Effect of high-pressure processing on bioactive compounds in pomegranate juice

Introduction

An observation that surprises many consumers: thermal pasteurisation of pomegranate juice destroys 25-40% of its anthocyanins and up to 50% of its vitamin C, yet it remains the industry standard because no affordable non-thermal alternative has been fully validated [1] . High-pressure processing (HPP) applies isostatic pressures of 100-600 MPa for seconds to minutes, inactivating vegetative bacteria, yeasts, and moulds while leaving small molecules-vitamins, pigments, flavour volatiles-largely intact [2] . Pomegranate juice is one of the richest dietary sources of punicalagins, ellagic acid, and acylated anthocyanins (cyanidin-, delphinidin-, and pelargonidin-3-glucosides), all of which contribute to its documented cardioprotective, anti-inflammatory, and antiproliferative activities [3] . These compounds are heat-labile, making pomegranate an ideal candidate for HPP. Earlier HPP trials on pomegranate juice focused on 300-600 MPa ranges but rarely measured bioactive retention at fine pressure intervals or tracked colour stability beyond 14 days [4] . This research therefore applied HPP at five pressure levels (200-600 MPa) to freshly extracted pomegranate juice and monitored bioactive compounds, microbial counts, and colour over 28 days of refrigerated storage to identify the minimum effective pressure for commercial-quality juice.

Material and Methods

Material

Ripe pomegranate fruits (cv. Bhagwa) were purchased from a wholesale market in İzmir, during July 2023. Arils were manually separated, juice extracted by hydraulic pressing, and filtered through muslin cloth. No preservatives or additives were used. Analytical chemicals were from Sigma-Aldrich (Germany) and Merck (Germany). HPP was performed on a laboratory-scale unit (Hiperbaric 55, Spain) at the Department of Food Science and Technology, Ege University, Bornova, İzmir [5] .

Methods

Juice (200 mL per pouch) was vacuum-sealed in flexible polyethylene pouches and treated at 200, 300, 400, 500, and 600 MPa for 5 min at 25 °C, with untreated juice as control. After treatment, pouches were stored at 4 ± 1 °C. Analyses at days 0, 7, 14, 21, and 28 included: total phenolic content (Folin-Ciocalteu, GAE), total anthocyanin content (pH differential method), vitamin C (DCPIP titration), DPPH radical scavenging (%), colour (L*, a*, b* by HunterLab), total plate count (pour plate, PCA, 37 °C, 48 h), and yeast/mould count (PDA, 25 °C, 5 days) [6] . All analyses in triplicate. Data were analysed by two-way ANOVA (pressure × storage) and Duncan's test (p<0.05) in SPSS v. 26.

Results

HPP at all pressure levels reduced microbial load relative to the untreated control. Bioactive retention decreased with increasing pressure, but the decline was modest up to 400 MPa and steepened markedly above 500 MPa. Comprehensive Interpretation HPP at 400 MPa achieved a 2.9-log microbial reductionsufficient for a 28-day refrigerated shelf life-while retaining over 90% of all measured bioactives. At 600 MPa, microbial kill increased to 4.2 log but vitamin C dropped to 79.2% of control values. Colour retention during storage was markedly better in all HPP-treated juices than in the untreated control, which lost 53.8% of its redness (a*) by day 28 compared with just 17.7% loss in the 400 MPa treatment.

Discussion

The 400 MPa optimum identified here aligns closely with findings by Ferrari et al. [4] , who reported 400 MPa / 5 min as the threshold for commercial sterility in pomegranate juice. The literature agrees that anthocyanin degradation under HPP follows a pressure-dependent first-order kinetic model, with the rate constant rising sharply above 500 MPa [7] . The greater sensitivity of vitamin C relative to anthocyanins can be explained by the oxidative pathway: pressure increases the solubility of dissolved oxygen, which accelerates ascorbic acid oxidation even at ambient temperature [8] .

Where this research adds to existing knowledge is in the 28day colour-stability data. Prior HPP trials on pomegranate rarely tracked colour beyond day 14 [4] . The present results show that the 400 MPa treatment maintained 82.3% colour retention at day 28, far superior to the untreated control

International Journal of Agriculture and Food Science https://www.agriculturaljournals.com (46.2%). This advantage likely arises from HPP inactivation of polyphenol oxidase and peroxidase, enzymes that catalyse anthocyanin browning during storage [9] . From a commercial standpoint, 400 MPa / 5 min is well within the operating range of existing industrial HPP units (up to 600 MPa), and the resulting juice would meet the microbial criteria (≤ 2 log CFU/mL) set by most coldpressed juice brands for a 30-day refrigerated shelf life [10] .

Conclusion

This research set out to determine the HPP pressure that best balances microbial safety with bioactive retention in pomegranate juice. Treatment at 400 MPa for 5 min retained over 90% of anthocyanins, TPC, and vitamin C while achieving a 2.9-log microbial reduction and maintaining colour stability for 28 days at 4 °C. Higher pressures (500-600 MPa) gave greater microbial kill but at the cost of meaningful vitamin C loss. For pomegranate juice processors, 400 MPa represents the sweet spot between safety and quality. Future work should validate this finding at pilot scale with commercial HPP equipment, assess consumer willingness-to-pay for HPP vs thermally pasteurised juice, and evaluate the fate of ellagitanninspomegranate's most bioactive polyphenol class-under HPP conditions.

References

1. Alper N, Bahceci KS, Acar J. Influence of processing and pasteurisation on colour values and total phenolics of pomegranate juice. Journal of Food Processing and Preservation. 2005;29(5-6):357-368.
2. Huang HW, Lung HM, Yang BB, Wang CY. Responses of microorganisms to high hydrostatic pressure processing. Food Control. 2014;40:250-259.
3. Gil MI, Tomás-Barberán FA, Hess-Pierce B, Holcroft DM, Kader AA. Antioxidant activity of pomegranate juice and its relationship with phenolic composition and processing. Journal of Agricultural and Food Chemistry. 2000;48(10):4581-4589.
4. Ferrari G, Maresca P, Ciccarone R. The application of high hydrostatic pressure for the stabilisation of functional foods: pomegranate juice. Journal of Food Engineering. 2010;100(2):245-253.
5. AOAC. Official methods of analysis. 20th ed. AOAC International, Rockville, MD. 2016.
6. Giusti MM, Wrolstad RE. Characterization and measurement of anthocyanins by UV-visible spectroscopy. Current Protocols in Food Analytical Chemistry. 2001;F1.2.1-F1.2.13.
7. Corrales M, García AF, Butz P, Tauscher B. Extraction of anthocyanins from grape skins assisted by high hydrostatic pressure. Journal of Food Engineering. 2009;90(4):415-421.
8. Oey I, Lille M, Van Loey A, Hendrickx M. Effect of high-pressure processing on colour, texture and flavour of fruit-and vegetable-based food products: a review. Trends in Food Science and Technology. 2008;19(6):320-328.
9. Terefe NS, Yang YH, Knoerzer K. High pressure and thermal inactivation kinetics of polyphenol oxidase and peroxidase in strawberry puree. Innovative Food Science and Emerging Technologies. 2010;11(1):52-60.
10. Balasubramaniam VM, Martínez-Monteagudo SI, Gupta R. Principles and application of high pressurebased technologies in the food industry. Annual Review of Food Science and Technology. 2015;6:435-462.
11. Calligaris S, Foschia M, Bartolomeoli I, Maifreni M, Manzocco L. Study on the applicability of highpressure homogenization for the production of banana juices. LWT-Food Science and Technology. 2012;45(1):117-121.
12. Tiwari BK, O'Donnell CP, Patras A, Brunton N, Cullen PJ. Anthocyanins and colour degradation in ozonetreated blackberry juice. Food and Chemical Toxicology. 2009;47(11):2824-2829.
13. Viuda-Martos M, Fernández-López J, Pérez-Álvarez JA. Pomegranate and its many functional components as related to human health: a review. Comprehensive Reviews in Food Science and Food Safety. 2010;9(6):635-654.
14. Varela-Santos E, Ochoa-Martínez A, Tabilo-Munizaga G. Effect of high hydrostatic pressure on the quality of two Pacific cold-water fish species. Innovative Food Science and Emerging Technologies. 2012;15:29-36.
15. Dede S, Alpas H, Bayindirli A. High hydrostatic pressure treatment and storage of carrot and tomato juices. European Food Research and Technology. 2007;225(5-6):899-905.
16. Keenan DF, Brunton NP, Gormley TR, Butler F. Effects of thermal and high hydrostatic pressure processing on antioxidant activity and colour of fruit smoothies. Innovative Food Science and Emerging Technologies. 2010;11(4):551-556.
17. Barba FJ, Esteve MJ, Frígola A. High pressure treatment effect on physicochemical and nutritional properties of fluid foods during storage. Comprehensive Reviews in Food Science and Food Safety. 2012;11(3):307-322.
18. Landl A, Abadias M, Sárraga C, Viñas I, Picouet PA. Effect of high pressure processing on the quality of acidified Granny Smith apple purée product. Innovative Food Science and Emerging Technologies. 2010;11(4):557-564.