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. 2025 Jun 25;14(13):2245.
doi: 10.3390/foods14132245.

Biorefining of Walnut Shells into Polyphenol-Rich Extracts Using Ultrasound-Assisted, Enzyme-Assisted, and Pressurized Liquid Extraction Coupled with Chemometrics

Busra Acoglu Celik  1 , Muhammed Alpgiray Celik  2 , Laura Jūrienė  3 , Jovita Jovaišaitė  3 , Rita Kazernavičiūtė  3 , Erturk Bekar  1 , Perihan Yolci Omeroglu  1 , Petras Rimantas Venskutonis  3 , Senem Kamiloglu  1
Affiliations

Biorefining of Walnut Shells into Polyphenol-Rich Extracts Using Ultrasound-Assisted, Enzyme-Assisted, and Pressurized Liquid Extraction Coupled with Chemometrics

Busra Acoglu Celik et al. Foods. .

Abstract

Walnut (Juglans regia L.) shells are valuable agro-industrial by-products rich in polyphenols. This study investigated traditional (maceration) and advanced extraction techniques-ultrasound-assisted extraction (UAE), enzyme-assisted extraction (EAE), pressurized liquid extraction (PLE), and combined ultrasound-enzyme extraction (US-EAE)-to recover bioactive compounds from walnut shells. Extraction efficiency, total phenolic content (TPC), antioxidant capacity (ABTS•+, DPPH•), and polyphenol composition were evaluated. UPLC-ESI-MS/MS identified key polyphenols including ellagic acid, 4-hydroxybenzoic acid, vanillin, taxifolin, and quercitrin. The highest TPC (5625 mg GAE/100 g dw) was found in extracts subjected to US-EAE, in which ultrasound pretreatment (200 W, 10 min) was followed by enzymatic extraction using 0.06 mL/g Viscozyme® L at pH 3.5 and 45 °C. Under the same extraction conditions, UAE alone yielded the second highest TPC (4129 mg GAE/100 g dw). The highest ABTS•+ scavenging activity (14,478 mg TE/100 g dw) and enhanced DPPH• activity (45.38 mg TE/100 g dw) were also observed in US-EAE extracts. Chemometric techniques (PCA and HCA) revealed meaningful clustering and variation patterns among methods. These findings highlight the potential of walnut shells as a sustainable source of polyphenols and demonstrate the effectiveness of innovative extraction technologies in maximizing bioactive compound recovery for potential functional applications.

Keywords: Juglans regia L. shell; antioxidant capacity; combined extraction; green extraction; phenolic compounds; principal component analysis.

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Conflict of interest statement

The authors declare no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Figures

Figure 1
Figure 1
(a) Walnut shell, (b) walnut shell powder, (c) PLE-EtOH, (d) PLE-AC, (e) EAE, (f) UAE, (g) US-EAE, (h) M-Water, (i) M-EtOH extractions.
Figure 2
Figure 2
Pearson correlation analyses.
Figure 3
Figure 3
Principal component analysis (PCA) loading plot of walnut shell extracts.
Figure 4
Figure 4
Principal component analysis (PCA) score plot of walnut shell extracts.
Figure 5
Figure 5
Cluster map of walnut shell extracts.
See this image and copyright information in PMC

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