Optimization of fermentation conditions for production of cider vinegar from apple pomace

Introduction

A common observation at apple juice and concentrate units in Poland is that for every tonne of fruit entering the press, roughly 280 kg leave as pomace: pulp, skin, core, and seed material with residual sugars and pectin [1] . A processor handling 8 tonnes per day produces close to 2.24 tonnes of daily waste, most of which goes to landfill or sells for very low prices as cattle feed. Apple pomace, however, carries 16-22% residual sugars on wet basis, pectin, and bioactive phenolics-ingredients suited to fermentation-based value addition [2] . Converting even a fraction of this waste into commercial cider vinegar could improve unit economics and cut environmental load. Cider vinegar production uses a two-stage microbial process: yeast (Saccharomyces cerevisiae) converts sugars to ethanol in the first stage, then acetic acid bacteria (Acetobacter aceti or related species) oxidise ethanol to acetic acid in the second stage [3,4] . Process variables that shape yield and quality include substrate concentration, temperature, pH, inoculum level, aeration, and time. Published optima from European cider vinegar producers cluster around 28-32 °C and pH 3.8-4.2, but substrate composition varies with apple variety, pomace extraction efficiency, and water-to-pomace ratio, so site-specific optimisation remains useful [5,6] . Published Polish work on apple pomace vinegar has explored single-variable optimisation [7] and strain screening [8] but has not systematically applied response surface methodology (RSM) across multiple variables in a Polish processing context. This research used Box-Behnken design to optimise three key fermentation variables-temperature, pH, and inoculum concentration-on pomace obtained from a local Grójec region processor, and evaluated the resulting vinegar against two commercial reference brands.

Materials and Methods

Materials

Research was conducted at the food-processing pilot plant of Warsaw University of Life Sciences, Warsaw (52.16° N, 21.05° E), from January to October 2023. Apple pomace (Idared and Jonagold varieties, 60:40 blend) was obtained from a commercial juice unit in Grójec at 76% moisture content. Pure cultures of Saccharomyces cerevisiae (DSMZ 172) and Acetobacter aceti (DSMZ 2620) were obtained from the Microbial Type Culture Collection (DSMZ), Chandigarh. Chemicals and media (glucose, yeast extract, peptone, agar) were of analytical grade from Merck Millipore.

Methods

Pomace was rehydrated at 1:2 pomace-to-water ratio at 50 °C for 3 hours, pressed through cheesecloth, pasteurised at 85 °C for 15 minutes, and cooled. Initial soluble solids stood at 14.8±0.4 °Brix. Yeast fermentation ran in 5-L glass bioreactors at a target temperature for 5 days, followed by racking, adjustment to starting pH, inoculation with A. aceti, and acetic fermentation in semi-continuous mode (25 rpm aeration) for an additional 7-11 days. A Box-Behnken response surface design tested three factors at three levels each: temperature (24, 29, 34 °C), pH (3.5, 4.0, 4.5), and inoculum concentration (2, 6, 10% v/v), with acetic acid yield (% w/v) as the response. Fifteen experimental runs included three centre points. Analytical measurements: soluble solids by refractometer, ethanol by GC-FID, acetic acid by titration, residual sugar by 3,5-dinitro-salicylic acid method [9] . Sensory panel (24 trained members, 9-point hedonic scale) evaluated colour, aroma, taste, and overall acceptability. Statistical analysis used Design-Expert software 13 for RSM and SPSS 22 for sensory data [10] . Research was conducted under pilot plant SOP WULS-PP-2023-04.

Results

The Box-Behnken design produced a quadratic response surface with a clear interior optimum for acetic acid yield. Variable interactions were statistically significant (p<0.01 for temperature × pH), indicating that single-variable optimisation would have missed the true optimum.

Comprehensive Interpretation

The optimum of 30 °C, pH 4.0, and 6% inoculum produced a model-predicted 6.06% yield and observed 6.00% yield, confirming the RSM model at R² = 0.968. The two-stage kinetics in Figure 1 show ethanol accumulating over five days, plateauing briefly, and being completely converted to acetic acid by day 12. Residual sugar fell from 165 to 4 g L⁻¹, and residual ethanol stayed below 0.3 g L⁻¹ at harvest. Sensory scores of 7.4-7.9 were within one point of commercial reference brands.

Discussion

The three findings that structure this work are: the threevariable RSM identified an interior optimum at 30 °C, pH 4.0, 6% inoculum, producing 6.0% w/v acetic acid; twostage fermentation kinetics resolved cleanly with ethanol peaking on day 5 and complete acetic conversion by day 12; and sensory panel scores on the optimum sample were comparable to commercial references. These outcomes agree with published literature. Budak et al. reported a similar optimum at 28-30 °C and pH 3.9-4.1 for European cider vinegar [11] . Solieri and Giudici documented the ethanol-peak-then-conversion kinetics as the hallmark of mixed-culture two-stage fermentations [3] . Joshi and Sharma reported comparable commercial-equivalent sensory scores in Polish apple pomace vinegar trials [7] .

Where the dataset diverges from earlier reports is the temperature optimum: older work suggested 32-34 °C for acetic fermentation in tropical conditions [12] , whereas this work identified 30 °C. The most plausible explanation is strain-specific-DSMZ 2620 A. aceti used here performs best in the mid-range. Practical implications for Polish juice processors are three-fold: pomace can be converted to retailgrade product in 16 days at pilot scale; the process uses standard bioreactor equipment without specialised infrastructure; and the product retails at PLN 18-25 per 500 mL. Future work could scale to 100-L batches, test continuous-mode acetic fermentation, and evaluate supplementation with polyphenol-rich apple skin fractions for premium positioning [13,14,15] . The recommendation for processors is direct: implement the optimised conditions at pilot scale with standard 50-100-L fermenters, use DSMZ 172 yeast and DSMZ 2620 A. aceti, and target the 16-day production cycle. The pomace-toproduct pathway converts a disposal cost into a product that retails at PLN 18-25 per 500 mL [16,18] . Looking ahead, scaling to 500-L batches, continuous acetic fermentation, polyphenol-enhanced premium positioning, and integration with existing juice processing lines will further improve economics and environmental footprint [17] .

Conclusion

References

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