Development of Chocolate Aroma from Underutilized Durian Seeds

Home , Cocoa bean, Durian, Durio zibethinus, Jackfruit

INTERNATIONAL JOURNAL OF SCIENTIFIC & TECHNOLOGY RESEARCH VOLUME 9, ISSUE 03, MARCH 2020 ISSN 2277-8616

Development Of Chocolate Aroma From Underutilized Durian Seeds

Mindula Kaumadi Wijayahena , Champa Disala Jayaweera

Abstract: Durian is an exotic tropical fruit containing seeds which are generally discarded after consumption of the flesh. The present study aimed at exploring the feasibility of the processed seed powder of underutilized durian (Durio zibethinus Murr.) seeds for the development of chocolate aroma. Durian seeds were fermented (FDS), acidified (ADS) and dried (DDS) to produce three types of seeds and they were roasted under different time/temperature conditions to produce eleven processed seed powder samples from each type according to the central composite design. The chocolate aroma of powder samples was ranked by a sensory panel having 180 panellists. The results indicated that FDS and ADS powders yielded the best chocolate aroma. Main odour active compounds which are responsible for chocolate aroma; 3-methylbutanal, phenylacetaldehyde, 2-phenylethyl acetate, trimethylpyrazine and 2,3-diethyl-5-methylpyrazine were identified in FDS powder using SPME/GC-MS. The FDS sample prepared under the moderate roasting condition was selected as the best powder with high chocolate aroma. The processed durian seed powder can be utilized as an alternative source for cocoa powder.

Index Terms: Chocolate aroma, Fermentation, Flavor development, Seed processing, Sensory evaluation, SPME/GC-MS, Underutilized Durian seeds ————————————————————

1 INTRODUCTION Durian and cocoa beans belong to the order Malvalves, and Aremarkable increase in the production, marketing, and both fruits shared a common ancestor about 65 million years consumption of exotic tropical fruits has been witnessed in the ago [10]. The demand for cocoa beans increases with the high world in the past decade and this trend can be accounted for consumption of chocolate products. Globally 4.5 millions of the growing recognition of the rich nutraceutical values and people are involved in cultivating cocoa and the cocoa harvest sensory attributes of such fruits [1]. Durian (Scientific name: is about 4.7 million mT/year [11],[12]. It is sensitive to changes Durio zibethinus Murr.; family: Bombacaceae) is a seasonal in environmental conditions and also susceptible to many tropical fruit either oblong or round in shape, with an outer diseases [13]. Therefore, fulfilling the future cocoa demand is spiky skin being green to brown colour. It is mainly distributed difficult as cocoa beans are the main source in chocolate in Southeast Asia (Malaysia, Philippines, Indonesia, and production as described in ―Fairtrade and Cocoa- Commodity Thailand) [1]. The Portuguese brought Durian to Sri Lanka in Briefing‖ [14]. Hence, it is important to seek for new alternative the 16th century from Southeast Asia and it is cultivated sources of chocolate aroma and flavour to maintain and to mainly in the southern part of the country as described by control this demand or else the chocolate industries may step Kanazaki et al [2] and in the webpage ―Durian Season in Sri towards a crisis in future. Chocolate contains a unique aroma Lanka 2019‖ [3]. The name originates from the Malaysian word and this is developed via the processes that carried out to ―duri‖, which means thorn. The ripe fruit is considered as the cocoa beans prior to the preparation of cocoa powder. Cocoa ‗King of Fruits‘ due to the fact that it has the highest fragrance bean processing contains three stages as fermentation, and flavour among fruits. Durian requires separating the skin drying, and roasting [15]. Fermentation plays the major role in from the flesh, the edible part is sweet in taste and the seeds formation of aroma and flavour precursors; amino acids and are usually round; the colour range from yellowish brown to reducing sugars. During fermentation, the embryo of the bean reddish brown. The durian harvest produces about 67-70% of is destroyed and lead to the production of ﬂavor precursors wastes where about 20-25% in the form of seeds and the rest [16]. Sucrose and its hydrolysis products release reducing is the non-edible spiky skin. Increasing efforts have being sugars and acid hydrolysis of proteins releases about 190 free made to convert these underutilized seed waste to useful amino acids and oligopeptides [16]. These flavour precursors; products [4],[5],[6],[7]. Seeds are roasted and boiled to amino acids and reducing sugars are used up during roasting produce snacks in countries where the durian fruit is abundant by the Maillard reaction as explained by the Hodge scheme to and sugar coated durian seed slices are used as candies. produce main odourant and flavour classes, aldehydes and Further, it has been reported that coconut oil fried spicy slices pyrazines [17]. During roasting, the odour active Strecker and curry prepared from seeds are consumed with rice aldehydes and aminoketones are formed via interacting - [1],[8],[9]. To the best of our knowledge, no reports were found dicarbonyl of 3-deoxyaldoketone with -amino acid. In the next in literature on their potential in generating flavours. We have step, these aminoketones are used up by condensation found for the first time that processing of durian seeds can reactions to produce the odour active pyrazines [15],[18]. The impart a flavour similar to that of chocolate. The most popular most odour active compounds that provide flavour in chocolate and favorite sweet food product chocolate is produced from are phenylacetaldehyde, 2,3-diethyl-5-methylpyrazine, 3- cocoa powder and cocoa butter found in cocoa beans. methylbutanal, trimethylpyrazine, and 2-phenylethyl acetate [19] and these compounds are used for the quantitative and qualitative determination of cocoa/chocolate flavor [20]. The ______main focus of this research is to explore the potential of durian seeds, an alternative source to produce the chocolate aroma • Champa D. Jayaweera, Senior Lecturer, Department of Chemistry, University of Sri Jayewardenepura, Sri Lanka. E-mail: by optimizing the conditions in a procedure similar to cocoa [email protected] bean processing. Acidified, fermented, and dried durian seeds • Mindula K. Wijayahena, Department of Chemistry, University of Sri will be roasted under different time/temperature conditions to Jayewardenepura, Sri Lanka. E-mail: [email protected] produce a powder. Chocolate aroma of the powder will be

assessed by sensory ranking tests and the key odourant compounds will be identified by SPME/GC-MS.

2 PROCEDURE

2.1 Durian Ripe durian fruits were collected between July to October 2017 from the durian trees authenticated as Durio Zibethinus Murr. by the Fruit Crops Research and Development Station, Gannoruwa, Peradeniya, Sri Lanka. The pulp around the seed was removed manually. All seeds were washed well with distilled water and stored at -4 °C.

2.2 Seed processing of durian seeds Three types of processed durian seeds were prepared as dried durian seeds (DDS), acidified durian seeds (ADS) and fermented durian seeds (FDS). Preparation of DDS: The washed seeds were dried in an oven (DHG-9030A, Henon TABLE 1 Touch Science Instruments Co., Ltd) with air circulation at 60 CENTRAL COMPOSITE DESIGN USING TWO FACTORS °C for 24 hours. Then the spermoderms were removed EACH AT FIVE LEVELS. manually and seeds were kept in the oven again at 60 °C for another 24 hours. Preparation of ADS: The washed seeds Temperature /°C were placed in a glass container and 500 mL of 1% v/v acetic Levels Time /min acid was added. The solution was removed after 5 days and the acidified seeds were separated and dried using the same DDS ADS FDS procedure used for DDS. Preparation of FDS: The washed -1.41 33 150 159 150 seeds were placed in a polyethylene container. Ripe jackfruit -1 35 156 165 154 pulp and banana leaves were added to it as a source for 0 40 171 180 165 microbial growth. For the first 7 days of fermentation, the 1 45 186 195 176 container was closed to provide the anaerobic condition and 1.41 47 192 201 180 then for the next 7 days the container was kept open and the fermenting mass was aerated daily by rolling the fermenting 2.4 Analysis of powder mass. After the 14th day the seeds were separated and dried using the same procedure used for DDS. The temperature of pH value the fermenting and acidifying mass was measured every 24 The pH values were determined using 1 g of powder dissolved hours. The pH was measured directly in ADS by taking small in 10 ml of distilled water [19]. portion of liquid extracted from the glass container. The pH of FDS was measured after mixing 2.5 g of the fermenting mass Colour in a 25 mL of distilled water [21]. The colour of commercial cocoa and processed durian seed powder samples were compared visually. Sensory analysis for 2.3 Roasting and grinding aroma in processed durian seed powderThree different groups The roasting conditions defined by the response surface containing 60 untrained panellists aged between 22 to 55 methodology carried out by Fernanda et al [19] to produce years participated for the sensory evaluation of the three types chocolate aroma from jackfruit seeds were followed as both of processed seed powder (DDS, FDS and ADS) samples. seeds have similar physical characteristics. For each Total number of panellists was 180. From 33 powder samples, treatment (DDS, ADS and FDS), 11 samples of seeds were about 5 g of processed seed powder was taken and heated in roasted in an air circulating oven with a digital temperature a water bath at 40 oC for 120 s. Firstly 9 ADS processed seed control according to the central composite design (Fig. 1). The powder samples were given to the first batch of panellists (60) roasting time and temperature are the two factors considered to smell. Each panellist received 9 samples where a set of 3 with three repetitions of the central point. Each type of samples were given at a time. The DDS and FDS processed processed seeds roasted under different conditions as shown seed powder samples were also given for the panellists for in Table 1, were milled using a motor and pestle to produce a sensory analysis in the same manner. powder. The powder samples were separately packed in food packaging polyethylene bags and stored at 4 °C. The yield of Volatile analysis the powder of each type was calculated using the following About 5 g of FDS / ADS processed seed powder was placed in equation, a SPME vial (20 mL). While the vial was in the water bath kept at 60 °C, the triple-phase gray colour code fiber (50/3 µm DVB/CAR/PDMS) was inserted to the space above the sample and the fiber was exposed for different times, i.e. 30 min. or 60 min. Water bath temperature, type of the triple-phase fiber and exposing time of the fiber were selected by trial and error tests.

The extracted volatile compounds by the fiber were analysed through GC-MS (GC – Agilent technologies, 7890A GC system) containing 19091S-433HP-5MS column. Absorbed compounds were desorbed for 5 min. during splitless injection at 240 °C. Oven was maintained at 40 °C at the beginning and the temperature was increased at 2 °C/min to 70 °C, 5 °C/min to 170 °C and finally 50 °C/min ramp rate to reach 240 °C. Flow rate of the carrier gas helium was 1 mL/min in constant flow. Volatile compounds in the samples were analysed by comparing their mass spectra with the mass spectra of authentic standards which were in the data base.

3 RESULTS AND DISCUSSION

3.1 Durian seeds The most interesting fact that was observed of the cleaned durian seeds was the absence of the most unusual flavour and aroma of the ripe durian pulp. 3.4 Roasting and grinding 3.2 Processing of durian seeds The treated durian seeds were roasted under different An alternative new source for cocoa beans was investigated in temperature and time conditions as discussed earlier. The this work. Hence, the condition that give the best flavour and roasted, processed seeds were ground manually to prevent the fragrance during durian seed fermentation, drying and roasting evaporation of volatile compounds from the powder samples. The has to be found. yields obtained for DDS, ADS and FDS powders were 58%, 39% and 37% respectively. A relatively moderate yield was obtained 3.3 Fermentation/ Acidification of durian seeds for the processed durian seed powders as the spermoderm of Cocoa beans are not big and hard as durian seeds and therefore durian seed was thick and it was removed before roasting. The easy to ferment within a short duration, about 5 to 8 days. yields of ADS and FDS powder types are less when compared to However, durian seeds have more carbohydrate to decompose; the yield of DDS powder type. more substrate for microorganisms to act upon. Therefore, 14 days were chosen to complete the fermentation process. The 3.5 Powder analysis destruction of embryo of the cocoa seed occurs around pH value pH value 4 during fermentation [22]. The variation of pH during the The pH range of FDS and DDS powders were 5.50 - 5.92 and fermentation process of the ADS and FDS types is shown in Fig. 5.63 - 5.86. They are slightly higher than the pH range of ADS 2a. In the FDS type, the initial pH value was around 5 and then it powders; 4.86 - 5.35. The lowest pH of FDS powder reported was decreased to around 4 (Fig. 2a(ii)). However, in ADS type, the pH 5.50. The pH of fermented cocoa seeds are reported around 4.75 started around 3 and fluctuated around 4 (Fig. 2a(i)). It is reported – 5.19 [23]. The pH of ADS was comparatively lower due to the that the well fermented cocoa seeds show a pH value more than acidification process. 4 [23]. The pH value of FDS type is higher than 4 indicating the similarity between the two fermentation processes. Further, both Colour FDS and ADS types have the pH value higher than 4. The The colour of a food is an important character for the recognition literature found for the processing of cocoa beans reports that the and enjoyment. When comparing the cocoa powder and durian fermentation takes place via two stages [16],[23]. During the first seed powders, cocoa powder has a blackish brown colour and stage, under anaerobic conditions, yeast and lactic acid bacteria durian seed powders exhibit a pale brown colour similar to biscuit are active and during the second stage, under aerobic conditions powder (Fig. 3). The colour of the cocoa powder (Fig. 3a) acetic bacteria become prominent. Therefore, the pH of the provides the unique colour to the chocolate. The DDS powder medium decreases and reaches to a value around 4 due to the samples were in darker shade, more brownish. (Fig. 3b) than formation of acidic products by the microorganisms present in the ADS and FDS powder samples (Fig. 3c and Fig, 3d). FDS and fermenting mass. The processes that occur during the aerobic ADS powders exhibit a very light brown colour. The FDS powder process are exothermic and therefore, the temperature of the sample has the lightest colour (Fig. 3d). fermenting mass increases at the end [24]. The temperature variations of the ADS and FDS processed seed types are shown in Fig. 2b illustrating that the temperature of the FDS seeds has increased during fermentation and has reached to 30 °C at the end (Fig. 2b(i)). It is reported that in cocoa fermentation, the temperature can reach to around 45 °C [23]. In the ADS type, such an increment for the temperature is not observed as there were no microbial exothermic reactions occurring in the solution (Fig. 2b(ii)). The pH values and temperature measurements in days 12, 13 and 14 were not taken for FDS type as the sample lost its juicy nature.

reaction. Some have described the aroma having fruity qualities (vanilla, fruity) and these are likely to be related to fruity aldehydes, alcohols, and esters, which are products of the fermentation process. Aroma attributes of the durian seed powder samples varies suggesting that there is a potential for obtaining different types of aroma other than chocolate.

TABLE 2 SOME AROMA ATTRIBUTES USED BY THE PANELISTS TO DESCRIBE THE FDS, ADS AND DDS POWDERS.

Type Aroma attribute

Chocolate, milky, malt, coffee, Nescafe, tea, biscuit, cheesy, FDS bee honey, fruity, cashew, oats, woody, fermented

Chocolate, biscuit, tea, malt, cereal, vanilla, spicy, wheat, ADS barley, roasted, woody, fermented, bitter

Chocolate, biscuit, coffee, peanuts, tea, malt, spicy, caramel, DDS milky, corn flakes, cashew, vanilla, cereal, coffee Sensory analysis of aroma in processed durian seed powder It is found from the ranking score of the sensory analysis that ADS and FDS powder samples have the most significant chocolate aroma character, when compared to DDS powder sample as Volatile analysis shown in Fig. 4. As the sensory score for ADS and FDS are higher The chocolate aroma comes from a mixture of odourant than for the DDS powder samples it confirms that fermenting or compounds and these are present at very low levels. The most acidifying process is required for durian seed processing to odour active compounds present in chocolate aroma are 3- produce the chocolate aroma more intensely. It is also reported in methylbutanal, phenylacetaldehyde, 2,3-diethyl-5- literature that fermentation process is necessary for cocoa bean methylpyrazine, trimethylpyrazine and 2-phenylethyl acetate. processing since it forms the flavouring and odourant compounds These compounds have a high OAV (odour active value) and [23]. The highest sensory ranking score for FDS, 32 was owned by a high FD (flavour dilution) factor, contributing to a significant both sample 1 roasted under 180 °C for 40 minutes and sample 2 aroma [19]. After analysing the data obtained by the sensory roasted under 176 °C for 45 minutes while a ranking score of 48 evaluation, the two samples that produced the most intense was obtained by ADS sample 1 which was roasted under 195 °C chocolate aroma from FDS and ADS powder samples were for 45 minutes. selected for SPME/GC-MS analysis. The sensory score for DDS powder sample was relatively poor as shown in Figure 3(c) and therefore it was not considered for volatile analysis. Table 3 illustrates the presence of two key chocolate odourant compounds; phenylacetaldehyde, trimethylpyrazine and three key chocolate odour active compounds; trimethylpyrazine, phenylacetaldehyde, 2,3-diethyl-5-methylpyrazine in FDS sample 2 when exposed the SPME fiber for 30 min. and 60 min. respectively. The FDS sample 1 shows one key chocolate odour active compound at a time when exposed the SPME fiber for 30 and 60 minutes (Table 3). The ADS sample 1 contains only one key chocolate odourant compound, 2,3- diethyl-5-methylpyrazine when exposed the fiber for 60 minutes. Additionally, FDS and ADS, both types of samples, contain some other odour active compounds that have been detected in pure chocolate.

A range of aroma attributes used by the panelists to describe the FDS, ADS and DDS powders are listed in Table 2. Chocolate, biscuit and malt were common to all the three treatment types. Some attributes were common to ADS and FDS (e.g. Woody, fermented) and spicy, vanilla attributes were common to ADS and DDS. The aroma of cashew was common to FDS and DDS. Some panellists described the aroma with roast attributes, e.g. roasted, caramel; suggesting a greater contribution from the Maillard 6874 IJSTR©2020 www.ijstr.org INTERNATIONAL JOURNAL OF SCIENTIFIC & TECHNOLOGY RESEARCH VOLUME 9, ISSUE 03, MARCH 2020 ISSN 2277-8616

TABLE 3 CHOCOLATE AROMA COMPOUNDS DETECTED IN PROCESSED 2-ethylhexan-1-ol[25] ND ND ND ND 72 2,3-dimethyl-5- DURIAN POWDER SAMPLES. THE KEY CHOCOLATE AROMA ND ND ND ND 78 COMPOUNDS ARE SHOWN IN BOLD LETTERS. ethylpyrazine[25],[26] FDS FDS ADS 1H-pyrrole[25] ND ND ND ND 50 Type and sample number sample sample sample Furfurylpyrrole[25] ND ND ND ND 83 1 2 1 4-methylthiazole[25] ND ND ND ND 43 Matching percentage % 2,6-dimethyl-3-(2-methyl-1- ND ND ND ND 45 Time exposed to butyl)pyrazine[25] the SPME 2,5-dimethyl-3-(3- fiber /min ND ND ND ND 94 30 60 30 60 60 methylbutyl)pyrazine[25] Compounds detected related to ND– not detected chocolate aroma N N N 2-methylbutanal[25] 90 ND D D D 4 CONCLUSION N N N 3-methylbutanal[19],[25],[26] 45 ND Underutilized durian seeds were fermented, acidified and dried D D D to produce three types of seeds and roasted under different Methyl 2- N N N 42 90 methylpropeonate[26] D D D time/temperature conditions to produce eleven different N N processed seed powder samples from each type. The 3-methylbutanoic acid[26] 38 53 ND D D temperature and pH variation during fermentation of durian N N N 3-methyl-2-pyridinamine[25] 58 ND seeds were much similar to the variation found for D D D fermentation of cocoa seeds. FDS powder sample which was N N N 2-ethyl-5-methylpyrazine[25] 38 ND D D D prepared under moderate roasting conditions had the highest Tetramethylpyrazine[19],[25] 87 90 74 76 ND sensory ranking score and the optimum SPME/GC-MS profile Nonanal[25],[26] 72 94 91 91 87 for chocolate aroma. Sensory and volatile analyses N N N demonstrated that the fermented durian seed (FDS) powder Disulfide[25] 64 ND D D D can be used as an alternative source for cocoa powder. N Benzaldehyde[25],[26] 94 95 97 96 D Trimethylpyrazine[19],[25],[26 N 5 ACKNOWLEDGMENT 53 43 53 ND ] D The authors thank the Instrument Centre at University of Sri N N N 3-cyclopenten-1-one[25] 78 ND Jayewardenepura for GC-MS and AAS analyses. D D D 2,5-dimethyl-3- N 91 90 91 47 ethylpyrazine[25],[26] D REFERENCES 3,5-diethyl-2- N [1]. R.B. Lee-Hoon Ho, "Exploring the Potential 91 87 78 ND methylpyrazine[19],[25],[26] D Nutraceutical Values of Durian Durio Zibethinus L an 2,5-dimethyl-3-(3- N N 90 91 ND Exotic Tropical Fruit", Food Chemistry, vol. 168, pp. methylbutyl)pyrazine[25] D D 80-89, 2015. N N N Furfural[25] 90 94 [2]. S.Y. Kanzaki, Keizo. Sugiura, Akira Andhu, Suranant D D D N N N Subhadrab, "Phylogenetic Relationships of the 4-methyl phenol[25] 53 ND D D D Common Durian (Durio Zibethinus Murray) to Other 5-methyl-2- N N 91 91 96 Edible Fruited Durio Spp. By Rflp Analysis of an furancarboxaldehyde[25] D D Amplified Region of Cpdna," The Journal of Benzeneacetaldehyde/phenyl N N 92 87 ND Horticultural Science and Biotechnology, vol. 73, pp. acetaldehyde[19],[25],[26] D D 317-321, 1998/01/01 1998. N N Decanal[26] 91 90 ND D D [3]. (2019, 10 August). Durian Season in Sri Lanka 2019 N N N – Rove.Me. Available: https://rove.me/to/sri- Methyl pyrazine[25] 90 ND D D D lanka/durian N N N 2-phenylethanol[25],[26] 93 ND [4]. A.S.A. Amiza Mat Amin, Yap YinYin, NorfarizaYahya, D D D Norhayat iIbrahim, "Extraction, Purification and 2,3-diethyl-5- N N N 93 94 Characterization of Durian (Durio Zibethinus) Seed methylpyrazine[19],[25],[26] D D D 2,5-dimethyl-3-(2- N N N Gum," Food Hydrocolloids, vol. 21, pp. 273-279, 64 35 methylpropyl)pyrazine[25] D D D 2007. 3,5-dimethyl-2- N N N [5]. SSSWW9 R.A. Amiza Mat Amin, "Proximate 70 ND isobutylpyrazine[25] D D D Composition and Pasting Properties of Durian (Durio 2,3-dimethyl-5-(3- N N N 91 ND Zibethinus) Seed Flour" Int. J. Postharvest methylbutyl)pyrazine[25] D D D 2,2-dimethyl N N N N Technology and Innovation, vol. 1, pp. 367-375 2009. 46 propanal[25],[26] D D D D [6]. V. Lipipun, Nantawanit, N. and Pongsamart, S., N N N N "Antimicrobial Activity (in Vitro) of Polysaccharide Gel Ethyl 2-methylpropeonate[26] 87 D D D D from Durian Fruit-Hulls," Songklanakarin J. Sci. N N N N Furfural alcohol[25] 94 Technol., vol. 24, pp. 31-38, 2002. D D D D [7]. K.A. Ghazali, Salleh, S. F., Riayatsyah, T. M. I., N N N N 2,5-dimethylpyrazine[25] 53 D D D D Aditiya, H. B., Mahlia, T. M. I., "The Effect of Dilute N N N N Acid Pre-Treatment Process in Bioethanol Production 2-ethyl-3-methylpyrazine[25] 47 D D D D from Durian (Durio Zibethinus) Seeds Waste," IOP N N N N Octanal[26] 55 D D D D 6875 IJSTR©2020 www.ijstr.org

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