF00067.23) were largely prominent in F26 fruit. These results suggested that C. gloeosporioides secreted some type of effector proteins in walnut fruit and appeared a different behavior based on the developmental stage of the walnut.
CONCLUSIONS: Our present results shed light on the arms race process by which C. gloeosporioides attacked host and walnut against pathogen infection, laying the foundation for the green prevention of walnut anthracnose.

Tree nut consumption has been widely associated with various health benefits, with walnuts, in particular, being linked with improved cardiovascular and neurological health. These benefits have been attributed to walnuts\' vast array of phenolic antioxidants and abundant polyunsaturated fatty acids. However, recent studies have revealed unexpected clinical outcomes related to walnut consumption, which cannot be explained simply with the aforementioned molecular hallmarks. With the goal of discovering potential molecular sources of these unexplained clinical outcomes, an exploratory untargeted metabolomics analysis of the isolated walnut pellicle was conducted. This analysis revealed a myriad of unusual lipids, including oxylipins and endocannabinoids. These lipid classes, which are likely present in the pellicle to enhance the seeds\' defenses due to their antimicrobial properties, also have known potent bioactivities as mammalian signaling molecules and homeostatic regulators. Given the potential value of this tissue for human health, with respect to its \"bioactive\" lipid fraction, we sought to quantify the amounts of these compounds in pellicle-enriched waste by-products of mechanized walnut processing in California. An impressive repertoire of these compounds was revealed in these matrices, and in notably significant concentrations. This discovery establishes these low-value agriculture wastes promising candidates for valorization and translation into high-value, health-promoting products; as these molecules represent a potential explanation for the unexpected clinical outcomes of walnut consumption. This \"hidden quality\" of the walnut pellicle may encourage further consumption of walnuts, and walnut industries may benefit from a revaluation of abundant pellicle-enriched waste streams, leading to increased sustainability and profitability through waste upcycling.

Introduction: MYC transcription factors are the basic regulators of the jasmonic acid signaling pathway and play important roles in plant growth and development and the response to adverse stress. In recent years, severe winter freezing and late spring frost in the main planting area of walnut in Xinjiang have affected the growth and development of walnut, which has become a prominent problem restricting walnut production. Xinjiang wild walnut is the only remaining wild species of walnuts in China, which contains a lot of genes with excellent traits, and is important for the cultivation and breeding. Methods: In this paper, the physicochemical properties and bioinformatics of MYC transcription factor members in walnut were analyzed, and the nine MYC were screened from the transcriptome data under low temperature stress. At last, we study the subcellular localizations and the expression patterns of the nine MYC members in Xinjiang wild walnut. Results: The results revealed that 30 MYC members were identified from published walnut whole-genome data, and their evolutionary relationships with Arabidopsis and poplar were divided into six groups according to clustering analysis, among which JrMYC22 and JrMYC23 had high homology with PtrMYC2b, which is induced by jasmonic acid in response to low-temperature stress. Walnut MYC members are unevenly distributed on 12 chromosomes. The prediction of promoter cis-acting elements of walnut MYC transcription factor family members revealed that cis-acting elements related to jasmonic acid and lowtemperature stress were the ones with the greatest number of members, with 12. In addition, all nine MYC family members in Xinjiang wild walnut plants responding to low-temperature stress exhibited strong fluorescence responses in the nucleus. The expression levels of these members in response to low-temperature stress revealed that JrMYC28, JrMYC31, JrMYC33, JrMYC34, and JrMYC35 were highly expressed, and it was hypothesized that JrMYC28, JrMYC31, JrMYC33, JrMYC34, and JrMYC35 might play a key role in the response to lowtemperature stress. Discussion: The results of this study provide a theoretical basis for further research on the functional mechanisms of the MYC transcription factor family members in walnut.