Impact of Storage Controlled Atmosphere on the Apple Phenolic Acids, Flavonoids, and Anthocyanins and Antioxidant Activity In Vitro

Apples are seasonal fruits, and it is important to prepare them adequately for storage and ensure proper storage conditions. In this study, we used ten different apple cultivars: ‘Alva’, ‘Auksis’, ‘Connell Red’, ‘Cortland’, ‘Ligol’, ‘Lodel’, ‘Noris’, ‘Rubin’, ‘Sampion’, and ‘Spartan’. We studied the qualitative and quantitative composition of phenolic compounds in the apple and apple extracts antioxidants activity before placing them in the controlled atmosphere chambers and again at the end of the experiment, eight months later. Different concentrations of O2, CO2, and N2, constant temperature, relative humidity, and removal of endogenous ethylene were continually maintained. HPLC analysis showed that the highest amount of 2265.7 ± 152.5 µg/g of chlorogenic acid was found in apple samples of the ‘Auksis’ cultivar stored under variant IV conditions. Different concentrations of gas in the controlled atmosphere chambers caused changes in antioxidant activity in whole apple and apple peel extracts. In our study, we found that the antioxidant activity of apple extracts varied between samples of different apple cultivars and depended on the composition of the controlled atmosphere. Determining the optimal storage conditions is beneficial to providing the consumers with apples that have a known and minimally altered chemical composition of phenolic compounds and the strongest antioxidant activity, which determine the use of apples in the healthy food chain.

Determination of morpholine residues in apples by liquid chromatography tandem mass spectrophotometry (LC-MS/MS)

Morpholine is an emulsifier that is normally used as a protective wax coating on the surfaces of fruit and vegetable. In order to keep fruits fresh, extend their shelf-life, against insect and fungal contamination for a long time, morpholine has been widely applied despite of the reported carcinogenic potential in previous studies and export prohibition in many countries. A highly sensitive method has been developed for the determination of this food additive in fruits using ultra-performance liquid chromatography tandem mass spectrophotometry. By using 1 % acetic acid in methanol for extraction and analyzing with binary ammonium formate 20 mM and acetonitrile, the method has been applied successfully to determine morpholine residues in apple samples collected at local markets in Hanoi, Vietnam. The good coefficient correlation was achieved in the linear range of 5 - 300 µg/L (R2 = 0.9998). The low limit of detection (LOD) and limit of quantification (LOQ) were 2 µg/kg and 5 µg/kg, respectively. The recoveries of morpholine were 83 – 108 % over three spike levels 20 - 60 μg/L for apple, peel and pulp matrices. The repeatability (RSDr %) was 1.1 % for apple matrix, 2.36 % for peel and 3.67 % for pulp. The reproducibility values (RSDR %) were 1.49, 3.50 and 4.08 % for apple, peel and pulp, respectively. This method could be further performed in different type of fruit and vegetable comiditities to determine the morpholine content.

Metabolomic and Transcriptomic Analysis of the Anthocyanin Regulatory Networks in Malus Domestica Borkh. Peel with Different Color Patterns

Background:Coloring is an important external quality of ‘Fuji’ apple (Malus domestica Borkh.) and there are two color patterns of apple peels, i.e., stripe and blush. The objectives of this study were to reveal the anthocyanin biosynthesis metabolic pathway in striped and blushed peels of Malus domestica using metabolomics and transcriptomics, to identify different anthocyanin metabolites, and to analyze the differentially expressed genes involved in anthocyanin biosynthesis.Result:The metabolite concentration and gene expression were profiled in the striped and blushed fruit peels of apple harvested at three ripening periods to elucidate the color formation mechanism. At the green fruit period, there were 83 DAMs,including 30 flavonoids, 674 DEGs (521 up-regulated and 153 down-regulated),including 3 MYB related genes (up-regulated, LOC103415449, LOC103421948, LOC103432338) and 2 bHLH genes(up-regulated, LOC103436250, LOC103437863) between striped and blushed apple.At the color turning period, there were 48 DAMs,including 20 flavonoids, 880 DEGs (274 up-regulated and 606 down-regulated), including 3 differentially expressed E2.3.1.133, HCT genes(down-regulated), 2 differentially expressed F3H genes (down-regulated), 1 differentially expressed BZ1 gene (down-regulated) and 2 differentially expressed ANS genes (up-regulated) and 2 up-regulated MYB related genes (LOC103411576, LOC103412495), 5 down-regulated MYB related genes(LOC103400953, LOC103408672, LOC103415404, LOC103420697, LOC103421948), 1 differentially expressed bHLH gene(down-regulated, LOC103400870). At the complete coloring period，there were 95 DAMs,including 34 flavonoids, 2258 DEGs (1159 up- and 1099 down-regulated), including 3 differentially expressed E2.3.1.133, HCT genes(down-regulated), 1 differentially expressed E2.3.1.133, HCT genes(up-regulated), 2 differentially expressed CYP98A genes (up-regulated), 4 differentially expressed CHS genes (up-regulated), 2 differentially expressed E5.5.1.6 genes(up-regulated), 2 differentially expressed CYP75B1 genes (up-regulated), 2 differentially expressed F3R genes (up-regulated), 2 differentially expressed ANS genes (up-regulated), 1 differentially expressed DFR genes (up-regulated), 2 differentially expressed BZ1 genes (up-regulated) and 1 differentially expressed MYB related gene (up-regulated, LOC103401575) .There were both 10 kinds of cyanidin in apple peel at color turning period and complete coloring period, Keracyanin and Cyanin were up-regulated at color turning period and Cyanidin-3-O-(6''-O-malonyl)glucoside was up-regulated at complete coloring period.Conclusions: Our researches provide important information on the anthocyanin metabolites and the candidate genes involved in the anthocyanin biosynthesis pathways of Fuji apple in M.domestcia.

Agricultural Waste Annona squamosa L. Peel and Seed Extract: Biosynthesis of Hand Sanitizer Gel against Skin Pathogens

Alcohol-based hand sanitizers are being recommended as an infection prevention measure for COVID-19. Washing hands is a simple matter to eliminate and minimize germs on the hands with water and by adding certain ingredients, the use of hand sanitizer becomes must in recent circumstances. We have developed reliable and eco-friendly process for the synthesis of hand sanitizer gel that is a really useful tool in the fight against SARS-CoV-2, the virus that causes coronavirus disease 2019 (COVID-19). Considering scanty literature available on the use of custard apple peel and seed for preparation of hand sanitizer gel, present study was undertaken to evaluate effect of hand sanitizer from custard apple peel and seed extract. The antimicrobial activity of the formulated gel was tested against Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Bacillus subtilis subsp. spizizenii, Salmonella typhimurinum and Shigella sonnei by The Quality Assurance and Testing Center 3 (QUATEST 3) in Ho Chi Minh city, Viet Nam.

MdBBX21, a B-Box Protein, Positively Regulates Light-Induced Anthocyanin Accumulation in Apple Peel

The red coloration of apple (Malus × domestica Borkh.) is due to the accumulation of anthocyanins in the fruit peel. Light is essential for anthocyanin biosynthesis in apple. In this study, we performed a transcriptome sequencing (RNA-seq) analysis of apple fruit exposed to light after unbagging. The identified differentially expressed genes included MdBBX21, which is homologous to Arabidopsis BBX21, suggesting it may be involved in light-induced anthocyanin biosynthesis. Additionally, MdBBX21 was localized in the nucleus and its gene was expressed earlier than MdMYB1 in apple peel treated with light. Overexpressing MdBBX21 in Arabidopsis and apple calli under light increased anthocyanin accumulation. Dual-luciferase and yeast one-hybrid assays confirmed that MdBBX21 binds to the MdHY5, MdBBX20, and MdBBX22-1/2 promoters and induces expression. At the same time, MdHY5 can also activate the expression of MdBBX21. Furthermore, bimolecular fluorescence complementation and yeast two-hybrid assays demonstrated that MdBBX21 can interact with MdHY5. This interaction can significantly enhance MdMYB1 promoter activity. These findings clarify the molecular mechanism by which MdBBX21 positively regulates light-induced anthocyanin accumulation in apple.

Vitamin-Containing Antioxidant Formulation Reduces Carcinogen-Induced DNA Damage through ATR/Chk1 Signaling in Bronchial Epithelial Cells In Vitro

Lung cancer has the highest mortality rate worldwide and is often diagnosed at late stages, requiring genotoxic chemotherapy with significant side effects. Cancer prevention has become a major focus, including the use of dietary and supplemental antioxidants. Thus, we investigated the ability of an antioxidant formulation (AOX1) to reduce DNA damage in human bronchial epithelial cells (BEAS-2B) with and without the combination of apple peel flavonoid fraction (AF4), or its major constituent quercetin (Q), or Q-3-O-d-glucoside (Q3G) in vitro. To model smoke-related genotoxicity, we used cigarette-smoke hydrocarbon 4-[(acetoxymethyl)nitrosamino]-1-(3-pyridyl)-1-butanone (NNKOAc) as well as methotrexate (MTX) to induce DNA damage in BEAS-2B cells. DNA fragmentation, γ-H2AX immunofluorescence, and comet assays were used as indicators of DNA damage. Pre-exposure to AOX1 alone or in combination with AF4, Q, or Q3G before challenging with NNKOAc and MTX significantly reduced intracellular reactive oxygen species (ROS) levels and DNA damage in BEAS-2B cells. Although NNKOAc-induced DNA damage activated ATM-Rad3-related (ATR) and Chk1 kinase in BEAS-2B cells, pre-exposure of the cells with tested antioxidants prior to carcinogen challenge significantly reduced their activation and levels of γ-H2AX (p ≤ 0.05). Therefore, AOX1 alone or combined with flavonoids holds promise as a chemoprotectant by reducing ROS and DNA damage to attenuate activation of ATR kinase following carcinogen exposure.

Using Biological Elicitation to Improve Type 2 Diabetes Targeted Food Quality of Stored Apple

Food quality improvements of fresh fruits targeting both food preservation and human health is essential to advance healthy dietary options and to mitigate imbalanced nutrition-linked non-communicable chronic disease (NCDs) challenges globally. Specifically, protective phenolic bioactives of fruits with dual functional benefits can be harnessed to advance innovations for improving nutritional quality and post-harvest shelf-life of perishable fruits. Based on this rationale the dual functional benefits of plant phenolics were harnessed using novel biological elicitation strategies to modulate phenolic bioactive-linked protective responses in apple during storage in two interrelated studies. Bioprocessed food-grade elicitors [water soluble chitosan oligosaccharide -(COS) and phenolic enriched oregano extracts-(OX)] were targeted as post-harvest dipping treatments (2 & 4 g/ L) and compared with diphenylamine (DPA) (1 & 2 g/L) to enhance phenolic-linked antioxidant and anti-diabetic (type 2 diabetes) relevant properties of Cortland apple during 3 months of storage (4°C). The selection of bio-elicitors and respective doses were based on the foundations of the previous related study, which resulted in reduction of superficial scald of Cortland apple during storage. Apples sampled over 3 months as aqueous and ethanol (12%) extracts of peel and pulp were analyzed separately for total soluble phenolic content, phenolic profile, antioxidant activity, and glucose metabolism relevant α-amylase and α-glucosidase enzyme inhibitory activities using in vitro assay models. Enhanced soluble phenolic content and associated antioxidant activity were observed in ethanol (12%) extracts of apple peel with 4 g/L COS elicitor treatments after 2 and 3 months of storage. High chlorogenic acid and quercetin derivatives were found in peel extracts of Cortland apple, while pulp extracts had high chlorogenic and gallic acids. Additionally, high α-glucosidase enzyme inhibitory activity, which is relevant for managing post-prandial hyperglycemia of type 2 diabetes was also observed in bio-elicited apple peel and pulp extracts. Therefore, results of these two interrelated studies indicate that bioprocessed food grade elicitor such as OX and COS can be recruited as a novel tool to enhance protective phenolic responses for improving type 2 diabetes targeted food quality and post-harvest storage quality of apple.