Exploring new applications of toasted chestnut shells as an oak alternative in distilled spirit aging

Abstract

This study investigated the potential of toasted chestnut shells as a novel and cost-effective material for aging distilled spirits. Results showed that aging whiskey with toasted chestnut shell chips moderately increased its acidity (0.14–0.18 g/L) and total phenolic content (48.28–109.58 mg/L), while imparting a color ranging from yellowish to light amber. HS-SPME-GC-MS, HS-GC-IMS, E-nose, and sensory evaluation analyses revealed the distinct aroma characteristics of whiskeys aged with toasted chestnut shell chips. Compared to whiskeys aged with toasted oak, which contained abundant ethyl esters and exhibited an obvious fruity aroma, aging whiskeys with toasted chestnut shell chips significantly increased the level of aldehydes while decreasing the levels of ethyl esters and acetate esters. 2-methyl butanal was identified as the key aroma compound that accounts for the pronounced roasted/smoky aroma in these whiskeys. This study provides valuable insights into the conversion of agro-industrial by-product into new oak alternatives in the oenological field.

Introduction

The chestnut, belonging to the Fagaceae family, is considered a significant tree in the agricultural and forestry economy¹. For centuries, chestnut seeds have been one of the most important food resources in North America, Europe, and East Asia². According to the Food and Agriculture Organization Corporate Statistical Database (FAOSTAT), China produced 1.52 million tons of chestnuts in 2023, accounting for more than 70% of the global total production³. The steady development of the chestnut sector has, on the one hand, greatly promoted the economy of China and, on the other hand, generated a large number of by-products, primarily shells. Chestnut shells, accounting for approximately 20% of the whole fruit by fresh weight, are generated as a byproduct during chestnut processing⁴. Due to their limited industrial or commercial applications, these shells are predominantly burned or discarded, undoubtedly leading to significant environmental concerns⁵. Therefore, it is crucial to develop effective approaches and strategies to reevaluate and utilize chestnut shells.

In recent years, numerous researchers have evaluated the nutritive value of chestnut shells and revealed their rich content of dietary fiber (hemicellulose, cellulose, and lignin), phenolic acids, flavonoids, and tannins^4,6. From the perspective of bioactive compounds, these primary and secondary plant metabolites have been proven to have anti-inflammatory, antimicrobial, antioxidant, hypoglycemic, hypolipidemic, and neuroprotective properties^4,7–12. Therefore, adding chestnut shells or their extracts as a nutritional supplement to food delivers outstanding health-promoting effects^13–15. From the perspective of flavor compounds, notably, these metabolites and/or their thermochemical reaction products may produce color, aroma, and taste characteristics that are essential for the quality of alcoholic beverages, such as whiskey¹⁶. Typically, these flavor compounds in distilled spirits can be obtained through aging with toasted oak woods^17–19. Although several materials (e.g., acacia, chestnut, cherry, ash, mulberry, and vine-shoot) can serve as oak alternatives for aging distilled spirits^20–22, chestnut shells have not, to our knowledge, been tested for this purpose. In view of this, we hypothesize that chestnut shells can develop distinct flavors when toasted at varying temperatures, and adding these toasted chestnut shells during the aging stage of distilled spirits may have a positive impact on their flavors.

Aroma is a critical sensory attribute of distilled spirits²³. Traditionally, the evaluation of this characteristic has predominantly relied on olfactory assessments, which require extensive expertise and frequently result in subjective outcomes²⁴. The electronic nose (E-nose), designed to mimic the human olfactory system, enables odor samples to interact with an array of internal sensors and outputs digital signals that represent the overall odor intensity²⁵. Since the signal output is typically a composite response of multiple compounds, the E-nose is unable to identify key aroma compounds or evaluate their contributions to the overall aroma profiles²⁶. Headspace solid-phase microextraction combined with gas chromatography-mass spectrometry (HS-SPME-GC-MS) is a commonly used technique for analyzing volatile compounds in distilled spirits because of its powerful qualitative and quantitative capabilities²⁷. However, this method requires a lengthy extraction process, which is very likely to lead to distortion of the results as well as reducing the efficiency of sample analysis²⁸. Nowadays, headspace combined with gas chromatography-ion mobility spectrometry (HS-GC-IMS), a novel analytical technique, provides a simplified pretreatment procedure, enables rapid and highly sensitive detection of volatile compounds, and facilitates the visualization and comparison of volatile compositions across different samples²⁹. However, the application of this method in the quantitative analysis of volatile compounds is limited due to the lack of a complete database²⁴. Considering the distinct characteristics of olfactory assessment, E-nose, HS-SPME-GC-MS, and HS-GC-IMS analytical methods, numerous studies have demonstrated that integrating these techniques enables both effective and comprehensive analysis of volatile compounds in alcoholic beverages^26,30–32.

Therefore, this study aims to evaluate the impact of aging with chestnut shell chips of varying toasting degrees on the aroma characteristics of whiskeys using HS-SPME-GC-MS, and HS-GC-IMS, E-nose, and artificial sensory analyses, and to compare these characteristics with those of whiskeys aged with toasted oak chips. The findings of this study could provide valuable insights into the utilization of chestnut shells as a novel oak alternative in the oenological field, and also contribute to the promotion of agro-industrial sustainability.

Results and discussion

Oenological parameters analysis

The oenological parameters of whiskeys aged with toasted chestnut shell chips and toasted oak chips were presented in Table 1. Compared to the control sample (0.11 g/L and 5.29), the whiskeys aged with toasted chestnut shell chips and toasted oak chips showed significantly higher titratable acidity levels (0.14 to 0.28 g/L) and, consequently, lower pH values (3.92 to 5.20). For alcohol content and volatile acidity of these whiskeys, neither aging with toasted chestnut shell chips nor aging with toasted oak chips significantly affects them (p > 0.05). This finding is consistent with reports by Duan et al.²¹, who observed similar results in spine grape brandies aged with various types of toasted wood chips. This can be explained by the fact that these whiskeys were produced through the same fermentation and distillation processes and aged under similar conditions. With respect to total phenolic content (5.03 mg/L), it was significantly enhanced after aging with toasted chestnut shell chips and toasted oak chips (48.28 to 427.61 mg/L), which was mainly due to the release of phenolic compounds from these chips. In fact, numerous studies have reported that both chestnut shells and oak wood are rich in phenolic compounds^6,7,33,34.

Table 1.

Oenological parameters of whiskeys aged with toasted chestnut shell chips and toasted oak chips

Parameters	Control	LS	MS	HS	LO	MO	HO
Pictures
Alcohol content (% vol)	62.35 ± 0.08	62.24 ± 0.12	62.32 ± 0.10	62.35 ± 0.06	62.26 ± 0.13	62.25 ± 0.07	62.29 ± 0.10
Titratable acidity (g acetic acid /L)	0.11 ± 0.02 d	0.18 ± 0.01b	0.18 ± 0.01b	0.14 ± 0.01c	0.20 ± 0.01b	0.28 ± 0.02a	0.18 ± 0.01b
Volatile acidity (g acetic acid /L)	0.11 ± 0.01	0.11 ± 0.01	0.12 ± 0.01	0.11 ± 0.01	0.11 ± 0.01	0.12 ± 0.01	0.11 ± 0.01
pH	5.29 ± 0.03a	4.97 ± 0.02 d	4.91 ± 0.02c	5.20 ± 0.03b	4.13 ± 0.03 f	3.92 ± 0.02 g	4.31 ± 0.02e
Total phenolic content (mg gallic acid /L)	5.03 ± 0.17a	106.26 ± 0.54 d	109.58 ± 0.41c	48.28 ± 0.17e	120.68 ± 0.71c	427.61 ± 3.80a	294.26 ± 3.24b
L*	97.05 ± 0.34a	90.58 ± 0.32c	90.12 ± 0.28c	94.48 ± 0.35b	94.13 ± 0.34b	84.37 ± 0.22e	87.05 ± 0.33 d
a*	−0.21 ± 0.06 d	1.50 ± 0.10b	-0.15 ± 0.06 d	−1.41 ± 0.11e	−1.65 ± 0.03 f	2.99 ± 0.12a	0.53 ± 0.07c
b*	0.37 ± 0.10e	21.34 ± 0.27 d	27.07 ± 0.26c	12.01 ± 0.25 f	14.80 ± 0.29e	55.15 ± 0.24a	45.52 ± 0.24b
ΔE	0.00	22.01 ± 0.19 d	27.58 ± 0.17c	11.99 ± 0.18 f	14.80 ± 0.27e	56.32 ± 0.33a	46.25 ± 0.30b

The values are expressed as mean ± standard deviation. The values followed by different letters in a row are significantly different (P < 0.05) by Duncan test.

After aging with two types of chips, the whiskeys present a mature hue that ranges from yellowish to amber. To be specific, both toasted chestnut shell chips aging and toasted oak chips aging significantly reduced the L* values and increased the b* values of the whiskeys. Additionally, aging with these chips also moderately affect the a* values of the whiskeys. Due to the relatively minor variations in these parameters, the whiskeys aged with toasted chestnut shell chips exhibited generally lower ΔE values (11.99 to 27.58), particularly that aged with HS chips. Conversely, the whiskeys aged with toasted oak chips demonstrated significantly higher overall ΔE values (14.80 to 56.32), especially those aged with MO and HO chips. These findings suggest that the compositional difference between chestnut shells and oak woods might be the primary reason for the significant color variation in whiskies aged with them. This is because such a difference not only causes variations in the Maillard reaction and thermal degradation products during roasting at different temperatures^35,36, but also gives rise to differences in the new pigments formed through condensation and oxidation reactions during the aging process^37,38. Furthermore, the differences in ΔE values among whiskeys aged with toasted chestnut shell chips and toasted oak chips may lead to a distinction in their expected fruity flavor, because tasters do associate deeper red-yellow hues of the beverage as ripening fruits^39,40.

E-nose analysis

To gain an initial understanding of the differences in volatiles between whiskeys aged with toasted chestnut shell chips and those aged with toasted oak chips, the E-nose was used to analyze the overall aroma profiles of these samples. As shown in Fig. 1a, compared to the control sample, the overall aroma profiles of whiskey were significantly altered after aging with either toasted chestnut shell chips or toasted oak chips, as evidenced by the variations in E-nose sensors response values. For the aged whiskeys, the elevated response values in the W1W, W2W, W1S, and W2S sensors and the decreased response values in the W5S and W6S sensors suggest that they may contain a higher concentration of sulfur-containing compounds, alcohols, and ethers, while having a lower concentration of nitrogen oxide compounds and hydrogen compounds. For the majority of sensors, comparable response values were observed between whiskeys aged with chestnut shell chips and oak chips of the same toasting degree. However, an obviously higher response value in the W1S sensor was found for whiskey aged with toasted chestnut shell chips. This phenomenon can be attributed to the distinct pyrolysis products formed by the different components in chestnut shells and oak during the toasting process. Additionally, the formation of these compounds in chestnut shells during the toasting process may be temperature-dependent, as evidenced by the increasing response value in the W1S sensor with the higher toasting degree. For the W3S sensor, its response value remained essentially unchanged before and after aging, which suggests that the aging of toasted chestnut shell chips and toasted oak chips may not significantly affect the content of ammonia and aromatic components in whiskey.

Fig. 1. Analysis of volatile compounds in whiskeys by E-nose.

a Radar chart of the E-nose sensors response data. b PCA biplot graph for the E-nose analysis.

To better classify the differences in overall aroma profiles between whiskeys aged with toasted chestnut shell chips and those aged with toasted oak chips, principal component analysis (PCA) was conducted using the E-nose sensor response values. As shown in Fig. 1b, the first two principal components explained 88.9% of the total variance, with PC1 of 73.9% and PC2 of 15.0%, respectively. The control sample and the aged samples were located in the opposite direction of PC1, which was consistent with the variations in E-nose sensors response values. This tendency was consistent with previous research conducted by Petrozziello et al.⁴¹. on marc distillates aged with toasted oak and poplar woods. For those aged whiskeys, differentiation was observed along PC2. Specifically, the whiskeys aged with toasted chestnut shell chips were distributed in the negative direction of PC2, while those aged with toasted oak chips were positioned in the positive direction of PC2. Such fact indicates the differences in volatile compositions between the whiskies aged with toasted chestnut shell chips and those aged toasted oak chips. To achieve an in-depth understanding of the differences in volatile compositions among these whiskeys, the volatile compounds were further analyzed using HS-SPME-GC-MS and HS-GC-IMS techniques.

Identification of volatiles in whiskeys by HS-SPME-GC-MS

To investigate the differences in volatile composition between whiskeys aged with toasted chestnut shell chips and those aged with toasted oak chips, HS-SPME-GC-MS was used to analyze the volatile compounds. A total of 65 volatile compounds were identified in all whiskeys via HS-SPME-GC-MS, including 15 higher alcohols, 17 esters (12 ethyl esters, 3 acetate esters, and 2 other esters), 14 carbonyl compounds (7 aldehydes, 2 ketones, and 5 acetals), 3 organic acids, 6 aromatic compounds, 2 phenols, 3 oxygenous heterocyclic compounds, and 5 terpenes. As shown in Supplementary Table 2 and Fig. 3, compared to the control sample, significant variations were observed in the volatile compositions of whiskeys aged with toasted chestnut shell chips, which were also different from those aged with toasted oak chips.

Fig. 3. Analysis of volatile compounds in whiskeys identified by HS-GC-IMS.

a The 2D topographic plots of volatile compounds. b The 2D difference comparison topographic plots of volatile compounds. c Fingerprints of volatile compounds. d Heatmap of volatile compound contents. e Concentrations of each group of volatile compounds. f PCA biplot graph for the HS-GC-IMS analysis (the volatile compounds represented by the numbers presented in Supplementary Table 3).

Higher alcohols are the major qualitative and quantitative components that detected in all whiskeys (from 241.66 to 263.81 mg/L). Compared with those in the control sample, the concentrations of 1-octen-3-ol and 2-ethyl-1-hexanol were increased after aging, particularly in whiskeys aged with MO, HO, MS, and HS chips. These results may be attributed to the extraction of certain higher alcohols from toasted chestnut shell chips and toasted oak chips, which is consistent with a few previous studies that have identified these compounds in both oak and chestnut woods^42–44. Furthermore, the possible adsorption of specific higher alcohols by the toasted chestnut shell chips could explain the decline of (Z)-3-Nonen-1-ol in whiskeys aged with LS, MS, and HS chips. However, for the majority of individual higher alcohols, their concentrations relatively unchanged after aging with toasted chestnut shell chips and toasted oak chips. This is mainly because the higher alcohols in these whiskeys are created and concentrated during fermentation and distillation^45,46, whereas a short-term aging process (30 days) may not significantly affect their concentrations.

Esters, including ethyl esters, acetate esters, and other esters, were the second most abundant compounds detected in all whiskeys (from 43.33 to 53.89 mg/L). In comparison with those in the control sample, ethyl 2-furancarboxylate and ethyl benzoate were the only compounds that showed increased concentrations after aging with toasted chestnut shell chips. In contrast, the concentrations of most individual ethyl esters decreased following aging with these chips. Notably, a significant increase in the concentrations of most ethyl esters was observed in whiskeys aged with toasted oak chips, which agree with the findings of Maria Balcerek et al.¹⁷. in Plum Distillate and Nie et al. ¹⁸. in persimmon brandy. However, for acetate esters, neither toasted chestnut shell chips aging nor toasted oak chips aging produces any significant effect on their concentrations. With respect to the two other esters: methyl thiolacetate and methyl 2-furoate, the latter compound was exclusively detected in whiskeys aged with toasted chestnut shell chips, and the usage of MS chips for aging demonstrated the most significant enhancement effect (Fig. 2a). For methyl thiolacetate, this compound was found in all whiskeys, and its concentration was decreased by both toasted chestnut shell chips aging or toasted oak chips aging (Fig. 2a). This phenomenon could be associated with the adsorption capacity of these aging materials, as the absorption ability of chestnut shells and toasted oak wood have been confirmed by many studies^47–49.

Fig. 2. Analysis of volatile compounds in whiskeys identified by HS-SPME-GC-MS.

a Heatmap of volatile compound contents. b Concentrations of each group of volatile compounds. c PCA biplot graph for the HS-SPME-GC-MS analysis (the volatile compounds represented by the numbers presented in Supplementary Table 2).

Aldehydes, ketones, and acetals, known as carbonyl compounds, are reported to contribute to the flavor of alcoholic beverages⁵⁰. Acetaldehyde is the most representative aldehyde in distilled spirits, and its concentration usually increases during the aging process due to the oxidation of ethanol²³. However, in comparison with the control sample, a decrease in its concentration was observed in all aged whiskeys. This phenomenon could be associated with the absorption effect of toasted chestnut shell chips and toasted oak chips. In addition, some antioxidant substances (e.g., phenols) existing in these chips may delay the oxidation of ethanol, resulting in the lower concentration of acetaldehyde in these aged whiskeys. For hexanal, octanal, and nonanal, their concentrations were increased to varying degrees after aging, typically in whiskeys aged with HO chips, which reflects that the formation of these compounds may be material and temperature-dependent.

Among the two ketones (acetone and 2,6-dimethyl-4-heptanone) quantified in this study, a significant reduction in the levels of 2,6-dimethyl-4-heptanone was observed in whiskeys aged with toasted chestnut shell chips. However, aging with toasted oak chips did not produce any significant effect on their levels.

In alcoholic beverages, acetals can be formed via the consecutive reaction of aldehydes with alcohols⁴⁶. As shown in Fig. 2a, both chestnut shell chips aging and toasted oak chips aging produce a significant enhancement in the total and individual concentrations of acetals in whiskeys. A Similar observation has been noticed by Maria et al.¹⁷. in oak chips aged plum distillate. We suspect that this phenomenon may be attributed to the catalytic effects of toasted chestnut shell chips and toasted oak chips on acetal formation.

The organic acids detected in this work are 2-methyl propanoic acid, hexanoic acid, and octanoic acid, and their concentrations all increased after aging with toasted chestnut shell chips and toasted oak chips (Fig. 2a). It is worth noting that toasted chestnut shell chips exhibited a more pronounced enhancement effect on the levels of hexanoic acid and octanoic acid compared to toasted oak chips during the aging process. Although the specific mechanism requires further exploration, the oxidation of hexanal and octanal during the toasting process could be one of the reasons resulting in the higher levels of hexanoic acid and octanoic acid in toasted chestnut shell chips, thereby significantly elevating their concentrations in whiskey after aging with these chips.

Regarding the aromatic compounds detected in this work, compared to the control samples, the concentration of benzaldehyde increased after aging with toasted chestnut shell chips and toasted oak chips. Conversely, the concentrations of 1,2,3-Trimethylbenzene and (2,2-Diethoxyethyl)-benzene decreased. However, for benzene acetaldehyde and naphthalene, opposite effects were observed between whiskeys aged with these two types of chips. In terms of 2-methylbenzofuran and guaiacol, these compounds were exclusively detected in whiskey aged with HS chips. This fact may relate to the presence of certain precursors, such as p-coumaric acid and ferulic acid, in chestnut shells⁵¹. These precursors could undergo thermal degradation to form guaiacol when the chestnut shells are roasted at higher temperatures^52,53.

For oxygenous heterocyclic compounds, furfural and 5-methyl-2-furancarboxaldehyde, their concentrations all increased after aging, notably in whiskeys aged with MS and MO chips. These findings are in agreement with those of Lu et al.⁵⁴, who reported that the ‘Merlot’ dry red wines aged in medium-toasted barrels contained higher levels of furanic compounds. As typical thermal dehydration products of carbohydrates, these compounds are abundantly present in roasted wooden materials, which has been confirmed by numerous studies^55,56.

Terpenes usually have a crucial influence on the overall aroma profiles of alcoholic beverages, and they typically originate from three potential sources: raw materials, microorganisms, and maturation materials^57–59. In this work, four monoterpenes and one norisoprenoid were detected in all whiskeys, and their concentrations were significantly lower in whiskeys aged with toasted chestnut shell chips compared to those in the control sample. This reduction may also be attributed to the adsorption effect of the toasted chestnut shells, as discussed in the previous section. Although previous studies demonstrated the presence of a variety of terpenes^19,60, such as α-terpineol, in oak woods, the obtained results indicated that aging with toasted oak chips did not contribute positively to their concentrations in whiskey.

To better understand the difference in volatile compositions among whiskeys aged with toasted chestnut shell chips and toasted oak chips, PCA was carried out using the concentrations of all volatiles as variables (Supplementary Table 2). The first two principal components accounted for a combined variance of 56.0%, with PC1 and PC2 explaining 38.1% and 17.9% of the variance, respectively. The results presented in Fig. 2c demonstrated a clear distinction between the aged and control whiskeys, which was attributed to the differences of numerous compounds with heavy loadings in the positive directions of PC2. For the aged whiskeys, the high loading values obtained for OE2, ALD6, ALD5, AC2, ACE1, AC3, OA3, and OA2 in the negative direction of PC1 were the main contributors to those aged with toasted chestnut shell chips. In contrast, AC5, EE8, KET2, AC1, EE5, EE2, TP5, TP1, HA15, TP4, OE1, TP3, EE6, EE3, TP2, EE7, EE10, and EE12 in the positive direction of PC1 corresponded to those aged with toasted oak chips. In addition, EE11, ACE4, ACE2, OA1, ACE5, HA8, OHC2, ALD4, OHC1, ALD1, which were positively correlated with PC2, were mainly associated with the whiskeys aged with two types of chips that have different toasting degrees.

Screen the key volatile flavor compounds from the volatiles in whiskeys identified by HS-SPME-GC-MS

To better understand the differences in overall aroma characteristics among the whiskeys aged with toasted chestnut shell chips and those aged with toasted oak chips, the variable importance in projection (VIP) values and the relative odor activity values (ROAV) were combined to identify the key volatile flavor compounds and assess their contributions to the overall aroma profile of these samples. The aroma descriptions and aroma types of volatile compounds with VIP ≥ 1.0 and ROAV ≥ 0.1 in these whiskeys are shown in Table 3. Among ten volatiles with VIP ≥ 1.0 and ROAV ≥ 1.0, five volatiles were existed in the control sample, namely: ethyl butyrate, ethyl hexanoate, ethyl octanoate, linalool, and β-damascenone. In addition to whiskey^16,61, these compounds have also been identified as key volatile flavor compounds in other types of distilled spirits^21,23. As expected, the control sample was dominated by fruity and floral aroma characteristics, and showed the lowest intensities of herbaceous, fatty, and roasted aroma characteristics. After aging with toasted chestnut shell chips, the intensities of roasted/smoky, herbaceous, and fatty aroma characteristics in whiskeys were obviously elevated, which was mainly related to the increased concentrations of 2-methyl butanal, guaiacol, hexanal, octanal, and nonanal in these samples. For whiskeys aged with toasted oak chips, the intensity of fruity aroma characteristic was further strengthened, which was mainly ascribed to the elevated levels of ethyl hexanoate and ethyl octanoate in these samples. In addition, for the volatile compounds with ROAV between 0.1 and 1.0, higher intensity values were also observed in whiskeys aged with toasted chestnut shell chips, which could potentially enhance the complexity of these whiskeys through auxiliary effects⁶².

Table 3.

Volatiles with VIPs ≥ 1.0 and ROAV ≥ 0.1 in whiskeys identified by HS-GC-IMS

No.	Compounds	Odor threshold (μg/L)	Aroma description	Aroma type	VIP	ROAV
						Control	LS	MS	HS	LO	MO	HO
ha4	(Z)-3-nonen-1-ol	1067	Green, fatty72	Herbaceous, Fatty	1.50	6.82	6.93	6.85	6.65	6.39	5.92	6.33
ha6	1-hexanol	537064	Fruity, alcoholic64	Fruity, Solvent	1.18	0.01	<0.1	<0.1	<0.1	0.01	0.01	0.01
ee1	ethyl acetate	81.567	Pineapple67	Fruity	1.02	2.39	1.95	2.10	2.09	1.95	1.57	1.83
ee2	ethyl 2-methylpropanoate	25.567	Berry67	Fruity	1.07	7.34	8.81	12.74	11.05	11.00	13.01	13.21
ee4	ethyl 3-methylbutanoate	39.667	Apple67	Fruity	1.56	100.00	100.00	100.00	100.00	100.00	100.00	100.00
ae2	butyl acetate	183064	Fruity64	Fruity	1.52	0.05	0.04	0.05	0.05	0.05	0.04	0.05
ald1	2-methyl butanal	1668	Cocoa, almond68	Roasted/Smoky	1.45	1.75	3.59	4.34	4.51	3.29	3.85	3.99
ket7	3-heptanone	45668	Cheese68	Fatty	1.36	0.33	0.29	0.28	0.28	0.29	0.27	0.31

Identification of volatiles in whiskeys by HS-GC-IMS

To further investigate the differences in volatile compositions between whiskeys aged with toasted chestnut shell chips and those aged with toasted oak chips, secondly, HS-GC-IMS was used to analyze the volatile compounds. A total of 74 volatile compounds were identified in all whiskeys via HS-SPME-GC-IMS, including 7 higher alcohols, 17 esters (4 ethyl esters, 4 acetate esters, and 9 other esters), 14 carbonyl compounds (3 aldehydes, 10 ketones, and 1 acetals), 2 organic acids, 3 aromatic compounds, 4 nitrogenous heterocyclic compounds, 6 oxygenous heterocyclic compounds, 6 sulfur-containing compounds, 4 terpenes, 2 other compounds, and 9 unidentified compounds. Among them, 1,1-diethoxy ethane (acetals) exhibited multiple signal peaks, indicating the presence of monomer, dimer, and trimer. The difference comparison topographic plots were obtained by using the topographic plot of the control sample as a reference (Fig. 3b). Compared to the control sample, concentrations of a variety of compounds significantly changed in the whiskeys aged with toasted chestnut shell chips. Moreover, substantial differences were also observed between these whiskeys and those aged with toasted oak chips.

To intuitively reflect the differences involatile compositions between the whiskeys aged with these two types of chips, the fingerprint of volatile compounds was constructed. As illustrated in Fig. 3c, the fingerprint plot can be clearly categorized into four distinct regions. Region A contains the volatiles that with elevated relative concentrations in whiskeys aged with toasted chestnut shell chips. Notably, the major representatives included (Z)-3-nonen-1-ol, 1-pentanol, ethyl 3-methylbutyrate, ethyl propanoate, butyl acetate, 2-methylbutanal, acetone, 2-pentanone, 3-heptanone, 1,1-diethoxyethane (both dimer and trimer forms), 3-octenoic acid, 2-furanmethanol, 4-methylthiazole, methyl 2-propenyl sulfide, 1-octene, and two unidentified compounds labeled as un1 and un4. Furthermore, compared to LS chips and HS chips, MS chips aging demonstrated a more pronounced enhancement effect on the majority of these compounds (Fig. 3c). The relative concentrations of volatiles, such as (E)-2-heptenal, 2-methylpropyl 2-methylpropanoate, 2-butanone, 3-ethyl-2-hydroxy-2-cyclopenten-1-one, 2-ethylbutanoic acid, furan, 2-methylfuran, myristicin, un3, and un9, which were included in Region B, were significantly increased and exhibited higher levels in whiskeys aged with toasted oak chips. In Region C, the relative concentrations of the four volatile compounds, 3-pentanone, 1,1-diethoxyethane, 2,3-diethyl-5-methylpyrazine, and un2, were consistently decreased after aging with both toasted chestnut shell chips and toasted oak chips. For the remaining 24 volatiles in Region D, however, their concentrations were largely unaffected by the aging of these two types of chips.

To better understand the difference in volatile compositions among whiskeys aged with toasted chestnut shell chips and toasted oak chips, PCA was carried out using the concentrations of all volatiles detected via HS-GC-IMS as variables (Fig. 3f). The first two principal components accounted for a combined variance of 56.1%, with PC1 and PC2 explaining 39.5% and 16.6% of the variance, respectively. As shown in Fig. 3f, a clear distinction was observed among the aged and control whiskeys, which was consistent with the PCA results obtained from HS-SPME-GC-MS data. For the control sample, the high loading values obtained for ket3 and un2 in both the negative direction of PC1 and PC2 were the primary contributors. For the aged whiskeys, a series of volatiles, such as oe3, un1, scc1, ee3, ket7, oc1, scc3, ee1, un4, ee4, oe1, ohc6, ket5, ket1, and ohc2, in the positive direction of PC1 corresponded to the samples aged with toasted chestnut shell chips. Conversely, several volatiles, for example scc2, ohc3, ald2, ohc1, oa1, un3, and oe4, in the negative direction of PC1 corresponded to the samples aged with toasted oak chips. Furthermore, due to the varying concentrations of these compounds in each sample, most of them also contribute to differentiating the whiskeys aged with two types of chips that have distinct toasting degrees.

Screen the key volatile flavor compounds from the volatiles in whiskeys identified by HS-GC-IMS

For volatiles detected by HS-GC-IMS, the VIP values and the ROAV were also combined to identify the key volatile flavor compounds and evaluate their contributions to the overall aroma profile of these samples. The volatiles with VIPs ≥ 1.0 were marked with star symbols in fingerprint plot. Among them, 8 volatiles with ROAV ≥ 0.1 were listed in Table 3. Compared to the control sample, both whiskeys aged with toasted chestnut shell chips and toasted oak chips demonstrated a more pronounced intensity of roasted/smoky aroma characteristic, particularly after aging with MS and HS chips, which can be attributed to the increased concentration of 2-methyl butanal. Meanwhile, the overall fruity aroma characteristic intensity was also enhanced in whiskeys after aging with these chips, with ethyl 2-methylpropanoate being the primary contributor. For herbaceous and fatty aroma characteristics in whiskeys, aging with toasted chestnut shell chips did not produce a notably effect on their intensities, whereas aging with toasted oak chips resulted in a slight decrease in their intensities, mainly attributable to variations in the ROAVs of (Z)-3-nonen-1-ol and 3-heptanone.

Comparative analysis of GC-MS and GC-IMS results

This study conducted a systematic and in-depth analysis of volatile compositions in whiskeys aged with toasted chestnut shell chips and toasted oak chips by integrating HS-SPME-GC-MS, and HS-GC-IMS techniques. According to the data obtained, a greater variety of other esters, ketones, nitrogenous heterocyclic compounds, oxygenous heterocyclic compounds, and sulfur-containing compounds were detected through HS-GC-IMS (Supplementary Table 3), whereas more varieties of higher alcohols, ethyl esters, aldehydes, aromatic compounds, and phenols were identified through HS-SPME-GC-MS (Fig. 2). Furthermore, the amounts of each group of volatiles also exhibited differences in the samples analyzed by these two methods. These discrepancies can be attributed to the distinct detection principles and varying sensitivities of GC-MS and GC-IMS toward the detected volatiles²⁶. Additionally, differences in the enrichment procedures between these two methods may also lead to variations in the types and amounts of the volatiles detected³⁰. Among the 131 volatiles detected in all whiskey samples, 57 were exclusively identified by HS-SPME-GC-MS, 66 were uniquely detected by HS-GC-IMS, and 8 were jointly identified by both methods (Figs. 2, 3). Despite the differences in both qualitative and quantitative results of volatiles between these two techniques, a similar distribution pattern was observed in the PCA results based on data obtained from HS-SPME-GC-MS and HS-GC-IMS. Moreover, the analysis of key volatile flavor compounds based on HS-SPME-GC-MS and HS-GC-IMS data both indicates that aging with toasted chestnut shell chips could considerably enhance the herbaceous, fatty and roasted/smoky aroma characteristics of whiskeys, which are distinct from those aged with toasted oak chips and dominated by fruity aroma characteristic. Therefore, integrating HS-SPME-GC-MS and HS-GC-IMS techniques indeed provide a more comprehensive analysis of the volatile composition differences between whiskeys aged with toasted chestnut shell chips and those aged with toasted oak chips.

Artificial sensory analysis

To better understand the differences of aroma sensory characteristics between whiskeys aged with toasted chestnut shell chips and those aged with toasted oak chips, the artificial sensory analysis was conducted using described the aroma terms from the aroma kit and subsequently scored on a 5-point scale. The MF% results presented in Fig. 4 revealed that both toasted chestnut shell chips aging and toasted oak chips aging significantly altered the aroma sensory attributes of the whiskeys. For the whiskeys aged with toasted chestnut shell chips, they were characterized by a higher level of green/plant and roasted/smoky aroma sensory attributes, and hexanal, 2-methyl butanal and guaiacol could be the contributors to these aroma characteristics (Tables 2 and 3). In contrast, the whiskeys aged with toasted oak chips exhibited higher intensities of fruity and Floral aroma sensory attributes, which could potentially be ascribed to the higher ROAVs of most ester compounds present in them (Tables 2 and 3). It is worth noting that the whiskeys aged with toasted chestnut shell chips showed a lower fusel/solvent aroma sensory attribute compared to those aged with toasted oak chips, suggesting that toasted chestnut shell chips may possess a stronger purification capacity for unpleasant aromas during the aging process.

Fig. 4.

Aroma sensory characteristics (Modified frequencies, %) of whiskeys aged with toasted chestnut shell chips and toasted oak chips.

Table 2.

Volatiles with VIPs ≥ 1.0 and ROAV ≥ 0.1in whiskeys identified by HS-SPME-GC-MS

No.	Compounds	Odor threshold (μg/L)	Aroma description	Aroma type	VIP	ROAV
						Control	LS	MS	HS	LO	MO	HO
EE2	Ethyl butyrate	81.566	Pineapple67	Fruity	1.24	1.15	1.37	1.13	1.24	1.09	1.07	1.10
EE5	Ethyl hexanoate	55.366	Fruity67	Fruity	1.24	16.30	17.38	16.33	15.73	17.70	17.36	17.52
EE8	Ethyl octanoate	12.966	Fruity67	Fruity	1.32	34.45	31.78	29.75	23.82	41.27	45.13	40.44
EE11	Ethyl benzoate	143066	Floral67	Floral	1.15	0.10	0.24	0.22	0.22	0.13	0.18	0.17
ALD2	2-Methyl butanal	1668	Cocoa, almond68	Roasted/Smoky	1.13	0.91	1.38	1.16	1.18	0.95	0.86	0.86
ALD4	Hexanal	25.567	Grassy67	Herbaceous	1.26	0.64	1.34	1.00	1.16	0.89	0.89	0.76
ALD5	Octanal	39.667	Fatty67	Fatty	1.57	Nd	1.40	1.15	3.64	Nd	Nd	Nd
ALD6	Nonanal	12266	Soapy67	Fatty	1.50	0.79	1.74	1.55	3.18	0.81	0.82	0.77
ACE1	1,1-Diethoxy ethane	209066	Fruity66	Fruity	1.51	0.11	0.24	0.22	0.21	0.11	0.13	0.11
AC3	Benzene acetaldehyde	26269	Floral67	Floral	1.04	0.13	0.20	0.21	0.22	<0.1	<0.1	<0.1
AC5	Naphthalene	15967	Mothball67	Fusel/Solvent	1.37	<0.1	<0.1	<0.1	<0.1	0.10	0.11	<0.1
PHE1	Guaiacol	13.468	Smoky68	Roasted/Smoky	1.10	Nd	Nd	Nd	1.67	Nd	Nd	Nd
TP1	Linalool	2370	Rose, flora21	Floral	1.14	7.37	6.07	4.07	7.40	7.76	7.36	7.05
TP5	β-Damascenone	0.1471	Floral, honey66	Floral	1.14	100.00	100.00	100.00	100.00	100.00	100.00	100.00

The result of this study confirmed that, after appropriate roasting, chestnut shells can transform into a novel and cost-effective material for aging distilled spirits, using the case of whiskey. Aging whiskeys with toasted chestnut shell chips moderately increases the titratable acidity and total phenol content, and gives them a color ranging from yellowish to light amber. These aged whiskeys also exhibit a high level of aldehydes and a low level of esters, and they present pronounced roasted/smoky aroma characteristic that is distinct from those aged conventionally with toasted oak woods. This study provides valuable insights into the conversion of agro-industrial by-product into new oak alternatives. In this way, they can acquire a new life in the oenological field and generate economic benefits.

Methods

Preparation of newly distilled whiskey

The Australian barley malt (4-5 EBC, purchased from GDH SUPERTIME QINHUANGDAO MALTING CO., LTD.) with a complete appearance was selected and crushed to powder using a pulverizer (TY-260, Hubei Shouxing Machinery Co. Ltd., Shishou, China). Then, the grain powder and water (in a ratio of 1:4) were added to an electric heating and insulation bucket (Y88, Chuangyouke Technology Co. Ltd., Shenzhen, China), maintained at 65 °C for 1.5 h to facilitate saccharification. After filtering and cooling, the saccharification solution was transferred to the fermenter. Then, the commercial whiskey yeast W19 (at a dosage of 0.8 g/L, Angel China) was activated and inoculated, and fermentation was carried out at 28 °C for 72 h. Subsequently, the fermentation liquid was transferred to a stainless-steel pot still (L10, Yishengyuan Industrial Co. Ltd., Pingdingshan, China) for the initial distillation process. After this stage, the alcohol content of crude liquor was approximately 20% vol. Then, the secondary distillation was initiated. The heads, which contained approximately 2% of the total alcohol content, were discarded. The resulting distillate, with a final alcohol content of approximately 62% vol was collected as newly produced whiskey and stored at room temperature for further use.

Preparation of toasted chestnut shell chips and toasted oak chips

The chestnut shells of “Qiuhong”, collected from the laboratory-owned plantation of Hebei Normal University of Science & Technology, were manually selected and cut into chips (approximately 10 × 5 × 1 mm) using scissors. Then, the chestnut shell chips were toasted in the oven at 150 °C, 175 °C, and 225 °C for 30 min, respectively, to obtain the lightly, medium, and heavily toasted chestnut shell chips for further use.

The lightly, medium, and heavily toasted oak chips, purchased from LAMOTHE-ABIETL, were manually selected and cut to the corresponding size of the chestnut shell chips for further use.

Experimental design

The trial was conducted on a laboratory scale using 500 mL glass bottles, comprising three aging modalities, each with three replicates:

Control: newly distilled whiskey with no treatment.

LS (lightly toasted chestnut shell chips), MS (medium toasted chestnut shell chips), HS (heavily toasted chestnut shell chips): newly distilled whiskey aged with 10 g/L of corresponding toasted chestnut shell chips.

LO (lightly toasted oak chips), MO (medium toasted oak chips), HO (heavily toasted oak chips): newly distilled whiskey aged with 10 g/L of corresponding toasted oak chips.

All above-mentioned bottles were stored in the incubator at a controlled temperature of 20 ± 1 °C for 30 days, with daily agitation at a rotational speed of 200 rpm for 3 min. After aging, LS, MS, HS, LO, MO, and HO samples were filtered to remove the chips for further analysis.

Physicochemical parameters analysis

The alcohol content (% vol), titratable acidity (expressed as acetic acid equivalent, g/L), volatile acidity (expressed as acetic acid equivalent, g/L), and pH value of whiskeys were measured according to the National Standard of the People’s Republic of China: GB 5009.225-2023, “Determination of alcohol content in wine and edible alcohol” and GB 12456-2021, “Determination of total acid in food”.

Total phenolic content analysis

The total phenolic content (expressed as of gallic acid equivalents, mg/L) was determined according to the method previously described by Lu et al.⁶³.

Chromatic characteristics analysis

The CIELab parameters, L* (lightness), a* (redness-greenness), and b* (yellowness-blueness), of the whiskeys were determined using a YS6003 colorimeter (3nh Technology Co. Ltd., Shenzhen, China). The ΔE (total color difference) was calculated using the following equation:

$$\mathrm{\Delta }\mathrm{E}={[{\left(\mathrm{\Delta }{\mathrm{L}}^{*}\right)}^2+{\left(\mathrm{\Delta }{\mathrm{a}}^{*}\right)}^2+{\left(\mathrm{\Delta }{\mathrm{b}}^{*}\right)}^2]}^{1/2}$$ 1

Electronic nose analysis

The electronic nose analysis of volatiles was performed by a PEN 3 E-nose system (Airsense Analytics Co. Ltd., Schwerin, Germany) equipped with ten sensors. The performances of these sensors are listed in Supplementary Table 1. The analysis of E-nose followed an established methodology with slight modifications²⁶. Briefly, the five-fold diluted whiskey sample (2.0 mL) was added to a 20 mL SPME glass vial and placed at room temperature (26 °C) for 40 min to attain equilibrium in the headspace. The gas in the headspace was then pumped over the sensors for analysis. The test conditions were set as follows: 5 s injection preparation time, 60 s detection time, 5 s autozero time, 200 s sensor cleaning time, and 400 mL/min carrier gas flow rate. Each sample was tested in triplicate.

HS-SPME-GC-MS analysis

The analysis of HS-SPME-GC-MS followed a previously described method with slight modifications⁵⁸. Briefly, 5.0 mL of five-fold diluted whiskey sample (approximately 12.5% v/v) was added to a 20 mL SPME glass vial with 1.5 g of sodium chloride, and 10 µL of the internal standard 4-methyl-2-pentanol (1.00 g/L). The vial was sealed with a PTFE-Silicone septum and equilibrated at 40 °C for 30 min under a rotational speed of 250 rpm. Subsequently, the 50/30 µm DVB/CAR/PDMS fiber (57328-U, Supelco, Bellefonte, PA, USA) was immersed in the headspace of an airtight vial to extract the volatiles at for 30 min with continued heating and rotation. Afterward, the SPME fiber was inserted into the GC injection port to be thermally desorbed for 10 min at 250 °C (in splitless mode) and began the GC-MS analysis.

The GC-MS analysis was carried out using Agilent 7890B GC system and 5977 A MS detector equipped with a DB-Wax capillary column (60 m × 0.250 mm × 0.25 µm, Agilent J&W, USA). The carrier gas was ultra-pure helium (99.999%) and the flow rate was 1 mL/min. The temperature of the transfer line and ion source was set as 250 °C. The column oven temperature was set as follows: the process started at 50 °C for 1 min and then increased to 220 °C at 3 °C /min with a final holding time of 5 min; the total run time was 62.7 min. The MS was operated in the electron ionization (70 eV) and full-scan model (30-450 m/z) at intervals of 0.2 s. Each sample was analyzed in triplicate.

The volatiles were identified by comparing their retention indices (RIs) and mass spectra with those of the pure standards in the NIST Standard Reference Database (NIST, Chemistry WebBook) and the standard NIST 20 library. The RIs were calculated using the C10-C24 n-paraffin Mix (all even, soluble in heptane, Sigma, Switzerland) under identical chromatographic conditions as for the diluted whiskey samples. The volatiles were quantified using the internal standard method by comparing the GC peak areas of the samples to that of the internal standards, and the results were expressed in μg/L.

HS-GC-IMS analysis

The analysis of HS-GC-IMS followed an established methodology with slight modifications⁶⁴. Briefly, 2.0 mL of five-fold diluted whiskey sample (approximately 12.5% v/v) was added to a 20 mL SPME glass vial with 10 µL of the internal standard 2-methyl-3-heptanone (10 mg/L). The vial was sealed with a PTFE-Silicone septum and equilibrated at 60 °C for 15 min under a rotational speed of 500 rpm. Subsequently, the syringe of the automatic headspace sampler unit (CTC Analytics AG, Zwingen, Switzerland) was immersed in the headspace of the airtight vial to extract 100 μL of the headspace gas at 85 °C, and then injected into the GC injection port in splitless mode and began the GC-IMS analysis.

The GC-IMS analysis was carried out using G.A.S. FlavourSpec GC-IMS system equipped with MXT-WAX capillary column (30 m × 0.53 mm × 1.0 μm, Restek, USA). The carrier gas was ultra-pure helium (99.999%) and the flow-rate was set as follows: the process started at 2 mL/min for 2 min, increased to 10 mL/min at 1 mL/min, increased to 100 mL/min at 9 mL/min with a final holding time of 39 min. The drift tube temperature was 45 °C, and the drift gas velocity was 150 mL/min. Each sample was analyzed in triplicate.

The volatiles were identified by comparing their RIs and drift time (Dt) with those of the pure standards in the NIST 20 library and the IMS Database (G.A.S., Dortmund, Germany). The RIs were calculated using the n-ketones C4-C9 (Sinopharm Chemical Reagent Beijing Co., Ltd., China) as external references. The volatiles were quantified using the internal standard method by comparing the GC peak volume of the samples to that of the internal standard, and the results were expressed in μg/L.

ROAV analysis

The relative odor activity value (ROAV) is a generally used parameter for evaluating the contribution of each volatile compound to the overall aroma of distilled beverage, and it is calculated using the following equation:

$${\mathit{ROAV}}_x=100\times ({\mathit{OAV}}_x/{\mathit{OAV}}_{max}$$

$$=100\times ({\mathrm{RC}}_x/{\mathrm{OT}}_x)/({\mathrm{RC}}_{\mathit{OA}Vmax}/{\mathrm{OT}}_{\mathit{OAV}max})$$

$$=100\times ({\mathrm{RC}}_x/{\mathrm{RC}}_{\mathit{OAV}max})\times ({\mathrm{OT}}_{\mathit{OAV}max}/{\mathrm{OT}}_x)$$ 2

RC_x: the relative content of the volatile compound to be measured

RC_OAVmax: the relative content of the volatile compound with the highest OAV

OT_x: the odor threshold value of the volatile compound to be measured

OT_OAVmax: the odor threshold value of the volatile compound with the highest OAV

Artificial sensory analysis

The aroma sensory evaluation of whiskeys was performed using the Modified Frequency (MF%) method⁶⁵. A group of 10 tasters (6 females and 4 males, aged between 20 and 36 years) from the College of Food Science at Hebei Normal University of Science & Technology had been previously trained with the “Master Wine Aroma Kit” before conducting the analysis. The aroma sensory evaluation was carried out in a tasting room at 20 °C, where each whiskey sample was presented in ISO standard tasting glasses for orthonasal evaluation. Each taster was instructed to describe the olfactory profiles of the wines using a set of 5 aroma terms from the “Master Wine Aroma Kit” (including “Green/Plant”, “Fruity”, “Floral”, “Fatty”, “Roasted/Smoky”, “Sweet”, “Fusel/Solvent”) and to score the intensity of each aroma term on a 5-point scale (1 = very weak; 2 = weak; 3 = medium; 4 = high; 5 = very high). Each sample was analyzed in triplicate. The final result of each aroma term was expressed as MF%, representing the combination of both “Intensity” and “Frequency” of detection, which was calculated using the following equation:

$$\mathrm{MF}(\%)={\left[F\right(\%)\times I(\%\left)\right]}^{1/2}$$ 3

F (%): the detection frequency of an aroma attribute expressed as a percentage

I (%): the average intensity expressed as a percentage of the maximum intensity

Statistical analysis

The data are expressed as the mean ± standard deviation of three independent experiments. One-way analysis of variance (ANOVA) was performed using IBM SPSS Statistics 19.0 software (SPSS Inc., Chicago, IL, USA), employing Duncan’s multiple range tests at a significance level of p < 0.05. Principal component analysis (PCA) was conducted using the Origin 2018 (OriginLab Corporation, Northampton, MA, USA). Variable importance in projection (VIP) values were obtained using SIMCA 14.1 software (Sartorius, Göttingen, Germany). Fingerprint plots and difference plots were generated using VOCal 0.4.03 software (G.A.S., Dortmund, Germany).

Author contributions

Yue Zhao: conceptualization, investigation, visualization, and writing-original draft. Xinyi Lu: investigation, and methodology. Qingyang Sun: investigation, visualization, and writing-review and editing. Ruiguo Cui: writing-review and editing. Haoran Wang: writing-review and editing. Yaya Yao: writing-review and editing. Xiaoyu Liu: writing-review and editing. Lijun Song: conceptualization, and writing-review and editing. All authors read and approved the final manuscript.

Data availability

The data used and/or analyzed during the current study belong to The Hebei Normal University of Science & Technology; any sharing needs to be evaluated and approved by the authors and the University.

Competing interests

The authors declare no competing interests.

Supplementary information

The online version contains supplementary material available at 10.1038/s41538-026-00715-9.