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Broccoli rapidly yellows and dehydrates post-harvest, causing a short storage life. Although arginine post-harvest has been shown to enhance broccoli's storage quality and shelf life, it remains unclear whether pre-harvest treatment yields similar benefits. The results showed that pre-harvest spraying of 5 mM arginine reduces the respiration rate and delays electrolyte leakage during broccoli's storage period, minimizing weight loss and yellowing and chlorophyll degradation. It also inhibits the degradation of ascorbic acid (ASA) and promotes the synthesis of total phenols, thereby increasing the total antioxidant capacity. Further studies showed that pre-harvest arginine spraying enhanced the activity of arginine decarboxylase (ADC), ornithine decarboxylase (ODC), and nitric oxide synthase (NOS). This accelerated the nitric oxide (NO) and the polyamine (PA) accumulation. Additionally, arginine aids in reactive oxygen species (ROS) balance and wound healing. Notably, pre-harvest arginine spraying was found to inhibit the hydrogen peroxide (H 2 O 2) accumulation and enhance the activities of superoxide dismutase (SOD) and catalase (CAT), while also activating the phenylalanine ammonia lyase (PAL) and cinnamic acid 4-hydroxylase (C4H) activities. In conclusion, pre-harvest arginine spraying extends the broccoli's storage life and improves its storage quality by promoting endogenous arginine catabolism, ROS metabolism, and the wounds healing. Therefore, this study establishes a theoretical foundation for pre-harvest handling in post-harvest broccoli storage. • The effect of pre-harvest arginine spraying on broccoli storage was studied. • Exogenous arginine promoted endogenous arginine degradation in broccoli. • Arginine spraying produced more NO in broccoli during storage. • Increased NO regulated ROS metabolism and promoted wound healing. • Pre-harvest arginine spraying was more effective than post-harvest spraying. [ABSTRACT FROM AUTHOR]

• Transcriptomics revealed the wound-healing mechanism of broccoli stems during SAS. • The promoted PPP produced large amounts of NADPH. • NADPH enhancement activated the shikimate and phenylpropane pathway. • The wound-healing process of broccoli stems was promoted by the enhancement of NADPH. • PPP is the crucial driving force for wound healing in cut harvest broccoli stems. Wound healing is a crucial metabolic process that occurs in fruit and vegetables within a short time after harvest, especially in cut-harvest ones. To reveal wound healing mechanisms in cut-harvest broccoli stems during the self-adaptive stage (SAS, 12 h after harvest), tissue samples from the stems at the incision were analyzed at the transcriptome level. With the extension of time, the expression levels of six differentially expressed genes (DEGs) in the pentose phosphate pathway (PPP) significantly increased in the stems after cut-harvest. Among the genes, the expression levels of PGLS and PGD , which are directly involved in nicotinamide adenine dinucleotide phosphate (NADPH) generation, were 2.62 and 5.82 times higher in the 12 h vs 0 h comparison group, respectively. The shikimate and phenylpropanoid pathways were significantly activated in the broccoli stems during SAS. Genes involved in NADPH degradation through the aforementioned two pathways, including DHQ-SDH1 (106323466), aroC1 (106295042), and PAL3 (106300639), were significantly upregulated. These findings certified that wound healing of cut-harvest broccoli stems was promoted by enhancing NADPH levels. Interestingly, glycolysis (EMP) was a non-significant change during SAS after cut-harvest that contributed to maintaining organic matter by suppressing respiration and acclimatizing the postharvest broccoli to become an independent unit. These results indicate that the focus should be on NADPH levels for promoting wound healing of fruit and vegetables during storage. [Display omitted] [ABSTRACT FROM AUTHOR]

Energy status is a vital factor in the physiology of postharvest vegetables. An integration profiling of transcriptome and metabolome was used to examine the effects of hydrogen peroxide treatment on carbohydrate and energy metabolism in broccoli at 20 ℃ for 4 d. The results showed that hydrogen peroxide (H 2 O 2) treatment maintained higher levels of ATP and energy charge during storage. The integration profiling of transcriptome and metabolome revealed that H 2 O 2 treatment promoted the degradation of 1,3-β-glucan to D-glucose and inhibited sucrose, D-raffinose, maltose, and cellulose degradation, thereby maintaining a long-term and stable supply of glucose for glycolysis (EMP) and pentose phosphate pathway (PPP)·H 2 O 2 treatment enhanced the EMP and PPP by upregulating the expression of related genes. Enhancing the tricarboxylic acid cycle and inhibiting alcohol fermentation in H 2 O 2 treatment promoted energy production. Therefore, the long-term and stable energy supply might be crucial for the delayed senescence of postharvest broccoli via H 2 O 2 treatment. [Display omitted] • H 2 O 2 treatment maintains a long-term and stable supply of glucose in broccoli. • H 2 O 2 treatment enhances the glycolysis and oxidative phosphorylation. • H 2 O 2 treatment enhances the TCA cycle and inhibits alcohol fermentation. • Stable energy supply is key reason for the delayed senescence of broccoli by H 2 O 2. [ABSTRACT FROM AUTHOR]

An integrated transcriptome and metabolome profiling was used to investigate how exogenous H2O2 affects amino acid metabolism and its underlying regulatory mechanism involved in post-harvest broccoli’s yellowing and senescence processes. This approach found 86 differentially expressed genes (DEGs) and 28 differentially accumulated metabolites (DAMs) associated with amino acid metabolism during broccoli storage in the Control, H2O2, and DPI (diphenylene iodonium, an H2O2 inhibitor) treatments on 2 or 4 d. Furthermore, H2O2 treatment promoted the biosynthesis and accumulation of amino acids (e.g., tryptophan, valine, isoleucine, arginine, lysine, histidine, and glutamate) while inhibiting the degradation of seven amino acids (e.g., tyrosine, valine, leucine, isoleucine, β-alanine, glutamate, and proline) into tricarboxylic acid (TCA) cycle precursor (pyruvate) and intermediates (fumarate, acetyl-CoA, and 2-oxoglutarate). This inhibition alleviated metabolic disorders and ammonium-induced toxicity of post-harvest broccoli. Additionally, H2O2 treatment promoted the NH3 conversion into glutamine and the urea cycle, further mitigating ammonium-induced toxicity. H2O2 treatment also down-regulated the expressions of S-adenosylmethionine synthases (SAM), 1-aminocyclopropane-1-carboxylate synthases (ACS), and 1-aminocyclopropane-1-carboxylate oxidases (ACO), which reduced the S-adenosyl-L-methionine content, thereby inhibiting ethylene biosynthesis from methionine. Therefore, this study provides insights into the role and potential underlying regulatory mechanism of H2O2 on amino acid metabolism during broccoli senescence, thus offering a theoretical basis for delaying senescence in post-harvest horticultural products by regulating their amino acid metabolism. [ABSTRACT FROM AUTHOR]

To investigate the effects and potential mechanism of exogenous H 2 O 2 on antioxidant metabolism and yellowing in postharvest broccoli, the transcriptome and metabolome integrated analysis were performed. A total of 61 differentially expressed genes and 6 differentially accumulated metabolites identified among of H 2 O 2 and diphenylene iodonium (DPI) treatments on 2 and 4 d. Exogenous H 2 O 2 promoted the accumulation of endogenous H 2 O 2 within 1 d by activating RBOHs expressions·H 2 O 2 treatment up-regulated the expressions of SOD , CAT and peroxiredoxin (Prx) genes and APX , DHAR and GPX genes involved in ascorbic acid-glutathione (AsA-GSH) cycle, thereby promoting the scavenging of O 2 -. and H 2 O 2 and alleviating the oxidative damage. Additionally, H 2 O 2 treatment promoted the biosynthesis of AsA and GSH, and increased the content of AsA, GSH, oxidized glutathione, and γ-glutamylcysteine·H 2 O 2 treatment also promoted glucosinolate biosynthesis, the accumulation of 3-(6′-methylthio)hexylmalic acid, and the production of sulforaphane by up-regulating MYS expression, thereby enhancing the health-care function of postharvest broccoli. This study provided the new insights into regulatory mechanism of H 2 O 2 on antioxidant system. [Display omitted] • Multi-omics revealed the regulatory mechanism of H 2 O 2 on antioxidant metabolism. • Exogenous H 2 O 2 promoted H 2 O 2 accumulation of broccoli in 1 d by activating RBOHs. • H 2 O 2 up-regulated SOD , CAT and Prx expression and enhanced the AsA-GSH cycle. • H 2 O 2 promoted the biosynthesis and accumulation of AsA and GSH. • H 2 O 2 promoted sulforaphane production by up-regulating MYS , and GSL biosynthesis. [ABSTRACT FROM AUTHOR]

To understand the effects and related molecular regulation mechanism of H 2 O 2 on nucleic acid metabolism in yellowing and senescence process of postharvest broccoli, an integration profiling of transcriptome and metabolome was performed. Results showed that 2 mM H 2 O 2 delayed the yellowing and increase of relative conductivity, whereas 3 μM diphenylene iodonium (DPI, an inhibitor of H 2 O 2 production) accelerated these processes·H 2 O 2 treatment promoted DNA replication and repair by up-regulating replication protein A , DNA polymerase , and DNA ligase 1 expressions and increasing the essential building blocks of DNA (e.g., cytosine, guanosine 5′-diphosphate, cytidine, and adenosine) contents, thereby ensuring the integrity of genome in broccoli·H 2 O 2 treatment promoted transcription and pre-mRNA processing by up-regulating the expressions of RNA polymerases and spliceosome-related genes, which promoted the biosynthesis of mature mRNA·H 2 O 2 treatment also up-regulated PABPs expression, and thus to promote translation process in broccoli. Additionally, H 2 O 2 promoted the recycle of mRNA and accumulation of adenosine, which was beneficial to timely change own gene profile in response to complex postharvest environmental changes and alleviate the energy deficiency suffered by postharvest broccoli. These results suggested that H 2 O 2 is involved in the positive regulation of nucleic acid metabolism and maintained higher contents of nucleic acid-related metabolites, DNA and RNA, ultimately delaying the senescence of postharvest broccoli, which provided new insight for the regulatory mechanism of the yellowing and senescence in broccoli. [Display omitted] • Multi-omics were used to reveal the mechanism of H 2 O 2 on nucleic acid metabolism. • H 2 O 2 delayed the degradation of 10 nucleic acid-related metabolites in broccoli. • H 2 O 2 enhanced the high-fidelity and repair capacity of DNA replication. • H 2 O 2 promoted siRNA biosynthesis by activating RNA polymerase IV and V expressions. • H 2 O 2 promoted the translation by activating BABP and ribonucleoprotein expressions. [ABSTRACT FROM AUTHOR]

Nucleic acids (DNA and RNA) play crucial roles in guiding protein synthesis and plant resistance to biotic and abiotic stresses, but our current available understanding concerning on the role and function of nucleic acid metabolism, transcription and translation processes in postharvest fruit and vegetables remains largely unclear. In this study, an iTRAQ-based proteomics was used to investigate changes of protein abundance in postharvest broccoli under 40% O 2 + 5% CO 2 , 20% O 2 + 5% CO 2 and 5% O 2 + 5% CO 2 treatments. A total of 128 differentially expressed proteins (fold change ≥ 1.2) were identified. High O 2 stress reduced the DNA level by down-regulating the expression of proteins involved in DNA replication and up-regulating the expression of proteins DNA degradation. The decrease of DNA content, inhibition of transcription and promotion of RNA degradation induced by high O 2 stress further led to the mass loss of RNA content in postharvest broccoli. Additionally, high O 2 stress suppressed RNA processing, transport and translation and activated the expression of 20 S and 26 S proteasome, thereby accelerating the decrease of protein levels in postharvest broccoli. The decrease of DNA and RNA content might be related to the early yellowing and senescence in postharvest broccoli. This study revealed that the effect and mechanism of high O 2 stress on DNA and RNA metabolism, transcription and translation processes, and the role of these metabolic pathway on senescence in postharvest broccoli. [Display omitted] • Proteomic revealed the effects of high O 2 stress on DNA, RNA and protein metabolism. • High O 2 stress accelerated the degradation of DNA and RNA. • High O 2 stress decreased protein by activating 20 S and 26 S proteasomes activities. • The decrease of DNA and RNA might be the key reason for broccoli senescence. [ABSTRACT FROM AUTHOR]

Yellowing is always accompanied by chlorophyll degradation and carotenoid accumulation, which is an important indicator for evaluating the quality and commercial value of postharvest broccoli. Elevated CO 2 treatment was applied in broccoli heads to investigate its effect and mechanism on pigment metabolism. The results demonstrated that 30% CO 2 treatment maintained higher nutritional quality by increasing phenols and flavonoids contents and delayed the yellowing by alleviating chlorophyll loss and carotenoid accumulation. The chlorophyll molecule could also be continuously synthesized in postharvest broccoli. The up-regulation of chlorophyll synthesis genes (BoMCase , BoChlM , BoChlE , BoPOR , BoDVR and BoChlG) and chlorophyll combination genes (BoCAB1 and BoCAB2) and the down-regulation of chlorophyll degradation genes (BoSGR1 , BoNYC1 , BoPPH , BoPAO and BoRCCR) were observed in postharvest broccoli treated with 30% CO 2. Additionally, 30% CO 2 treatment down-regulated the expression of BoPSY , BoLCYB , BoCHYB and BoVDE , thereby inhibiting zeaxanthin and β-carotene biosynthesis. This study provided new insights into chlorophyll biosynthesis after harvest and the regulation of elevated CO 2 on pigment metabolism in postharvest broccoli. • This study revealed the effect of CO 2 treatment on yellowing of postharvest broccoli. • Proper CO 2 delayed chlorophyll degradation by suppressing chlorophyll catabolism. • The expression of chlorophyll anabolic genes and BoCAB was induced by 30% CO 2. • 30% CO 2 inhibited the accumulation of zeaxanthin and β-carotene. • Broccoli yellowing is due to the of chlorophyll decrease and carotenoid increase. [ABSTRACT FROM AUTHOR]

• Proteomic analysis was used to reveal the senescence mechanism under elevated O 2. • A total of 54 proteins related to broccoli senescence were identified and classified. • Elevated O 2 up-regulated the EMP, the TCA cycle, and OXPHOS to produce more ATP. • Elevated O 2 destroyed the vesicle transport system of synthetic proteins and lipids. • Energy waste is responsible for rapid broccoli senescence under high O 2 stress. Ripening and senescence of horticultural products involve in different metabolic processes, but the understanding of these processes at the proteomics level remains unknown. In this study, a proteomic technique was used for investigating the expression changes of proteins in broccoli heads under 5% O 2 + 5% CO 2 , 20% O 2 + 5% CO 2 and 40% O 2 + 5% CO 2 treatments. A total of 54 differentially expressed proteins (fold change ≥1.5) associating with ripening and senescence were identified. The results indicated that high O 2 treatment promoted the degradation of carbohydrate, lipid, amino acid and protein, thereby providing more substrates for energy production, in turn, accelerated the biosynthesis of lipid, amino acid, and protein. However, high O 2 treatment disrupted vesicle and transmembrane transport and inhibited the transport of these metabolites to specific organelles, thereby resulting in energy waste in postharvest broccoli heads. Therefore, we concluded that energy waste caused by the damage of transmembrane transport might be responsible for broccoli head ripening and senescence under high O 2 stress. In summary, this study revealed a comprehensive and interacting metabolic network of postharvest broccoli head ripening and senescence under high O 2 stress, which could be used for the development of postharvest preservation technologies to control ripening and senescence of horticultural products. [Display omitted] [ABSTRACT FROM AUTHOR]

[Display omitted] • Amino acid metabolism in broccoli was revealed by proteomics under high O 2 stress. • High O 2 stress induced amino acid degradation to support the TCA cycle. • High O 2 stress disrupted the urea cycle , thereby aggravating ammonium toxicity. • Amino acids, the precursors of glucosinolate, were reduced under high O 2 stress. • High O 2 stress induced ethylene anabolism and methionine catabolism. To better understand the global changes of amino acid catabolism and anabolism in broccoli in response to high O 2 stress, iTRAQ-based proteomics combined with amino acid analysis was used to investigate the broccoli proteome at 0 and 4 d after treatment with different O 2 concentrations (5% O 2 + 5% CO 2 , 20% O 2 + 5% CO 2 and 40% O 2 + 5% CO 2) at 20℃. A total of 106 proteins with changes ≥ 1.2-fold in abundance were observed. Amino acid anabolism was significantly suppressed by high O 2 stress, while catabolism was enhanced. High O 2 stress-induced amino acid metabolism promoted the conversion of Met to ethylene and the degradation of amino acids to intermediate metabolites of the TCA cycle, thereby suppressing glucosinolate biosynthesis. However, the up-regulation of arginase and urease induced by high O 2 stress aggravated ammonium toxicity. These findings enhance our understanding of high O 2 stress-induced amino acid metabolism, as well as the effects of amino acid metabolism on broccoli senescence. [ABSTRACT FROM AUTHOR]

• Proteomic analysis was used to reveal the yellowing mechanism under elevated O 2. • 189 proteins related to broccoli yellowing were identified and classified. • Elevated O 2 might damage the thylakoid structure and thus to promote yellowing. • Elevated O 2 destroyed the photosystem and led to chlorophyll photobleaching. • The oxidative damage of chloroplast was aggravated by high O 2 stress. Controlled atmosphere storage has been proved to be an effective measure to regulate chlorophyll degradation. The obvious inhibitory effect of CO 2 on chlorophyll degradation was widely studied, but information available on the promotion of chlorophyll degradation by elevated O 2 was limited. The present work aimed to investigate the regulation of different O 2 concentrations on yellowing of broccoli heads during storage by physiology and proteomics. As expected, 40% O 2 + 5% CO 2 treatment promoted the chlorophyll degradation and carotenoid synthesis, and thus to accelerate the yellowing of broccoli heads. In addition, elevated O 2 decreased the ascorbic acid, glucosinolate, and flavonoid contents and increased the MDA content. A total of 189 proteins related to the yellowing of broccoli heads were identified as differentially expressed proteins (fold change ≥1.20 or ≤0.83). These proteins were mainly associated with pigment metabolism, thylakoid structure, photosynthetic system, chloroplast ribosome, antioxidant substance metabolism, and antioxidant enzyme metabolism. The proteomics results showed 40% O 2 + 5% CO 2 treatment disrupted antioxidant metabolism and aggravated oxidative stress compared with 5% O 2 + 5% CO 2 and 20% O 2 + 5% CO 2 treatments, thus leading to the destruction of photosystems I and II, thylakoid membrane, and chloroplast ribosome in broccoli heads. In addition, 40% O 2 + 5% CO 2 treatment accelerated the yellowing of postharvest broccoli heads by down-regulating chlorophyll b iosynthetic pathway and the expression of apolipoproteins (light-harvesting complex I, II, and oxygen-evolving enhancer protein) and ribulose-1,5-bisphosphate carboxylase (Rubisco), and up-regulating chlorophyll degradation and carotenoid biosynthetic pathways. This study might improve our understanding of yellowing acceleration mechanisms of broccoli heads by elevated O 2 , thereby promoting the development of new broccoli heads preservation technology. [ABSTRACT FROM AUTHOR]

• Elevated O 2 concentration accelerated chlorophyll degradation. • H 2 O 2 content decreased along with O 2 concentration elevated. • H 2 O 2 down-regulated the expression of BoSGR1, BoNYC, BoPPH, BoPAO , and BoRCCR. • Elevated O 2 concentration enhanced Chl-POX activity at the transcription level. • Chl oxidative dominated the Chl degradation under elevated O 2 concentration. Postharvest broccoli heads undergo yellowing quickly in unsuitable conditions, which diminishes their quality and commercial value. We investigated the pattern and molecular regulatory mechanism of chlorophyll (Chl) degradation in broccoli under elevated O 2 controlled atmosphere. As results, the 40% O 2 + 5% CO 2 treatment accelerated the Chl degradation and the change in color of postharvest broccoli heads, while the 5% O 2 + 5% CO 2 treatment delayed the yellowing. The results indicated that Chl degradation pattern of postharvest broccoli heads concurrently involved the Pheide a oxygenase (PAO)/phyllobilin and oxidative degradation pathways, and the latter might be the dominant pathway of Chl degradation. The regulatory mechanism of elevated O 2 on Chl degradation may be related to its ability to accelerate cell damage and death, destroy the light-harvesting complex II, and promote up-regulation of genes in the PAO/phyllobilin and oxidative degradation pathways. In contrast, low O 2 treatment may down-regulate the expression of Chl degradation-related genes by increasing H 2 O 2 content, thus delaying the yellowing of broccoli heads. [ABSTRACT FROM AUTHOR]