Physicochemical, antioxidant and sensory profiling of dark chocolate fortified with Terminalia Arjuna Bark powder

Introduction

Medicinal plants have long served as an important source of therapeutic agents worldwide, contributing to the growing interest in safe, effective, and economically accessible healthcare alternatives with fewer adverse effects (Jain et al., 2009) [1] . According to the World Health Organization (WHO), nearly 80% of the world's population depends on traditional systems of medicine to meet their primary healthcare needs (WHO, 2002) [2] . In developing nations like India, approximately 60% of the rural population turns to herbal remedies as a primary treatment source due to rising healthcare costs and cultural acceptance (WHO, 2002; Amalraj et al., 2017; Sewell et al., 2014; Sharma et al., 2008) [2,3,4,5] . Factors such as the increasing cost of conventional medical treatment, accessibility, and longstanding cultural acceptance have contributed to the continued use of herbal therapies (WHO, 2002; Amalraj & Gopi, 2017; Sewell et al., 2014; Sharma et al., 2008) [4] . However, only a small fraction of these bioactive rich resources has been scientifically explored or integrated into modern healthcare systems (Jeyanthi et [7,6,8,9,10] . Terminalia arjuna (Roxb.) Wt. & Arn. (Combretaceae), widely referred to as Arjuna, is a notable deciduous tree found in seasonal forests and semi-arid regions of Asia, Africa, and Australia (Gaikwad et al., 2018; Sivalokanathan et al., 2006) [11,12] . In the Indian subcontinent, it is particularly abundant along riverbanks, where it thrives in riparian soils (Sivalokanathan et al., 2006) [12] . The stem bark of T. arjuna is its most pharmacologically valuable part (Warrier et al., 1993-1996) [13] . Biochemically, the bark boasts a high density of secondary metabolites including phenolic compounds, flavonoids, tannins, triterpenoids, glycosides, saponins, and sterols, which impart exceptional antioxidant, free radical scavenging, cardioprotective, antimicrobial, and anti-inflammatory activities (Warrier et al., 1993 [13] -1996; Dhingra et al., 2013 [14] ; Sivalokanathan et al., 2006 [12] ; Cock, 2015 [15] ; Jain et al., 2009 [1] ; Viswanatha et al., 2010 [16] ; Anka et al., 2019 [17] ; Gupta & Jain, 2011) [18] . Confectionery products, particularly chocolate, represent highly popular food matrices globally (Goldstein, 2015) [19] . Modern chocolate processing creates a complex food system where cocoa solids, sugar, and milk components are dispersed in a continuous fat phase (Shafi et al., 2018; Singh et al., 2024; Özbal et al., 2022) [21, 22, 20] . While raw cocoa naturally contains health-promoting antioxidants, standard manufacturing processes like roasting and alkalization degrade these native compounds (Shafi et al., 2018; Singh et al., 2024) [21] . Consequently, reformulating chocolate by incorporating external plant-derived bioactive ingredients offers an excellent platform to restore and enhance its nutritional profile (Singh et al., 2024) [22] . Recent studies validate the viability of chocolate as a functional carrier, demonstrating successful fortification using Arthrospira platensis (Özbal et al., 2022) [20] as well as functional spices like liquorice, cinnamon, cardamom, and rosemary (Reddy et al., 2024) [23] . Despite the profound therapeutic properties of Terminalia arjuna bark, its direct consumption faces steep challenges because the raw bark powder possesses an intense bitterness, severe astringency and high moisture sensitivity, which reduce consumer acceptance (Khan et al., 2023; Damale et al., 2026) [25,24] . Incorporating Terminalia arjuna bark extract into a compound chocolate matrix strategically addresses these multi-dimensional challenges. The lipid-continuous phase of chocolate can effectively mask the undesirable sensory attributes of the bark while protecting its moisture-sensitive bioactive compounds. This formulation represents an innovative approach to creating a value-added functional confectionery product. However, creating such a product requires a systematic evaluation of processing conditions, raw material standardization, and final product quality.

Materials and Methodology

Materials

Raw materials and reagents

Healthy, mature outer bark specimen of Terminalia arjuna were harvested from the VNMKV, Parbhani university campus and authenticated by competent botanical authorities to ensure specimen integrity before experimental processing. Commercial-grade dark compound chocolates were procured from local commercial markets. All chemicals, solvents (including food-grade 70% ethanol and citric acid) and reagents utilized during the investigation were of analytical grade.

Equipment and instrumentation

Processing and analytical instrumentation included a laboratory-scale mixer grinder, cabinet dryer, mechanical sieve shaker, hot-air oven, digital analytical balance, and a water bath. Product characterisation was performed using a HunterLab ColorFlex EZ Colorimeter.

Processing of raw material

A. Cleaning, size reduction, and sieve analysis

The collected raw Terminalia arjuna bark samples were processed using the following sequence: 1. Cutting: The Terminalia arjuna stem bark was detached from the tree using a knife. 2. Cleaning: Bark samples were washed thoroughly with distilled water to remove adhering dust, soil, microbial surface contamination, and extraneous matter. Cleaned barks were oven-dried at a controlled temperature to a constant weight.

Pulverization:

The dried bark samples were subjected to mechanical size reduction using a laboratory-scale grinder. 4. Sieving: Sieve analysis was conducted using a standard mechanical sieve shaker to isolate a homogenous, uniform particle size distribution and eliminate oversized fibrous particles.

B. Experimental pre-treatments and optimization

To optimize the yield and maximize the retention of bioactive compounds, the processed bark powder was evaluated under two distinct partial hydrolysis extraction pre-treatments:

Method A: Citric acid hydrolysis 3.0 g of citric acid was dissolved in 300 ml of distilled water to prepare acid solution. A 30.0 g aliquot of the processed bark powder was mixed with the prepared solution (1:10 w/v solid-to-solvent ratio) in a glass vessel. The vessel was tightly sealed and thermally incubated in a water bath at 50-60 °C for 30-45 minutes with periodic shaking. After cooling to room temperature, the mixture was filtered through Whatman No. 1 filter paper. The resulting filtrate was dried in an oven at 50 °C for 12-24 hours to obtain the final concentrated extract powder, as given by Puspitasari et al. (2025) [26] with some modifications.

Method B: Alcohol treatment (partial hydrolysis)

A 30.0 g aliquot of the fine bark powder was placed into an extraction vessel, and 150 ml of food-grade 70% ethanol was added to achieve a 1:5 w/v solid-to-solvent ratio. Extraction was carried out via maceration at room temperature (25 ± 2 °C) for 24 hours under intermittent shaking to enhance mass transfer. The mixture was then filtered through Whatman No. 1 filter paper to remove insoluble plant debris. The resulting filtrate was concentrated via solvent evaporation and dried in a hot-air oven at 40-45 °C. Finally, the dried extract was ground into a homogeneous powder and stored for further analysis (Parmar et al., 2017) [27] .

C. Pre-treatment selection criteria

Weight of recovered pre-treated powder Following the execution of both pre-treatments, fractions were dried, recovered, and quantitatively evaluated for total yield percentage using the standard formula: Weight of sample used Yield (%) = ×100 Based on comparative statistical analysis, Method B (Alcohol treatment) demonstrated a significantly superior yield profile (P < 0.05) compared to Method A. Consequently, the alcohol-treated substrate was selected as the optimized substrate for subsequent chocolate matrix incorporation.

D.

Recipe formulation and standardization

The functional chocolate formulations were standardized by replacing portions of the dark compound chocolates with varying concentrations (1%, 2%, and 3%) of the optimized Terminalia arjuna bark extract. Unfortified compound chocolates served as control.

Where, S 0: 100g dark chocolate, without Terminalia arjuna bark powder

Preparation of functional chocolate

The functional chocolates were prepared by melting dark chocolate using double-boiler method (45-50 °C, 5-10 min). Then the alcohol treated Terminalia arjuna bark powder was added as per the formula-

Sensory evaluation of functional dark chocolate

The sensorial characteristics was studied using the 9-point hedonic scale by the 10 trained panelist member for prepared samples (S 0 , S 1 , S 2 , S 3 ). The sensory attributes evaluated included color and appearance, taste, flavor and overall acceptability.

Evaluation of chemical composition of functional chocolates

The moisture content was estimated by hot air oven method (AOAC, 2000) [28] , fat content using Soxhlet apparatus (AACC, 2000) [29] , protein content estimated by Micro-kjeldhal apparatus as described in AACC (2000) [29] , ash, crude fiber AOAC (2000) [28] and carbohydrate by difference method were evaluated.

Evaluation of functional properties of functional chocolates

The phytochemical profile and antioxidant activity of the chocolates (S 0 and S 2 ) were evaluated using established quantitative methods. Total Phenolic Content (TPC) was determined by the Folin-Ciocalteu colorimetric assay following the protocol described by Sharma et al. (2023) [30] , while Total Flavonoid Content (TFC) was measured using the aluminum chloride method proposed by Chang et al. (2002) [31] . Tannin content was quantified by the titrimetric procedure established by Sharma et al. (2023) [30] . Saponin and alkaloid contents were both determined gravimetrically, adhering to the methodologies of Obadoni et al. (2001) [32] and Harborne (1998) [33] , respectively. Finally, the antioxidant activity was assessed by measuring the percent DPPH radical scavenging axtivity according to the method developed by Brand-Williams et al. (1995) [34] .

Statistical analysis

Each analysis was performed three times (n = 3), with the results reported as mean values along with their standard deviations. The organoleptic sensorial analysis of the prepared chocolates was conducted using 9-point hedonic scale. The S 2 (Selected) chocolate sample outscored all the other formulations, for color and appearance (8.8), taste (8.4), flavor (8.4), texture (8.4) and overall acceptability (8.6). The proximate composition of the prepared S 0 (Control) and S 2 (Selected) chocolates was carried out to understand how Terminalia arjuna bark powder affects the basic nutritional setup of the final product. The prominent increase was seen in the ash content rising from 1.11 to 1.24%, mainly due to the mineral content present in the bark powder. The same trend was observed for the carbohydrate con-tent. The fat content dropped from 34.28 to 33.60%, this was due to the amount of fat in the Terminalia arjuna in powder is less and substituting the chocolate with bark powder dilutes the overall fat content. Similar occurs in case of the fiber and protein content. The data highlights the upward trend for magnesium (130.84 mg/100g), sodium (22.20 mg/100g) and phosphorous content (208.49 mg/100g) in selected (S 2 ) variants, due to the reason that the raw Terminalia arjuna bark powder is naturally a rich source of these macro-minerals. While the calcium, potassium and zinc showed mi-nor decrease into the fortified chocolates, as the small portion of cocoa solids was swapped for the herbal bark powder of Terminalia arjuna which slightly lowers the percentage of these native minerals. The colorimetric values demonstrated a distinct shift towards a darker visual profile upon the addition of the Terminalia arjuna bark powder as L * value decreases. The a * value decreases meaning that the red shifted slightly towards the green. The b * value also decreases meaning the color value shifted from yellow to blue.

Result and Discussion

Organoleptic evaluation of functional milk chocolate

Proximate of prepared functional chocolate

Determination of functional properties of standardized chocolates

Mineral composition of standardized chocolates

Color characteristics of standardized chocolates

Conclusion

Teminalia arjuna is the medicinal tree that possess a wide range of biological activities like anti-oxidant, anti-viral, cardio-tonic, etc. Incorporating this in the suitable food matrix creates a food with functional benefits. The functional chocolate prepared by using Terminalia arjuna bark powder in dark compound chocolate enhances the overall nutritional, mineral and functional profile of the chocolate. Also the chocolate is the confectionery preferred by children, adult and teenagers for consumption. Therefore, the standardized chocolate offers a way of enriching the chocolate with a number of beneficial properties.

Figures

References

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36. How to cite this article: Belode AP, Gadhe KS, Kshirsagar RB, Girhepunje C, Misal RV. Physicochemical, antioxidant and sensory profiling of dark chocolate fortified with Terminalia Arjuna Bark powder. International Journal of Agriculture and Food Science. 2026;8(10):184-189. doi: https://www.doi.org/10.33545/ 2664844X.2026.v8.i10c.2018