Acetic Acid Assay Kit (ACS Manual Format)

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00:05   Introduction
00:52   Principle
01:21     Reagent Preparation
02:04   Procedure
06:04   Calculations

Acetic Acid Assay Kit ACS Manual Format K-ACET Scheme
   
Reference code: K-ACET
SKU: 700004254

53 assays per kit

Content: 53 assays per kit
Shipping Temperature: Ambient
Storage Temperature: Short term stability: 2-8oC,
Long term stability: See individual component labels
Stability: > 2 years under recommended storage conditions
Analyte: Acetic Acid
Assay Format: Spectrophotometer
Detection Method: Absorbance
Wavelength (nm): 340
Signal Response: Increase
Linear Range: 0.3 to 20 µg of acetic acid per assay
Limit of Detection: 0.14 mg/L
Reaction Time (min): ~ 14 min
Application examples: Wine, beer, fruit and fruit juices, soft drinks, vinegar, vegetables, pickles, dairy products (e.g. cheese), meat, fish, bread, bakery products (and baking agents), ketchup, soy sauce, mayonnaise, dressings, paper (and cardboard), tea, pharmaceuticals (e.g. infusion solutions), feed and other materials (e.g. biological cultures, samples, etc.).
Method recognition: Methods based on this principle have been accepted by EN, ISO,ICUMSA, IFU and MEBAK

The Acetic Acid (ACS Manual Format) test kit is a simple method for the rapid and reliable measurement and analysis of acetic acid/acetate in foods, beverages and other materials.

Note for Content: The number of manual tests per kit can be doubled if all volumes are halved.  This can be readily accommodated using the MegaQuantTM  Wave Spectrophotometer (D-MQWAVE).

See our full range of organic acid test kits.

Scheme-K-ACET ACET Megazyme

Advantages
  • No wasted ACS solution (stable suspension supplied) 
  • PVP incorporated to prevent tannin inhibition 
  • All reagents stable for > 2 years after preparation
  • Very competitive price (cost per test) 
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing
Documents
Certificate of Analysis
Safety Data Sheet
FAQs Assay Protocol Data Calculator Other automated assay procedures Product Performance Validation Report
Publications
Megazyme publication

Megazyme “advanced” wine test kits general characteristics and validation.

Charnock, S. J., McCleary, B. V., Daverede, C. & Gallant, P. (2006). Reveue des Oenologues, 120, 1-5.

Many of the enzymatic test kits are official methods of prestigious organisations such as the Association of Official Analytical Chemicals (AOAC) and the American Association of Cereal Chemists (AACC) in response to the interest from oenologists. Megazyme decided to use its long history of enzymatic bio-analysis to make a significant contribution to the wine industry, by the development of a range of advanced enzymatic test kits. This task has now been successfully completed through the strategic and comprehensive process of identifying limitations of existing enzymatic bio-analysis test kits where they occurred, and then using advanced techniques, such as molecular biology (photo 1), to rapidly overcome them. Novel test kits have also been developed for analytes of emerging interest to the oenologist, such as yeast available nitrogen (YAN; see pages 2-3 of issue 117 article), or where previously enzymes were simply either not available, or were too expensive to employ, such as for D-mannitol analysis.

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Megazyme publication

Grape and wine analysis: Oenologists to exploit advanced test kits.

Charnock, S. C. & McCleary, B. V. (2005). Revue des Enology, 117, 1-5.

It is without doubt that testing plays a pivotal role throughout the whole of the vinification process. To produce the best possible quality wine and to minimise process problems such as “stuck” fermentation or troublesome infections, it is now recognised that if possible testing should begin prior to harvesting of the grapes and continue through to bottling. Traditional methods of wine analysis are often expensive, time consuming, require either elaborate equipment or specialist expertise and frequently lack accuracy. However, enzymatic bio-analysis enables the accurate measurement of the vast majority of analytes of interest to the wine maker, using just one piece of apparatus, the spectrophotometer (see previous issue No. 116 for a detailed technical review). Grape juice and wine are amenable to enzymatic testing as being liquids they are homogenous, easy to manipulate, and can generally be analysed without any sample preparation.

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Publication

Isolation and characterization of a novel methanogen Methanosarcina hadiensis sp. nov. from subsurface Boom Clay pore water.

Giménez, F. J., Peeters, E., Honty, M., Leys, N. & Mijnendonckx, K. (2024). Environmental Microbiology26(12), e70004.

Safe geological disposal of radioactive waste requires a thorough understanding of geochemical conditions in the host formation. Boom Clay is a potential candidate in Belgium, where active methanogenesis has been detected in its deep subsurface, influencing the local geochemistry. However, the pathways driving this process and the characteristics of the methanogenic archaea involved remain unclear. We isolated a distinct archaeal strain from Boom Clay pore water and characterized it geno- and phenotypically. Isolate TD41E1-1 belongs to a novel species of the Methanosarcina genus, for which the name Methanosarcina hadiensis sp. nov. is proposed. TD41E1-1 cells are coccus-shaped, irregularly sized cells enveloped by extracellular polymer substances. Growth and substrate utilization experiments and genomic analysis demonstrated that the strain prefers methylated compounds or hydrogen as substrates for methane production. Although it possesses a complete acetoclastic pathway, no growth was observed in the presence of acetate in the tested conditions. Based on its phylogenetic relation to other known Methanosarcina species and on the presence of c-type cytochromes, it can be concluded that the strain likely occupies an intermediate position between type I and type II Methanosarcina species. These findings provide valuable insights for assessing Boom Clay's suitability for geological disposal of radioactive waste.

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Publication

Dextran-enriched pea-based ingredient from a combined enzymatic and fermentative bioprocessing. Design of an innovative plant-based spread.

Perri, G., Difonzo, G., Wang, Y., Verni, M., Caponio, G. R., Coda, R., Blandino, M. & Pontonio, E. (2024). Future Foods, 10, 100502.

In this study a plant-based spread was developed using dextran-enriched ingredients derived from pea flours, supplemented with defatted durum wheat germ and almond flour. Optimization of fermentation with Leuconostoc pseudomesenteroides DSM 20193, both with and without enzymatic hydrolysis, aimed to enhance exopolysaccharide production and the nutritional value of pea flours. Best results were achieved through enzymatic hydrolysis with Veron PS protease followed by fermentation at 25°C, resulting in elevated dextran levels and increased peptides and total free amino acid concentration in green and yellow pea-based ingredients. The yellow pea-based ingredient was selected for the final plant-based spread formulation, blended at 35% w/w, with 45% w/w defatted durum wheat germ, and 20% w/w almond flour. The resultant spread exhibited elastic and solid-like characteristics like milk-based spreadable cheese and yogurt, boasting 'high protein' (12.49 g/100g) and 'high fiber' (11.01 g/100g) designations. It maintained chemical, biochemical, and microbiological stability over a 10-day shelf-life under refrigerated conditions. Sensory evaluation confirmed the acceptability of the plant-based spread (PBS), highlighting a well-balanced aroma and a grainy, adhesive texture. This research underscores the potential of an integrated approach utilizing food-grade enzymes and fermentation for the in-situ production of dextran to create innovative, clean label, and plant-based foods.

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Publication

Acetyl-CoA synthetase activity is enzymatically regulated by lysine acetylation using acetyl-CoA or acetyl-phosphate as donor molecule.

Qin, C., Graf, L. G., Striska, K., Janetzky, M., Geist, N., Specht, R., Schulze, S., Palm, G. J., Girbardt, B., Dörre, B., Berndt, L., Kemnitz, S., Doerr, M., Bornscheuer, U. T., Delcea, M. & Lammers, M. (2024). Nature Communications, 15(1), 6002.

The AMP-forming acetyl-CoA synthetase is regulated by lysine acetylation both in bacteria and eukaryotes. However, the underlying mechanism is poorly understood. The Bacillus subtilis acetyltransferase AcuA and the AMP-forming acetyl-CoA synthetase AcsA form an AcuA•AcsA complex, dissociating upon lysine acetylation of AcsA by AcuA. Crystal structures of AcsA from Chloroflexota bacterium in the apo form and in complex with acetyl-adenosine-5'-monophosphate (acetyl-AMP) support the flexible C-terminal domain adopting different conformations. AlphaFold2 predictions suggest binding of AcuA stabilizes AcsA in an undescribed conformation. We show the AcuA•AcsA complex dissociates upon acetyl-coenzyme A (acetyl-CoA) dependent acetylation of AcsA by AcuA. We discover an intrinsic phosphotransacetylase activity enabling AcuA•AcsA generating acetyl-CoA from acetyl-phosphate (AcP) and coenzyme A (CoA) used by AcuA to acetylate and inactivate AcsA. Here, we provide mechanistic insights into the regulation of AMP-forming acetyl-CoA synthetases by lysine acetylation and discover an intrinsic phosphotransacetylase allowing modulation of its activity based on AcP and CoA levels.

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Publication

Monitoring corn stover processing by the fungus Ustilago maydis.

Robertz, S., Philipp, M., Schipper, K., Richter, P., Miebach, K., Magnus, J., Pauly, M. & Ramírez, V. (2024). Bioresources and Bioprocessing, 11(1), 87.

A key aspect of sustainable bioeconomy is the recirculation of renewable, agricultural waste streams as substrates for microbial production of high-value compounds. One approach is the bioconversion of corn stover, an abundant maize crop byproduct, using the fungal maize pathogen Ustilago maydis. U. maydis is already used as a unicellular biocatalyst in the production of several industrially-relevant compounds using plant biomass hydrolysates. In this study, we demonstrate that U. maydis can grow using untreated corn stover as its sole carbon source. We developed a small-scale bioreactor platform to investigate U. maydis processing of corn stover, combining online monitoring of fungal growth and metabolic activity profiles with biochemical analyses of the pre- and post-fermentation residues. Our results reveal that U. maydis primarily utilizes soluble sugars i.e., glucose, sucrose and fructose present in corn stover, with only limited exploitation of the abundant lignocellulosic carbohydrates. Thus, we further explored the biotechnological potential of enhancing U. maydis´ lignocellulosic utilization. Additive performance improvements of up to 120 % were achieved when using a maize mutant with increased biomass digestibility, co-fermentation with a commercial cellulolytic enzyme cocktail, and exploiting engineered fungal strains expressing diverse lignocellulose-degrading enzymes. This work represents a key step towards scaling up the production of sustainable compounds from corn stover using U. maydis and provides a tool for the detailed monitoring of the fungal processing of plant biomass substrates.

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Publication

Functional and biochemical characterization of pre-fermented ingredients obtained by the fermentation of durum wheat by-products.

Rossi, S., Gottardi, D., Siroli, L., Giordani, B., Vitali, B., Vannini, L., Patrignani, F. & Lanciotti, R. (2024). Journal of Functional Foods, 116, 106136.

This work was aimed to characterize functional and biochemical parameters of a bakery ingredient prepared with durum wheat by-products (micronized bran and middling) fermented by a selected microbial consortium composed of yeasts and lactic acid bacteria. The unfermented milling by-products mixture and the mixture fermented by a baker’s yeast were used as reference. The innovative ingredient showed more stable colour indexes compared to the references, a more complex profile in volatile molecules characterized by a higher presence of alcohols, ketones and acids compared to the references. A significant increase in the content of peptides, short chain fatty acids, total phenols, antioxidant activity and prebiotic activity together with a reduction in phytic acid content was observed in the samples fermented by the selected microbial consortium compared to the references. This work provides information on the impact of lactic acid bacteria and yeasts on functional and biochemical characteristics of fermented milling by-products.

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Publication

Design of a Plant-Based Yogurt-Like Product Fortified with Hemp Flour: Formulation and Characterization.

Montemurro, M., Verni, M., Rizzello, C. G. & Pontonio, E. (2023). Foods, 12(3), 485.

Plant-based milk alternatives have gained massive popularity among consumers because of their sustainable production compared to bovine milk and because of meeting the nutritional requests of consumers affected by cow milk allergies and lactose intolerance. In this work, hemp flour, in a blend with rice flour, was used to design a novel lactose- and gluten-free yogurt-like (YL) product with suitable nutritional, functional, and sensory features. The growth and the acidification of three different lactic acid bacteria strains were monitored to better set up the biotechnological protocol for making the YL product. Hemp flour conferred the high fiber (circa 2.6 g/100 g), protein (circa 4 g/100 g), and mineral contents of the YL product, while fermentation by selected lactic acid bacteria increased the antioxidant properties (+8%) and the soluble fiber (+0.3 g/100 g), decreasing the predicted glycemic index (-10%). As demonstrated by the sensory analysis, the biotechnological process decreased the earthy flavor (typical of raw hemp flour) and increased the acidic and creamy sensory perceptions. Supplementation with natural clean-label vanilla powder and agave syrup was proposed to further decrease the astringent and bitter flavors. The evaluation of the starter survival and biochemical properties of the product under refrigerated conditions suggests an estimated shelf-life of 30 days. This work demonstrated that hemp flour might be used as a nutritional improver, while fermentation with a selected starter represents a sustainable and effective option for exploiting its potential.

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High-throughput mass spectrometry analysis revealed a role for glucosamine in potentiating recovery following desiccation stress in Chironomus.

Thorat, L., Oulkar, D., Banerjee, K., Gaikwad, S. M. & Nath, B. B. (2017). Scientific Reports7(1), 3659.

Desiccation tolerance is an essential survival trait, especially in tropical aquatic organisms that are vulnerable to severe challenges posed by hydroperiodicity patterns in their habitats, characterized by dehydration-rehydration cycles. Here, we report a novel role for glucosamine as a desiccation stress-responsive metabolite in the underexplored tropical aquatic midge, Chironomus ramosus. Using high- throughput liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QToF-MS) analysis, biochemical assays and gene expression studies, we confirmed that glucosamine was essential during the recovery phase in C. ramosus larvae. Additionally, we demonstrated that trehalose, a known stress-protectant was crucial during desiccation but did not offer any advantage to the larvae during recovery. Based on our findings, we emphasise on the collaborative interplay of glucosamine and trehalose in conferring overall resilience to desiccation stress and propose the involvement of the trehalose-chitin metabolic interface in insects as one of the stress-management strategies to potentiate recovery post desiccation through recruitment of glucosamine.

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Publication

Continuous fed-batch strategy decreases acetic acid production and increases volatile ester formation in wines under high-gravity fermentation.

Deng, H., Wang, M. & Li, E. (2023). OENO One, 57(1), 363-374.

High sugar fermentation elevates acetic acid levels in wines, which can be avoided by applying the continuous fed-batch strategy. In this study, yeast gene expressions and wine volatile compounds were evaluated by quantitative real-time PCR (RT-qPCR) and gas chromatograph mass spectrometry (GC-MS) in high-gravity (HG, 320 g/L sugars) fermentations with different batch strategies. The acetic acid concentration in continuous fed-batch fermentation wine was reduced by 51.69 %, compared with that in whole-batch fermentation wine. The acetyl-CoA synthase gene (ACS2) expression was up-regulated, whereas the glycerol-3-phosphate dehydrogenase gene (GPD1) expression was down-regulated on day 3 and day 7 during the continuous fed-batch fermentation. The volatile ester concentration in continuous fed-batch fermentation wine was 36.74 % higher than that in whole-batch fermentation wine. Overall, the continuous fed-batch strategy can modulate the expression of yeast genes involved in acetic acid metabolism and can increase volatile esters in wine under high sugar fermentation. Our findings provide a reference for the application of a continuous fed-batch strategy in high-sugar fermentation so as to improve the quality of the wine.

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Up-cycling grape pomace through sourdough fermentation: Characterization of phenolic compounds, antioxidant activity, and anti-inflammatory potential.

Torreggiani, A., Demarinis, C., Pinto, D., Papale, A., Difonzo, G., Caponio, F., Pontonio, E., Verni, M. & Rizzello, C. G. (2023).. Antioxidants12(8), 1521.

Despite its appealing composition, because it is rich in fibers and polyphenols, grape pomace, the major by-product of the wine industry, is still discarded or used for feed. This study aimed at exploiting grape pomace functional potential through fermentation with lactic acid bacteria (LAB). A systematic approach, including the progressively optimization of the grape pomace substrate, was used, evaluating pomace percentage, pH, and supplementation of nitrogen and carbon sources. When grape pomace was used at 10%, especially without pH correction, LAB cell viability decreased up to 2 log cycles. Hence, the percentage was lowered to 5 or 2.5% and supplementations with carbon and nitrogen sources, which are crucial for LAB metabolism, were considered aiming at obtaining a proper fermentation of the substrate. The optimization of the substrate enabled the comparison of strains performances and allowed the selection of the best performing strain (Lactiplantibacillus plantarum T0A10). A sourdough, containing 5% of grape pomace and fermented with the selected strain, showed high antioxidant activity on DPPH and ABTS radicals and anti-inflammatory potential on Caco2 cells. The anthocyanins profile of the grape pomace sourdough was also characterized, showing qualitative and quantitative differences before and after fermentation. Overall, the grape pomace sourdough showed promising applications as a functional ingredient in bread making.

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Publication

Agaricus bisporus chitosan influences the concentrations of caftaric acid and furan-derived compounds in Pinot noir juice and base wine.

Mederios, J., Xu, S., Pickering, G. & Kemp, B. (2023). Oeno One, 57(3), 255-268.

Chitosan is a fining agent used in winemaking, although its use in juice and wine beyond fining has been limited until now. Therefore, this study's first aim was to determine if chitosan derived from Agaricus bisporus (button mushrooms) could reduce caffeic and caftaric acid concentrations in Pinot noir grape juice (Study A). The second aim was to determine if chitosan, when added to base wine, could influence the synthesis of furan-derived compounds during storage (Study B). In Study A, Pinot noir grape juice was stored at 10°C for 18 hours after the following treatments: control (no addition), bentonite/activated charcoal (BAC), low molecular weight (< 3 kDa; LMW) chitosan, med. MW (250 kDa; MMW) chitosan, and high MW (422 kDa; HMW) chitosan (all 1 g/L additions). Caftaric acid was decreased, and total amino acid concentration was increased in the LMW chitosan-treated juice, while the estimated total hydroxycinnamic acid content, turbidity, and browning were decreased in the MMW chitosan-treated juice compared to the control. In Study B, Pinot noir base wine destined for sparkling wine was stored at 15 and 30°C for 90 days with the following treatments: control (no addition), LMW chitosan, MMW chitosan, and HMW chitosan (all 1 g/L additions). The three chitosan treatments stored at 30°C had increased furfural, homofuraneol, and 5-methylfurfural formation in the base wine compared to the control. At 15°C, furfural and homofuraneol had greater concentrations in all chitosan-treated wines after 90 days of storage. Our results demonstrate the potential of mushroom-derived chitosan to remove caftaric acid from grape juice and suggest that chitosan can influence the synthesis of furan-derived compounds in wine after short-term storage.

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Growth-coupled anaerobic production of isobutanol from glucose in minimal medium with Escherichia coli.

Boecker, S., Schulze, P. & Klamt, S. (2023). Biotechnology for Biofuels and Bioproducts, 16(1), 148.

Background: The microbial production of isobutanol holds promise to become a sustainable alternative to fossil-based synthesis routes for this important chemical. Escherichia coli has been considered as one production host, however, due to redox imbalance, growth-coupled anaerobic production of isobutanol from glucose in E. coli is only possible if complex media additives or small amounts of oxygen are provided. These strategies have a negative impact on product yield, productivity, reproducibility, and production costs. Results: In this study, we propose a strategy based on acetate as co-substrate for resolving the redox imbalance. We constructed the E. coli background strain SB001 (ΔldhA ΔfrdA ΔpflB) with blocked pathways from glucose to alternative fermentation products but with an enabled pathway for acetate uptake and subsequent conversion to ethanol via acetyl-CoA. This strain, if equipped with the isobutanol production plasmid pIBA4, showed robust exponential growth (µ = 0.05 h−1) under anaerobic conditions in minimal glucose medium supplemented with small amounts of acetate. In small-scale batch cultivations, the strain reached a glucose uptake rate of 4.8 mmol gDW−1 h−1, a titer of 74 mM and 89% of the theoretical maximal isobutanol/glucose yield, while secreting only small amounts of ethanol synthesized from acetate. Furthermore, we show that the strain keeps a high metabolic activity also in a pulsed fed-batch bioreactor cultivation, even if cell growth is impaired by the accumulation of isobutanol in the medium. Conclusions: This study showcases the beneficial utilization of acetate as a co-substrate and redox sink to facilitate growth-coupled production of isobutanol under anaerobic conditions. This approach holds potential for other applications with different production hosts and/or substrate–product combinations.

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Combined use of Trametes versicolor extract and sourdough fermentation to extend the microbiological shelf-life of baked goods.

Torreggiani, A., Beccaccioli, M., Verni, M., Cecchetti, V., Minisci, A., Reverberi, M., Pontonio, E. & Rizzello, C. G. (2023). LWT, 189, 115467.

Fungal spoilage is the main responsible for the short shelf-life of baked goods. Indeed, many chemical preservatives (e.g., calcium propionate and ethanol) are often included in their formulation leading to consumer dissatisfaction. Here, an in-vitro and in-situ integrated approach was used to investigate the potential antifungal activity of the extract obtained from Trametes versicolor as biological preservative. An intense inhibition towards most of the Aspergillus and Penicillium species and a broad spectrum of activity on typical spoilage fungi of the bakery products, as well as high thermal stability and intense activity at pH 4.00 characterized the Trametes extract. The antifungal potential of the extract has been exploited in sour bread made with organic acids (chemical) or type II-sourdough (biological). Compared to baker's yeast bread produced as control (pH 5.6), the acidified samples were characterized by a longer mold-free shelf-life, with indicator mycelia that became visible after 6–9 days of storage at room temperature. Sourdough was effective to counteract the negative effect of the extract supplementation on the leavening performances, textural and sensory features of the bread samples.

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Expression and Molecular Modification of Chitin Deacetylase from Streptomyces bacillaris.

Yin, L., Wang, Q., Sun, J. & Mao, X. (2023). Molecules, 28(1), 113.

Chitin deacetylase can be used in the green and efficient preparation of chitosan from chitin. Herein, a novel chitin deacetylase SbCDA from Streptomyces bacillaris was heterologously expressed and comprehensively characterized. SbDNA exhibits its highest deacetylation activity at 35°C and pH 8.0. The enzyme activity is enhanced by Mn2+ and prominently inhibited by Zn2+, SDS, and EDTA. SbCDA showed better deacetylation activity on colloidal chitin, (GlcNAc)5, and (GlcNAc)6 than other forms of the substrate. Molecular modification of SbCDA was conducted based on sequence alignment and homology modeling. A mutant SbCDA63G with higher activity and better temperature stability was obtained. The deacetylation activity of SbCDA63G was increased by 133% compared with the original enzyme, and the optimal reaction temperature increased from 35 to 40°C. The half-life of SbCDA63G at 40°C is 15 h, which was 5 h longer than that of the original enzyme. The improved characteristics of the chitin deacetylase SbCDA63G make it a potential candidate to industrially produce chitosan from chitin.

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Safety Information
Symbol : GHS08
Signal Word : Danger
Hazard Statements : H319, H360
Precautionary Statements : P201, P202, P280, P308+P313, P405, P501
Safety Data Sheet
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