96 ISSN 1120-1770 online, DOI 10.15586/ijfs.v33i1.1874 P U B L I C A T I O N S CODON Italian Journal of Food Science, 2021; 33 (1): 96–105 P U B L I C A T I O N S CODON Influence of sugar concentration and sugar type on the polyphenol content and antioxidant activity in spiced syrup preparation Mohd Nazri Zayapor1,2, Aminah Abdullah3, Wan Aida Wan Mustapha2* 1Center of Food Science and Technology Research, MARDI Johor Baharu, Johor, Malaysia; 2Department of Food Sciences, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, Selangor, Malaysia; 3Natural Medicine Research Center, Universiti Islam Malaysia, Selangor, Malaysia *Corresponding Author: Wan Aida Wan Mustapha, Department of Food Sciences, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor Darul Ehsan, Malaysia. Email: wanaidawm@ukm.edu.my Received: 26 April 2020; Accepted: 25 January 2021; Published: 22 February 2021 © 2021 Codon Publications OPEN ACCESS PAPER Abstract Besides their culinary roles, spices in the eastern traditional medical practices serve as a medicinal diet therapy. The polyphenol-rich content of these spices contributes to their antioxidant properties, which convey the thera- peutic elements. Traditionally, these phytonutrients are orally delivered via herbal decoction, including sugar-syr- up-based decoctions. However, the impact of the addition of sugar into polyphenol content and their antioxidant activities are still insufficiently researched. Therefore, this study aimed to evaluate the influence of sugar concen- tration and its types (refined and unrefined) on the polyphenol content and their antioxidant activities. The results of the principal component analysis (PCA) did not exhibit any specific trend on either sugar concentration or its type. As indicative evidence, reducing sugar by less than 25% in such products can be considered for lower-calorie beverage development as a means for healthier diet choice. Keywords: decoction syrup; low-sugar; polyphenol–sugar interaction; sugar-sweetened Introduction Spices are traditionally served as culinary condiments and habitually used in Ayurveda, Persian Tebb-e Sonnati, Traditional Chinese Medicine (TCM), and Unani (Graeco-Arabic) medicinal preparations. Phytochemicals in spices can provide a great deal of medicinal therapy in preventing or treating common ailments. For instance, a mixture of cinnamon, clove, ginger, allspice, nutmeg, and star anise is used in the sugar-sweetened beverage for diet therapy in alleviating metabolic syndrome (Block, 2015). Their phytotherapeutic benefits are administered mostly in crude forms, either orally as infusions (herbal teas), tinctures (alcoholic extracts), decoctions (boiled extracts), and syrups (extracts of herbs made with syrup or honey), or topically applied as poultices, balms, and essential oils (Ahmad et al., 2016; Ramalingum and Mahomoodally, 2014). Among the common liquid forms, Sharbat or Sérbét, which has the Arabic root word shariba or “to drink,” is made from either decoction, infusion of herbal remedies, fruit juice, or flower petal mixtures. There are various types of Sharbat preparation, depending on the type of the substrate to be extracted. Commonly, dry substrates, like most culinary spices, are boiled until one-third of the water is left and allowed to cool before being filtered. The filtered decoction is usu- ally sugar sweetened with two to three parts and further boiled to obtain a syrupy consistency. mailto:wanaidawm@ukm.edu.my Italian Journal of Food Science, 2021; 33 (1) 97 Influence of sugar concentration and sugar type cause an alarming glycemic response in patients with metabolic syndrome. Nevertheless, the negative health impact of commonly consumed sugary beverages on the risk factors for meta- bolic syndrome cannot be validated with adequate clini- cal evidence (Angelopoulos et al., 2016; Della Corte et al., 2018) to confirm that the added sugar products are the chief culprits for the metabolic imbalance. While it is health-wise beneficial to consume added sugars beyond the average level, the reduction in sugar intake accompa- nied by restricted calorie intake may be more beneficial (Rippe and Angelopoulos, 2016). Therefore, this prelimi- nary study was to evaluate the effect of sugar concentra- tion and the sugar type by examining their influence on polyphenol contents and their antioxidant activities. Material and Method Four culinary spices comprising Chinese cinnamon (Cinnamomum cassia), dried ginger (Zingiber officinale), green cardamom (Elettaria cardamomum), sweet fen- nel (Foeniculum vulgare), and six types of sugar, namely, refined white sugar (RWS), filtered sugarcane brown sugars (FBS), unfiltered sugarcane brown sugar (NBS), organic molasses (OMO), Javanese coconut sugar (JCS), and Malaccan coconut sugar (MCS) were acquired from Jaya Grocer store, Bangi Gateway. Spiced syrup preparation Dirt and debris were removed from the spices by clean- ing under running tap water, and the excess water was drained before they were air-dried. A mixture of spices, comprising 10% (w/v) cinnamon and 5% (w/v) other spices, was added to boiling sugar syrup (110°C) before it was simmered for 240 min at 80°C. The spice mixture was directly used without pounding into smaller sizes based on a traditional technique to avoid a strong flavor. The spiced syrup was prepared (Figure 1) according to a traditional recipe, whereby sugars were added to filtered water at ratios of 1:2 kg/L, 1:3 kg/L, 1:4 kg/L, and 1:6 kg/L to produce syrups at the corresponding concentrations (Table 1). The range of sugar additions was according to traditional practices in preparing a sugar-based decoc- tion. However, the most common practice is to mix one part of sugar with four parts of water (25%, w/v). The non-spiced 25% sugar syrups (all selected sugar types) were boiled and simmered in the same way as the spiced syrup was prepared and were considered as the posi- tive control (control, CTRL [sugar type]), and the spice decoction without sugar syrup (0% sugar) was considered as the negative control. The spiced syrup was then fil- tered with a clean muslin cloth before being bottled and Addition of sugar to the phytochemical extraction step is relatively similar to the fruit osmotic dehydration tech- nique, whereby sugar syrup is used as an osmotic agent (Pereira et al., 2014) to produce an aromatic and flavor- ful herbal syrup or extract (Geng and Zhou, 2010). The range of osmotic agent additions were from 2 to 50% of the herbal substrate, but the study did not indicate as to when the sugar should be added. This syrup extraction was traditionally meant to improve the flavor and aroma of the herbal extract but was not intended to enhance the extractability of antioxidant compounds or specif- ically the polyphenols. Previously, it was known that a hot aqueous extract of several spices (non-sugar addi- tion) had a high antioxidant activity that was probably due to the phenolic and flavonoid compounds (Kim et al., 2011). Later, a study revealed that the addition of sugar before or during processing might play a significant role in enhancing the polyphenol extraction, and simulta- neously the antioxidant activities instead of improving only the gastronomic qualities (Loncaric et al., 2014). However, studies that evaluate the impact of sugar addi- tion on polyphenols’ content and their antioxidant activ- ities are still limited to ensure this positive enhancement. The latest findings showed that there were only slight polyphenol losses during food processing (Szymanowska et al., 2017; Zeng et al., 2017) to unapprehensive changes in antioxidant capacities (Baroni et al., 2018) upon the introduction of sugar, which was probably due to some degree of protective mechanism. A similar mechanism was observed in chokeberry juice that was added with polysaccharide as the clarifying agent (Lachowicz et al., 2018). However, the polyphenolic content became less as soon as the natural sedimentation appeared, resulting in polyphenol binding to polysaccharides. A list of sugar-loaded syrups showed promising thera- peutic potentials as novel sources of antioxidant polyphe- nols. Among the examples are the traditional prickly pear (Opuntia ficus indica) and the pink Tecoma (Tabebuia impetiginosa) inner-bark syrups were advocated for their anti-tumor potential (Dhaouadi et  al., 2013; Pires et al., 2015), and the English plantain (Plantago lanceo- late) leaf syrup was recommended for combating com- mon cold (Mansoor et al., 2017). A polyphenol-rich syrup can potentially be a novel functional ingredi- ent like the bog bilberry (Vaccinium uliginosum) syrup (Malvidin-3-O-glucoside, petunidin-3-O- glucoside, and delphinium-3-O-glucoside), which is the chief flavonoid glucoside that is used to enrich wine for antioxidant capacities (Liu et al., 2015). Carbohydrates can enhance the bioactivity and the bioavailability of polyphenol com- pounds through increased fermentation by microbiota in the large intestine, especially the oligosaccharides (Zhang et al., 2014). However, the primary concern is monosaccharide and disaccharide, which are readily absorbed and digested in the small intestine and may 98 Italian Journal of Food Science, 2021; 33 (1) Zayapor MZ et al. Table 1. Sugar ratio to sugar percentage conversion. Sugar to water ratio (kg/L) Sugar used (g/1000 mL) Sugar percentage, % (w/v) 1:2 500.0 50.0 1:3 333.3 33.3 1:4 250.0 25.0 1:6 166.7 16.7 Sugar and filtered water mixture Boiling Simmering at 80°C, 240 minutes Filtration Bottling Addition of cleaned and air-dried mixture of spices Figure 1. Flow chart of spiced syrup processing stored at room temperature until use. Each spiced syrup was prepared in triplicates. The uncertainty due to spice sampling heterogeneity was kept below 13%, as recom- mended by Bodnar et al. (2013). Determination of the total phenolic content The Folin–Ciocalteu (FC) method (Singleton et al., 1999) was used to quantify the phenolic content with slight modification on the reactant volumes for 96-well micro- plate suitability. A hundred microliters of 10% FC reagent were mixed with 20 µL of blank (deionized water)/ standard/filtered syrup (diluted 50-fold with deion- ized water). After 5 min, 80 μL of 7.5% sodium carbon- ate was added and incubated at 37⁰C for 120 min. This spectrophotometric measurement was carried out at λ = 765 nm (Spectrostar Nano, BMG Labtech, Germany). The quantification of TPC by using the calibration curve of gallic acid was calculated by MARS Data Analysis Software Version 2.01. The concentration for the cali- bration curve ranged from 250 to 4000 ppm. The results were expressed as micrograms of gallic acid equivalents (GAE)/mL of syrup. Antioxidant activity determination Multiple spectrometric techniques were employed to evaluate the spice polyphenol antioxidant activities by three different mechanisms: organic radical scav- enging, ferric reducing capacity, and peroxyl radical scavenging. It was essential to evaluate the antioxidative compounds by using various techniques due to the unique identity of each polyphenol with different action mechanisms against oxidative substances. With these multiple antiox- idant determinations, an almost complete profile of anti- oxidants can be attained. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activities assay The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scav- enging activities were determined according to Kodama et al. (2010). The 0.5 mM DPPH was pre-adjusted with methanol (1.00 ± 0.1 absorbance unit) before being mixed with diluted syrup (20-fold with distilled water) at a ratio of 9:1. After 30 min, the absorbance of mixtures was measured at 515 nm, and the DPPH radical scaveng- ing activities were calculated as mM Trolox equivalent (TE)/mL of syrup. Ferric reducing antioxidant power assay A ferric reducing antioxidant power (FRAP) assay of spiced syrup was based on Musa et al. (2011), which required the FRAP reagent to be freshly prepared by sequentially mixing one part of 20 mM FeCl3. 6H2O with one part of 10 mM 2,4,6-tri(2-pyridyl)-s-triazine (TPTZ) and thoroughly mixing with 10 parts of 300 mM acetate buffer, at pH 3.6. A freshly prepared FRAP reagent at a 1950 μL was mixed with 50 μL diluted syrup before a 30-min incubation at room temperature. The absorbance changes against the blank (distilled water) were mea- sured at 595 nm. The Trolox calibration curve (3.125 mM to 100 mM) was used to equivalently estimate the FRAP reducing capacities, with results expressed as mM Trolox equivalents (TE)/mL of syrup. Oxygen radical absorbance capacity assay For the oxygen radical absorbance capacity (ORAC) value determinations, 150 µL of 10 nM fluorescein solution was mixed with 25 µL diluted syrup in a 96-well black microplate, as described by Payne et al. (2013). After incubation for 30 min at 37⁰C, the fluorescence degra- dation measurements were executed at the emission and excitation of 520 nm and 485 nm, respectively, by using a microplate reader (FLUORstar Omega, BMG Labtech, Germany). Automatic injection of 240 mM AAPH (25 µL) at the fourth cycle was made before measurement resumption, and the calculated results were expressed as μM Trolox equivalents (TE)/mL of syrup. Italian Journal of Food Science, 2021; 33 (1) 99 Influence of sugar concentration and sugar type Statistical analysis Data collected as means of triplicate readings were sub- jected to analysis of covariance (ANCOVA), correla- tion analysis by Pearson’s correlation coefficient, and data analysis by principal component analysis (PCA) by using XLSTAT Premium 2018.1.4913 (Addinsoft Inc., New York, NY, USA). The ANCOVA was applied to evaluate whether TPCs were varied with sugar con- centrations (quantitative) and types (qualitative), and verify a sensible linear model. The exploratory statis- tical tool, PCA, was applied to observe any typical or divergent features or degree of similarities among a set of variables. Results and Discussions Effect of sugar concentration and sugar type on the TPC of spiced syrups Figure 2 shows that all CTRL sugars had a TPC that ranged from 500 to 1000 GAE mg/mL, except for RWS, which was the least with < 500 GAE mg/mL. Java coco- nut sugar, JCS had the highest TPC among the selected sugar types of approximately 1000 GAE mg/mL. Most brown or unrefined sugars contained a significant amount of polyphenol (P < 0.05), compared to refined sugar (Kongkaew et al., 2014; Choong et al., 2016). Brown sugar from sugarcane and coconut are commonly used in traditional medicine preparation as a sweetener because it is believed to possess therapeutic benefits. However, the 0% sugar decoction had 2-fold to 5-fold higher TPC than each CTRL sugar, and showed that sugar syrup of any type could contribute to a smaller portion of TPC value in the spiced syrup. Overall, the higher the sugar concentration, the higher was the polyphenol content, except for OMO, which showed an opposite trend. This contradictory effect on OMO might be attributed to the OMO polyphenol antagonistic effect, reflecting the reduction of content. Meanwhile, in the RWS syrup, the TPC showed a unique bell-shaped trend, whereby intermediate sugar concen- trations of 25 and 33.3% were exhibited among the highest contents (> 2500 GAE mg/mL). At higher sugar concen- trations, the TPC high and low values were possibly due to the osmotic pressure, which allowed for desorption or prevention of spice polyphenols into the sugar syrup. It was postulated that the TPC decrement in the spiced syrup as the sugar concentration decreased was due to spice polyphenol degradation in the low solute matrix. However, the enhancement of the polyphenolic content in pumpkin by using concentrated flowering quince juice was attributed to the penetration of the polyphenol-rich osmotic solution into the pumpkin, which provided some stability (Lech et al., 2018). The inconsistent variance among TPC in a syrup of different sugar concentrations and types might occur due to the interaction of Folin– Ciocalteu reagents with any reducing or readily oxidiz- able substances of nonphenolic compounds, causing an aberration of the total phenolic content (Essawet et al., 2015; Ho et al., 2017; Ramachandran and Nagarajan, 2014). It was postulated that different profiles of mono- saccharide and disaccharide could play some interaction with the spice-polyphenols via a covalent or non- covalent bond, and thus, reflecting some variation in free polyphe- nol content. The ANCOVA analysis showed that 63% of TPC vari- ability (Table 2) was explained by sugar concentrations and types. However, the latter variability and interac- tion between sugar concentrations and types did not 0.00 C T R L ( 1 0 % S U G A R ) C T R L R W S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L F B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L N B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S Sugar concentration (%, w/v) 1 6 .7 % R W S C T R L O M O ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L M C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L J C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S 500.00 1000.00 1500.00 2000.00 T P C ( G A E µ g /m L ) 2500.00 3000.00 3500.00 4000.00 Figure 2. Total phenolic content (TPC) of spiced syrups prepared from different type of sugars and concentrations. 100 Italian Journal of Food Science, 2021; 33 (1) Zayapor MZ et al. significantly explain the TPC variability. Furthermore, the equation of the TPC linear model is proposed, TPC = 733.60 × Type of Sugar-C-FBS + 1070.85 × Type of Sugar-C-JCS + 602.73 × Type of Sugar-C-MCS + 872.03 × Type of Sugar-C-NBS + 880.13 (1) Since the type of sugar did not carry significant infor- mation, the proposed model (Equation 1) was rejected. Based on Type 3 sum of squares, the interaction vari- able (type of sugar × sugar concentration) was the most influential contributor to the insignificant variation. Therefore, RWS can still be utilized in the production of the spiced syrup without a considerable loss in the TPC. One can avoid brown sugar in the mass production of the spiced syrup as it was uneconomically feasible. Antioxidant capacities (DPPH scavenging, FRAP reducing activities, and ORAC values) of spiced syrups From the summary of variable analysis (Table 2), 44% of the ORAC value variability was explained by the two explanatory variables (sugar concentration and type). The variability of the ORAC values in the spiced syrup was similar to TPC variability, which was not signifi- cantly explained by the type of sugar and the interaction variables of sugar concentration and sugar type. It may explain that the TPC content may have a correspond- ing influence on the ORAC values. All the spiced sugar syrups showed decrement trends in the ORAC values, excluding three negligible samples that showed incre- ments (Figure 3). Meanwhile, the DPPH scavenging and the FRAP reducing capacities had 40 and 27% variabil- ities, respectively, explained. The sugar concentration did not give significant variation information in DPPH scavenging capacity (P > 0.05). However, the type of sugar played an influential role in explaining the variation in the capacity based on Type 3 sum of squares. It con- tradicted the FRAP reducing capacity, whereby the sugar concentration significantly influenced its variability. The DPPH scavenging capacity of the spiced syrup showed decreasing trends in the RWS (0.6 to 72%) but the opposite trend in the MCS-based spiced syrup (31–53%), as compared to the control zero-sugar spiced decoction (Figure 4). The rest of the four-brown sug- ar-based decoctions exhibited mixed trends in the scav- enging capacities. All FRAP reducing capacities showed reductions as negatively influenced by sugar, excluding two samples from the different types of sugar (Figure 5). Visualization of the correlations between variables by using PCA For distinguishing the spiced syrup-based TPC and anti- oxidant activities, the PCA was employed on the Pearson correlation matrix. As a classification/discrimination method, PCA permits a simplified data representation over a complex data matrix of the spiced syrup with different sugar concentrations and types of sugar use. The first two principal components (PC), PC1 and PC2, explained a total of 78.4% variance (Figure 6). Both PCs were almost equally explained, whereby the initial PC, explained up to 41.7% and the latter up to 36.7%. PC1 has a strong positive correlation with the TPC and all antiox- idant variables, whereby the ORAC values have the high- est correlation with the TPC (P < 0.514) (Table 3). The DPPH scavenging and FRAP reducing capacities have the least correlation with the other two variables and are negatively correlated with each other (P < −0.465). Table 2. Summary of ANCOVA for all variables. TPC DPPH SC FRAP RC ORAC V R² 0.627 0.403 0.273 0.439 F 13.098 14.186 2.019 5.681 Pr > F < 0.0001 < 0.0001 0.067 0.000 Sugar concentration (%) F 0.730 10.740 Pr > F 0.401 0.003 Sugar type F 1.814 2.726 1.106 Pr > F 0.150 0.018 0.395 Sugar concentration (%) * Sugar type F 2.234 1.226 1.262 Pr > F 0.084 0.321 0.320 ANCOVA, analysis of covariance; TPC, total phenolic content; ORAC, oxygen radical absorbance capacity; DPPH, 2, 2-Diphenyl-1- picrylhydrazyl; FRAP, ferric reducing antioxidant power. Italian Journal of Food Science, 2021; 33 (1) 101 Influence of sugar concentration and sugar type O R A C a ct iv ity ( T rx E µ g /m L ) C T R L ( 1 0 % S U G A R ) C T R L R W S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L F B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L N B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S Sugar concentration (%, w/v) 1 6 .7 % R W S C T R L O M O ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L M C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L J C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S 0.00 5000.00 10000.00 15000.00 20000.00 25000.00 30000.00 35000.00 40000.00 45000.00 50000.00 Figure 3. ORAC values of spiced syrups prepared from different type of sugar and concentration. C T R L ( 1 0 % S U G A R ) C T R L R W S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L F B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L N B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S Sugar concentration (%, w/v) 1 6 .7 % R W S C T R L O M O ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L M C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L J C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S 0.00 10 00.00 2500.00 3000.00 4000.00 6000.00 5000.00 D P P H r a d ic a l s ca ve n g in g a ct iv ity (T rx E µ g /m L ) Figure 4. DPPH radical scavenging activities of spiced syrups prepared from different type of sugar and concentration. C T R L ( 1 0 % S U G A R ) C T R L R W S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L F B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L N B S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S Sugar concentration (%, w/v) 1 6 .7 % R W S C T R L O M O ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L M C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S C T R L J C S ( 5 0 % ) 5 0 .0 % R W S 3 3 .3 % R W S 2 5 .0 % R W S 2 0 .0 % R W S 1 6 .7 % R W S 0.00 –2.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 F R A P a ct iv ity ( T rx E m M /m L ) Figure 5. FRAP values of spiced syrups prepared from different type of sugar and concentrations. 102 Italian Journal of Food Science, 2021; 33 (1) Zayapor MZ et al. Table 3. Correlation matrix (Pearson [n – 1]). Variables TPC DPPH SC FRAP RC ORAC V TPC 1 0.201 0.255 0.514 DPPH SC 0.201 1 −0.465 0.220 FRAP RC 0.255 −0.465 1 0.156 ORAC V 0.514 0.220 0.156 1 Values in bold are different from 0 with a significance level alpha = 0. ANCOVA, analysis of covariance; TPC, total phenolic content; ORAC, oxygen radical absorbance capacity; DPPH, 2, 2-Diphenyl-1- picrylhydrazyl; FRAP, ferric reducing antioxidant power. Figure 6. PCA biplots of spiced syrups made from different type of sugar and concentrations versus TPC and antioxidant parameters. From the biplot (Figure 6), there was no specific classifi- cation based on either concentration of sugar or the type of sugar exhibited on both the PCs. However, two clus- ters can be observed in the positive and negative loadings of PC2 without specific classification in each cluster. The PCA analysis results showed that either sugar concentra- tion or sugar type significantly influences the TPC and their antioxidant activities. Only hard ingredients, either from bark, seed, or heart- wood, are traditionally subjected to simmering. In con- trast, the soft or leafy ingredients are infused in boiled water to extract bioactive compounds. The duration of the former may take hours as compared to the latter, which only takes a few minutes. The decoction process started after the sugar syrup was boiled and the spices were added and terminated after 4 h of simmering. The spices were introduced into the boiled sugar syrup to obtain the glassy clear syrup in culinary practice. The point at which the sugar was added did not significantly impact taste (Madhavamenon and Maliakel, 2015). However, from the study observation, the addition of sugar toward the end of the decoction process only retained the liquid turbid- ity, which was visually less appealing. Previous studies showed that mild thermal exposure, such as in the fortification of sugar syrup with functional fruit- based ingredients, retained its high polyphenol content (Tarko et al., 2015). Those that were traditionally prepared fruit-based beverages also showed more pronounced anti- oxidant capacities than the commercially processed juice (Marjanović et al., 2015). A traditional Calabrian fig syrup made from fresh figs’ (Ficus carica) aqueous slurry, pro- long boiling until the syrup concentrate has a high poly- phenol content (3.92 mg of GAE per gram of syrup) with desirable sensorial qualities (Puoci et al., 2011). It seemed to be not much affected by the lengthy thermal treatment. Therefore, the effect of the thermal treatment on the poly- phenol content of the spiced syrup was not carried out. The decrement trend in radical scavenging capacities Italian Journal of Food Science, 2021; 33 (1) 103 Influence of sugar concentration and sugar type the oxidation of phenolic compounds (Le Bourvellec and Renard, 2012), which probably occurred in the spiced syrups and highly corresponded to the low antioxidant activities. It showed that both types of association have a determinant effect on the quality level of polyphenol-rich beverages. Sugars engage in the stability of polyphenols in the form of conjugated sugar or glycoside via a glyco- sidic bond (excluding the subclass of catechins) to one or more hydroxyl groups (Martin, 2009). Therefore, the addition of sugar may inhibit the autoxidation of poly- phenols and contribute to high antioxidant activities in some spiced syrups. However, the profound antioxidant activities were not solely contributed by inhibition of autoxidation. It may partially be due to the advantage of additive or synergistic effect of different phenolics, fla- vonoids, reducing sugars, and vitamin C profiles like in the must of dried grapes (Peinado et al., 2010), whereby an antagonist effect may be found in the spiced syrup for the reduced antioxidant activities. The glucoside forms (sugar conjugated) are likely to offer more therapeutic benefits than the aglycones (nonsugar conjugated poly- phenol), as they are more bioaccessible, bioavailable, and stable under in vivo conditions (Fernandes et al., 2017). In addition, the sugar moiety of glycoside can enhance the bioavailability, but dimerization weakens the ability. However, glycosides are usually weaker in vitro antioxi- dants than aglycones (Shashank and Pandey, 2013); this explains why some spiced syrups have lower antioxidant activities than the zero-sugar spiced decoction. Conclusions Various sugar concentrations and different types of sugar used in the extraction of polyphenols to produce the spiced syrup may influence the TPC and antioxidant activities to a particular extent with mixed variabilities, as observed from the results of the ANCOVA and the PCA. The weaving trends may uniquely contribute to the different profile components of sugar and water-soluble phytochemicals from the spice mixtures. These biochem- ical complexities of molecular weight and structure- related reaction produced a different level of antioxidant activities. From the compiled results of the ANCOVA and the PCA, it can be suggested that a sugar concen- tration of less than 25% is adequate to prepare the spiced syrup with many antioxidant activities, even while using the RWS. Moreover, further studies are required to bet- ter conclude the sugar–polyphenol interaction as in vivo. Acknowledgment The authors are grateful for the research grant from the Natural Medicine Research Center, Universiti Islam Malaysia, Cyberjaya, and Antioxidant Lab, Faculty might be attributed to the glycosidic bond formation through the condensation reaction between a hydroxyl group of sucrose molecule and the hydroxyl group of phenolic compounds to form glycoside compounds. However, the increments were probably the result of phenolic reduced-form compounds due to the oxidation of phenolic compounds and sucrose molecules (Shalaby et al., 2016). In addition, Jlassi et al. (2016) suggested that the dose-dependent manners of the polyphenol bioactiv- ities may vary due to the polyphenol functional site reac- tion with ionized sucrose. Besides, the polyphenol profile may influence disaccharide action and give variability in antioxidant activities (Pengseng et al., 2011). Sugar-sweetened beverage consumption still gives a stir to the national health issues. Simultaneously, polyphe- nol-rich food intakes are linked with a positive impact on consumers’ health. However, sugar addition to polyphe- nol-rich food products, especially beverages, can change the health benefits. Some suggested that the presence of sugars generally increases the bioavailability of polyphe- nols, but product formulation may influence the sugar impact to a certain degree (Ackar et al., 2013). This study observed a specific range of sugar concentration and sugar type that accounts for a certain extent of the TPC. These two variables may introduce different profiles of disaccharides, which in turn react to a certain extent. It was not only the type of disaccharide that was reported to have an essential impact on phenolics but also their chemical isomers since the chemical behavior can be dif- ferent in complex food matrices (Zlatić et al., 2017). For example, maltose and trehalose are isomers of sucrose. The latter isomer has a more hydrophilic site than the former, leading to a higher interaction with hydrophilic polyphenols. In the study spice syrup, hydroxycinnamic acids derived from cinnamon, which was hydrophilic (Teixeira et al., 2013), probably had more sugar inter- action. In addition to the saccharide structure-related factors, evidence showed that the polyphenol interac- tion with saccharides was influenced by the polyphenol molecular weight (monomeric or polymeric) (Amoako and Awika, 2016). It was found that polymeric polyphe- nol (molecular weight, MW > 1000) polysaccharides via hydrogen bonding and hydrophobic interactions formed nondigestible complexes. This positive interaction can reduce calorie density and may help modulate glucose metabolism. Therefore, future research on the conse- quences of polyphenol–sugar interaction should uncover a positive impact on health. In the food matrix, polyphenols can have either a covalent or noncovalent association with polysaccharides. A com- bination of hydrophobic interaction and hydrogen bond (between the phenolic hydroxyl group and the polysac- charide oxygen atom) can result in a noncovalent associ- ation. Simultaneously, the covalent association involved 104 Italian Journal of Food Science, 2021; 33 (1) Zayapor MZ et al. Dhaouadi, K., Raboudi, F., Funez-Gomez, L., Pamies, D., Estevan, C., Hamdaoui, M. and Fattouch S., 2013. Polyphenolic extract of barbary-fig (Opuntia ficus-indica) syrup: RP-HPLC-ESI-MS analysis and determination of antioxidant, antimicrobial and cancer-cells cytotoxic potentials. Food Analytical Methods 6: 45–53. https://doi.org/10.1007/s12161-012-9410-x Essawet, N., Cvetkovic, D., Velicanski, A., Canadanovic-Brunet,  J., Vulic, J., Maksimovic, V. and Markov, S., 2015. Polyphenols and antioxidant activities of Kombucha beverage enriched with Coffeeberry® extract. Chemical Industry and Chemical Engineering Quarterly 21: 399–409. http://www.doiserbia.nb.rs/ Article.aspx?ID=1451-93721400042E Fernandes, I., Pèrez-Gregorio, R., Soares, S., Mateus, N., De Freitas,  V., Santos-Buelga, C. and Feliciano, A.S., 2017. Wine flavonoids in health and disease prevention. Molecules 22 (2): 1–30. https://doi.org/10.3390/molecules22020292 Geng, L. and Zhou, Y.L., 2010. Use of saccharides in extraction of tea or herbaceous plants. 1–8. http://v3.espacenet.com/ textdoc?DB=EPODOC&IDX=US2010136191 Ho, G.T.T., Yen Nguyen, T.K., Kase, E.T., Tadesse, M., Barsett, H. and Wangensteen, H., 2017. Enhanced glucose uptake in human liver cells and inhibition of carbohydrate hydrolyzing enzymes by nordic berry extracts. Molecules 22 (10): 1–15. https://doi. org/10.3390/molecules22101806 Jlassi, H., Dhaouadi, K. and Laaribi, M., 2016. Short cooking time increases, but sucrose or sweetener counteracts the radical scavenging activities of tea decoction as compared to tea infu- sion. Journal of Food and Nutritional Disorders 5, available at: http://www.scitechnol.com/peer-review/short-cooking-time- increasesbut-sucrose-or-sweetenercounteracts-the-radicals- cavenging-activities-of-teadecoction-as-compared-to-te-RY0i. php?article_id=5574 Kim, I.S., Yang, M.R., Lee, O.H. and Kang, S.N., 2011. Antioxidant activities of hot water extracts from various spices. International Journal of Molecular Sciences 12: 4120–4131. https://doi. org/10.3390/ijms12064120 Kodama, D., Gonçalves, A., Lajolo, F. and Genovese, M., 2010. Flavonoids, total phenolics and antioxidant capacity: com- parison between commercial green tea preparations. Ciência e Tecnologia de Alimentos 30: 1077–1082. https://doi. org/10.1590/S0101-20612010000400037 Kongkaew, S., Chaijan, M. and Riebroy, S., 2014. Some character- istics and antioxidant activity of commercial sugars produced in Thailand. KMITL Science and Technology Journal. 14: 1–9. http://stj.kmitl.ac.th/Archives/vol14no1/00089_reformatted.pdf Lachowicz, S., Oszmiański, J. and Kalisz, S., 2018. Effects of various polysaccharide clarification agents and reaction time on con- tent of polyphenolic compound, antioxidant activity, turbidity and colour of chokeberry juice. Lebensmittel-Wissenschaft & Technologie 92: 347–360. https://doi.org/10.1016/j.lwt.2018. 02.054 Lech, K., Figiel, A., Michalska, A., Wojdylo, A. and Nowicka, P., 2018. The effect of selected fruit juice concentrates used as osmotic agents on the drying kinetics and chemical proper- ties of vacuum-microwave drying of pumpkin. Journal of Food Quality 2018: 1–11. https://doi.org/10.1155/2018/7293932 of Science and Technology, Universiti Kebangsaan Malaysia for providing the best-equipped facilities for the antioxidant analysis. The authors would also like to express their great appreciation to Assoc. Prof. Dr. Yusof Maskat for reviewing this paper and giving his thought- ful comments. References Ackar, D., Valek Lendić, K., Valek, M., Šubarić, D., Miličević, B., Babić, J. and Nedić, I., 2013. Cocoa polyphenols: can we con- sider cocoa and chocolate as potential functional food? Journal of Chemistry. 2013: 1–7. https://doi.org/10.1155/2013/289392 Ahmad, I., Shamsi, S. and Zaman, R., 2016. Sharbat: an Important Dosage Form of Unani System of Medicine Methodology of preparation of Sharbat of different drugs in USM. Medical Journal of Islamic World Academy of Sciences 24: 83–88. https://doi.org/10.5505/ias.2016.91129 Amoako, D. and Awika, J.M., 2016. Polyphenol interaction with food carbohydrates and consequences on availability of dietary glucose. Current Opinion in Food Science 8: 14–18. https://doi. org/10.1016/j.cofs.2016.01.010 Angelopoulos, T., Lowndes, J., Sinnett, S. and Rippe, J., 2016. Fructose containing sugars at normal levels of consumption do not effect adversely components of the metabolic syndrome and risk factors for cardiovascular disease. Nutrients 8: 179. http:// www.mdpi.com/2072-6643/8/4/179 Baroni, M.V., Gastaminza, J., Podio, N.S., Lingua, M.S., Wunderlin, D.A., Rovasio, J.L., Dotti, R., Rosso, J.C., Ghione, S. and Ribotta, P.D., 2018. Changes in the antioxidant properties of quince fruit (Cydonia oblonga Miller) during Jam Production at Industrial Scale. Journal of Food Quality 2018: 1–9. https://doi. org/10.1155/2018/1460758 Block, E.F., 2015. Food therapy for the Traditional Chinese Medicine (TCM) diagnosis of dampness. International Journal of Complementary and Alternative Medicine 2: 2–4. https://doi. org/10.15406/ijcam.2015.02.00050 Bodnar, M., Namieśnik, J. and Konieczka, P., 2013. Validation of a sampling procedure. TrAC Trends in Analytical Chemistry 51: 117–126. https://doi.org/10.1016/j.trac.2013.06.011 Choong, C. C., Anzian, A, Che Wan Sapawi, C. W. N. S., Meor Hussin, A. S., 2016. Characterization of Sugar from Arenga pinnata and Saccharum officinarum sugars. International Food Research. Journal 23:1642–52. http://ifrj.upm.edu.my/23%20 (04)%202016/(39).pdf Le Bourvellec, C. and Renard, C.M.G.C., 2012. Interactions between polyphenols and macromolecules: quantification methods and mechanisms. Critical Reviews in Food Science and Nutrition 52: 213–248. https://doi.org/10.1080/10408398.2010.499808 Della Corte, K., Perrar, I., Penczynski, K., Schwingshackl, L., Herder,  C. and Buyken, A., 2018. Effect of dietary sugar intake on biomarkers of subclinical inflammation: a systematic review and meta-analysis of intervention studies. Nutrients 10: 606. http://www.ncbi.nlm.nih.gov/pubmed/29757229%0A; http:// www.mdpi.com/2072-6643/10/5/606 https://doi.org/10.1007/s12161-012-9410-x� http://www.doiserbia.nb.rs/Article.aspx?ID=1451-93721400042E� http://www.doiserbia.nb.rs/Article.aspx?ID=1451-93721400042E� https://doi.org/10.3390/molecules22020292� http://v3.espacenet.com/textdoc?DB=EPODOC&IDX=US2010136191� http://v3.espacenet.com/textdoc?DB=EPODOC&IDX=US2010136191� https://doi.org/10.3390/molecules22101806� https://doi.org/10.3390/molecules22101806� http://www.scitechnol.com/peer-review/short-cooking-time-increasesbut-sucrose-or-sweetenercounteracts-the-radicalscavenging-activities-of-teadecoction-as-compared-to-te-RY0i.php?article_id=5574� http://www.scitechnol.com/peer-review/short-cooking-time-increasesbut-sucrose-or-sweetenercounteracts-the-radicalscavenging-activities-of-teadecoction-as-compared-to-te-RY0i.php?article_id=5574� http://www.scitechnol.com/peer-review/short-cooking-time-increasesbut-sucrose-or-sweetenercounteracts-the-radicalscavenging-activities-of-teadecoction-as-compared-to-te-RY0i.php?article_id=5574� http://www.scitechnol.com/peer-review/short-cooking-time-increasesbut-sucrose-or-sweetenercounteracts-the-radicalscavenging-activities-of-teadecoction-as-compared-to-te-RY0i.php?article_id=5574� https://doi.org/10.3390/ijms12064120� https://doi.org/10.3390/ijms12064120� https://doi.org/10.1590/S0101-20612010000400037� https://doi.org/10.1590/S0101-20612010000400037� http://stj.kmitl.ac.th/Archives/vol14no1/00089_reformatted.pdf� https://doi.org/10.1016/j.lwt.2018.02.054� https://doi.org/10.1016/j.lwt.2018.02.054� https://doi.org/10.1155/2018/7293932� https://doi.org/10.1155/2013/289392� https://doi.org/10.5505/ias.2016.91129� https://doi.org/10.1016/j.cofs.2016.01.010� https://doi.org/10.1016/j.cofs.2016.01.010� http://www.mdpi.com/2072-6643/8/4/179� http://www.mdpi.com/2072-6643/8/4/179� https://doi.org/10.1155/2018/1460758� https://doi.org/10.1155/2018/1460758� https://doi.org/10.15406/ijcam.2015.02.00050� https://doi.org/10.15406/ijcam.2015.02.00050� https://doi.org/10.1016/j.trac.2013.06.011� http://ifrj.upm.edu.my/23%20(04)%202016/(39).pdf� http://ifrj.upm.edu.my/23%20(04)%202016/(39).pdf� https://doi.org/10.1080/10408398.2010.499808� http://www.ncbi.nlm.nih.gov/pubmed/29757229%0Ahttp://www.mdpi.com/2072-6643/10/5/606� http://www.ncbi.nlm.nih.gov/pubmed/29757229%0Ahttp://www.mdpi.com/2072-6643/10/5/606� Italian Journal of Food Science, 2021; 33 (1) 105 Influence of sugar concentration and sugar type N., 2011. Antioxidant activity of a mediterranean food prod- uct: “fig syrup.” Nutrients 3: 317–329. https://doi.org/10.3390/ nu3030317 Ramachandran, P. and Nagarajan, S., 2014. Quality character- istics, nutraceutical profile, and storage stability of Aloe gel-papaya functional beverage blend. International Journal of Food Sciences and Nutrition 2014: 1–7. https://doi. org/10.1155/2014/847013 Ramalingum, N. and Mahomoodally, M.F., 2014. The therapeutic potential of medicinal foods. Advances in Pharmacological and Pharmaceutical Sciences 2014: 1–18, available at: http://www. ncbi.nlm.nih. gov/pubmed/24822061%5Cn Rippe, J.M. and Angelopoulos, T.J., 2016. Relationship between added sugars consumption and chronic disease risk factors: cur- rent understanding. Nutrients 8: 1–19. https://doi.org/10.3390/ nu8110697 Shalaby, E.A., Mahmoud, G.I. and Shanab, S.M.M., 2016. Suggested mechanism for the effect of sweeteners on radical scaveng- ing activity of of phenolic compounds in black and green tea. Frontiers in Life Science 9: 241–251. http://www.agriculture- journals.cz/publicFiles/105870.pdf Shashank, K. and Pandey, A.K., 2013. Chemistry and biologi- cal activities of flavonoids. Hindawi Scientific World Journal 2013: 533–548. http://linkinghub.elsevier.com/retrieve/pii/ S0924224405002049 Singleton VL, Orthofer R, Lamuela-Raventós, 1999. Analysis of total phenols and other oxidation substrates and antioxidants by means of folin-ciocalteu reagent. In: Oxidants and Antioxidants Part A. Academic Press, pp 152–78. https://www.sciencedirect. com/science/article/pii/S0076687999990171 Szymanowska, U., Baraniak, B. and Gawlik-Dziki, U., 2017. Changes in the level and antioxidant activity of polyphenols during stor- age of enzymatically treated raspberry juices and syrups. Acta Scientiarum Polonorum. Technologia Alimentaria 16: 269–282. https://doi.org/10.17306/J.AFS.2017.0491 Tarko, T., Duda-Chodak, A., Semik, D. and Nycz, M., 2015. The use of fruit extracts for production of beverages with high antioxidative activity. Potravinarstvo 9: 280–283. https://doi. org/10.5219/480 Teixeira, J., Gaspar, A., Garrido, E.M., Garrido, J. and Borges, F., 2013. Hydroxycinnamic acid antioxidants: an electrochemical overview. BioMed Research International 2013: 1-11. https:// doi.org/10.1155/2013/251754 Zeng, Z., Li, Y., Yang, R., Liu, C., Hu, X., Luo, S., Gong, E. and Ye, J., 2017. The relationship between reducing sugars and pheno- lic retention of brown rice after enzymatic extrusion. Journal of Cereal Science 74: 244–249. https://doi.org/10.1016/j. jcs.2017.02.016 Zhang, H., Yu, D., Sun, J., Liu, X., Jiang, L., Guo, H. and Ren, F., 2014. Interaction of plant phenols with food macronutrients: characterisation and nutritional-physiological consequences. Nutrition Research Reviews 27: 1–15. https://doi.org/10.1017/ S095442241300019X Zlatić, E., Pichler, A. and Kopjar, M., 2017. Disaccharides: influence on volatiles and phenolics of sour cherry juice. Molecules 22 (11): 1-10. https://doi.org/10.3390/molecules22111939 Liu, S.X., Yang, H.Y., Li, S.Y., Zhang, J.Y., Li, T., Zhu, B.Q., Zhang,  B.L., 2015. Polyphenolic compositions and chromatic characteristics of bog bilberry syrup wines. Molecules 20: 19865–19877. https://doi.org/10.3390/molecules201119662 Loncaric, A., Dugalic, K., Mihaljevic, I., Jakobek, L. and Pilizota, V., 2014. Effects of sugar addition on total polyphenol content and antioxidant activity of frozen and freeze-dried apple Purée. Journal of Agricultural and Food Chemistry 62: 1674–1682. https://doi.org/10.1021/jf405003u Madhavamenon, K.I. and Maliakel, B.P., 2015. Instant water sol- uble bioactive dietary phytonutrients composition of spice/ herb extracts and a process for its preparation. United States Application Publication. US 2015/0352173 A1. 1–8. Mansoor, K., Qadan, F., Schmidt, M., Mallah, E., Abudayyih, W. and Matalka, K., 2017. Stability study and a 14-day oral dose toxic- ity in rats of plantain leaf extract (Plantago lanceolata L.) syrup. Scientia Pharmaceutica. 85 (1): 1-5. https://doi.org/10.3390/ scipharm85010015 Marjanović, A., Đeđibegović, J., Brčaninović, M., Omeragić, E., Čaklovica, F., Dobrača, A. and Šober, M., 2015. Antioxidant potential of selected traditional plant-based beverages in Bosnia and Herzegovina. https://www.academia.edu/36703896/ Antioxidant_potential_of_selected_traditional_plant-based_ beverages_in_Bosnia_and_Herzegovina Martin, K.R., 2009. Polyphenols as dietary supplements: a dou- ble-edged sword. Nutrition and Dietary Supplements 2: 1–12. https://doi.org/10.2147/NDS.S6422 Musa, K.H., Abdullah, A., Jusoh, K. and Subramaniam, V., 2011. Antioxidant activity of pink-flesh guava (Psidium guajava L.): effect of extraction techniques and solvents. Food Analytical Methods 4: 100–107. https://doi.org/10.1007/s12161-010-9139-3 Payne, A.C., Mazzer, A., Clarkson, G.J.J. and Taylor, G., 2013. Antioxidant assays – consistent findings from FRAP and ORAC reveal a negative impact of organic cultivation on antioxidant poten- tial in spinach but not watercress or rocket leaves. Food Science & Nutrition 1: 439–444. http://doi.wiley.com/10.1002/fsn3.71 Peinado, J., de Lerma, N.L. and Peinado, R.A., 2010. Synergistic anti- oxidant interaction between sugars and phenolics from a sweet wine. European Food Research and Technology 231: 363–370. https://doi.org/10.1007/s00217-010-1279-6 Pengseng, N., Siripongvutikorn, S., Usawakesmanee, W. and Wattanachant, S., 2011. Combined effect of carbohydrate and ther- mal processing on antioxidant activity of galangal coconut-milk paste extract, Tom-Kha. International Food Research Journal 18: 907–914. Pereira, W., Silva, C.M.D.P.S., Lígia, K., De Almeida, C. and Aires, F., 2014. Description of guava osmotic dehydration using a three-dimensional analytical diffusion model. Journal of Food Processing 2014: 1-7. https://doi.org/10.1155/2014/157427 Pires, T.C.S.P., Dias, M.I., Calhelha, R.C., Carvalho, A.M., Queiroz, M.J.R.P., Barros, L., Ferreira, I.C.F.R. and McPhee, D.J., 2015. Bioactive properties of tabebuia impetiginosa-based phyto- preparations and phytoformulations: a comparison between extracts and dietary supplements. Molecules 20: 22863–22871. https://doi.org/10.3390/molecules201219885 Puoci, F., Iemma, F., Spizzirri, U.G., Restuccia, D., Pezzi, V., Sirianni, R., Manganaro, L., Curcio, M., Parisi, O.I., Cirillo, G. and Picci, https://doi.org/10.3390/nu3030317� https://doi.org/10.3390/nu3030317� https://doi.org/10.1155/2014/847013� https://doi.org/10.1155/2014/847013� http://www.ncbi.nlm.nih. gov/pubmed/24822061%5Cn http://www.ncbi.nlm.nih. gov/pubmed/24822061%5Cn https://doi.org/10.3390/nu8110697� https://doi.org/10.3390/nu8110697� http://www.agriculturejournals.cz/publicFiles/105870.pdf� http://www.agriculturejournals.cz/publicFiles/105870.pdf� http://linkinghub.elsevier.com/retrieve/pii/S0924224405002049� http://linkinghub.elsevier.com/retrieve/pii/S0924224405002049� https://www.sciencedirect.com/science/article/pii/S0076687999990171� https://www.sciencedirect.com/science/article/pii/S0076687999990171� https://doi.org/10.17306/J.AFS.2017.0491� https://doi.org/10.5219/480� https://doi.org/10.5219/480� https://doi.org/10.1155/2013/251754� https://doi.org/10.1155/2013/251754� https://doi.org/10.1016/j.jcs.2017.02.016� https://doi.org/10.1016/j.jcs.2017.02.016� https://doi.org/10.1017/S095442241300019X� https://doi.org/10.1017/S095442241300019X� https://doi.org/10.3390/molecules22111939� https://doi.org/10.3390/molecules201119662� https://doi.org/10.1021/jf405003u� https://doi.org/10.3390/scipharm85010015� https://doi.org/10.3390/scipharm85010015� https://www.academia.edu/36703896/Antioxidant_potential_of_selected_traditional_plant-based_beverages_in_Bosnia_and_Herzegovina https://www.academia.edu/36703896/Antioxidant_potential_of_selected_traditional_plant-based_beverages_in_Bosnia_and_Herzegovina https://www.academia.edu/36703896/Antioxidant_potential_of_selected_traditional_plant-based_beverages_in_Bosnia_and_Herzegovina https://doi.org/10.2147/NDS.S6422� https://doi.org/10.1007/s12161-010-9139-3� http://doi.wiley.com/10.1002/fsn3.71� https://doi.org/10.1007/s00217-010-1279-6� https://doi.org/10.1155/2014/157427� https://doi.org/10.3390/molecules201219885� _GoBack