Physicochemical and dynamic sensory characterization of a Yerba mate/Blackcurrant instant beverage powder rich in natural antioxidants

Abstract

BACKGROUND:

Yerba-mate (YM) and blackcurrant (BC) are rich in polyphenols and anthocyanins with proven health benefits therefore they could be used in functional beverage preparation.

OBJECTIVE:

To (a) develop a palatable powdered beverage with high physicochemical and nutritional quality using YM/BC, (b) determine the effect of in-vitro gastrointestinal digestion on antioxidant content and activity and (c) evaluate beverage’s acceptance temporal changes and dynamic profile during multiple intakes.

METHODS:

We determined powder’s water-activity (aw), moisture content, flowability, micro-morphology, color, cold water solubility, glass-transition temperature (Tg), total polyphenols (TP)/anthocyanins (MAC) and ascorbic-acid contents, antiradical activity (DPPH).

RESULTS:

Powder’s aw (0.089) ensured biochemical/microbiological stability. Tg(55.64°C) indicated that YM/BC’s vacuum-storage at 20–25 °C was possible without losing glassy state. Powder’s cold water’s solubility was 90%. 1 L (60 g/L) provided 556.8 mg TP/198.11 mg monomeric-anthocyanins. Ascorbic- acid dose was half the recommended daily intake. In-vitro gastrointestinal digestion reduced total polyphenols/monomeric-anthocyanins 59–70 %, but antiradical activity losses were 9%. Overall-acceptance and Time-Intensity curves results were in the Hedonic scale’s “Like moderately to like very much” zone. Berries/Sweet were the only dominant attributes Astringency accumulative effects not detected.

CONCLUSIONS:

The YM/BC powder had good physical/biochemical/microbiological stability under industrial storage conditions and high antiradical activity after simulated gastrointestinal digestion. Sensory scores suggested that consumer’s acceptance could be high.

Keywords

Yerba mate blackcurrant antioxidant activity dynamic sensory methods

1 Introduction

The current dietary concerns of both the aging population and those with fast-paced lifestyles have moved from foods that prevent nutritional deficiency and their associated diseases to those offering long-term prevention of chronic ailments. The growing awareness of the link between diet and health has resulted in new views and perceptions about the effect of dietary compounds [1] that steadily enhanced functional foods consumption.

Yerba mate (Ilex paraguariensis; YM), has a high content of polyphenols and flavonoids with antioxidant and hepato-protective properties and the capacity to improve the cardiovascular and central nervous systems [1, 2].

Blackcurrant (Ribes nigrum; BC) is an excellent source of ascorbic acid, anthocyanins, flavonols, procyanidins, and phenolic acids [3] with a wide range of health benefits including antioxidant, antimicrobial, anti-carcinogenic and neuroprotective activities [4].

Marriott [5] reported that functional food’s palatability, format, convenience, and familiarity are key factors in consumers’ acceptance. Consumption of yerba mate infusions alone or combined with juices is very popular in Argentina, Brazil, and Uruguay, therefore, producing a functional beverage combining it with blackcurrant juice may be a simple and effective mean for providing their health benefits to a high number of consumers.

Although phenolics’ content and composition in foods can influence the effectiveness of their health benefits, the digestion effect is more relevant [1] In vitro digestion models are useful tools for simulating the different stages occurring in the human digestive tract. These techniques are a good alternative to in vivo studies because they are faster, less expensive and free of ethical issues [6].

Recent studies by Correa et al. [7] using different yerba mate infusions popular in Brazil, Uruguay and Argentina indicated that although in vitro gastrointestinal digestion decreased their phytochemicals content, YM’s antioxidant, antibacterial and antitumor activities were statistically significant. Olejnik et al. [8] demonstrated that digested BC extracts reduced reactive oxygen species generation and oxidative DNA damage. On the other hand, in a complex mix containing polyphenols and other antioxidants from different plants, the bioactives may act additively and/or synergistically, therefore, a combination of YM and BC may improve the health benefits detected in the original ingredients.

The process of eating or drinking implies dynamic phenomena that take place over a time lapse ranging from a few seconds to several minutes. During this period, chewing, interaction with saliva and oral mucosa and heat exchanges result in modifications in the product’s structure and texture as well as in its patterns of flavor release. Some stimuli may induce very short sensory signals while others, like bitter substances, can cause persisting sensations capable of conditioning the way the product is perceived [9]. There are several methodologies for measuring consumer’s hedonic response to food during a one bite/sip consumption event from a dynamic perspective. Methven et al. [10] used Hedonic Time-Intensity (TI) and Multi-Attribute Time-Intensity methods to provide information about the onset and decay of the hedonic attributes, their maximal intensity and duration. However, Rocha-Parra et al. [11] concluded that analyzing the acceptance’s dynamic changes over several sips provided a more realistic hedonic experience.

There is an increasing interest in determining the evolution of sensations during the sensory analysis of food products. Pineau et al. [12] concluded that Temporal Dominance of Sensation (TDS) methodology was appropriate for obtaining the temporal information, since it allowed the simultaneous evaluation of several attributes and their dominance sequence during a certain time period. In these tests, participants received a complete list of attributes and asked to continuously identify and score the different dominant sensations appearing and disappearing over a fixed period. This method was successful for analyzing study dairy products [12].

In this study we aimed to: a) examine the feasibility of using yerba mate infusions and black currant pulp to prepare a functional instant beverage powder combining good palatability with high physicochemical and nutritional quality. b) determine the effect of in vitro gastro intestinal digestion on their antioxidant content and antiradical scavenging capacity and c) evaluate the temporal changes in YM/BC’s acceptance through multiple-sip temporal-liking and to measure its dynamic profile over multiple intakes.

2 Materials and methods

2.1 Preparation of the blackcurrant pulp and the yerba mate infusion

The organic ripe blackcurrant berries (Ribes nigrum cv. Silvergieter; Chacras Cuyen, El Bolson, Chubut, Argentina) were harvested during January 2012 and stored at – 20°C for 270 days. 24 h before the beverage preparation, the fruit was defrosted and processed in an industrial fruit pulper (Filter net pore diameter: 2 mm) to obtain the pulp (BC; 40 °Brix; pH = 3.21). To prepare the yerba mate infusion (YMI), 60 g L^–1 of commercial yerba mate leaves (Ilex paraguariensis St Hil; La Unión Suave, Est. Las Marias SAIC, Gov. Virasoro, Argentina) were heated at 100°C for 15 min, decanted (15 min, 25°C) and filtered.

2.2 Freeze-drying process

The BC and YMI, in a 3:1 ratio, were mixed with 15% w/v Maltodextrin Dextrose Equivalent 10 (MD; Productos de Maíz S.A., Buenos Aires, Argentina) and freeze dried at room temperature with a FIC L1-1-E300-CRT freeze dryer (Buenos Aires, Argentina). After freeze-drying, the powder was homogenized with sugar (4.80 %; Ledesma, Jujuy, Argentina), a commercial diet sweetener (0.10 %; Cyclamate 5700 mg 100 g^-¹; Saccharin 2000 mg/Dextrose) and a commercial “red berries” aroma (0.10 %; Symrise AG, Tortuguitas, Argentina). The powder’s formulation (% w/w) was: YM/BC/MD (53.3 %), sugar (44.9 %); commercial sweetener (0.9 %), red berries Aroma (0.9 %) while its concentration in the beverage was 60 g L^-1.

2.3 Characterization of the freeze-dried YM/BC instant beverage powder

2.3.1 Flowability tests

The freeze-dried YM/BC’s flowability was determined by measuring the dynamic angle of repose with the rotating cylindrical chamber from the Solids Handling Study Bench (CEN, Armfield, United Kingdom), the results were the average of 10 repeats. The control was a mix of commercial sugar and MD₁₀ (5 %). To determine the tapped bulk density, a known mass of the powder (40 g) was delivered by gravity into a measuring cylinder and tapped 100 times at roughly 60 taps min^-¹ [13].

The density was calculated in triplicate with equation (1), ρ is the tapped density, P_i is the weight of the sample (g) and V the average volume of the powder (mL). $ρ_{volumetric} = \frac{P_{i}}{V}$ (1)

2.3.2 Water activity and moisture content

The water activity (a_w) was determined at 25°C in an AquaLab series 3 (Decagon Device, Pullman, Washington, USA). The moisture content was analyzed gravimetrically in triplicate samples with the AOAC method [14].

2.3.3 Color analysis

Powder’s colour was measured on triplicate samples with a Minolta CR-400 Chroma Meter (Minolta, Osaka, Japan), each value was the average of 9 measurements. Colour was expressed by its lightness (L*), redness (a*) yellowness (b*), saturation index (SI) and hue angle (HA; [15]).

An increase in HA towards more positive values or a reduction to more negative indicated an enhancement in yellowness or blueness respectively.

2.3.4 Cold water solubility

The YM/BC freeze-dried powder's solubility (Equation 2) was determined in duplicate according to [16].

$% Solubility = \frac{(p_{ss} * DF * 100)}{p_{l}}$ (2)

pss and pl = dried weights (g) of the supernatant and the sample respectively; FD = dilution factor.

2.3.5 Glass transition temperature

The glass transition temperature (T_g) was determined with a differential scanning calorimeter (DSC, Q-100 TA 5000; TA Instruments; USA), The samples (2–4 mg) hermetically sealed in aluminum pans were heated at 5°C min^-¹ from –50 to 120°C using an empty pan as a reference. The onset (T_gonset), mid (T_gI) and end (T_gend) temperatures of glass transition, were determined with the TA Universal Analysis 2000 software (TA Instruments Waters LLC DE USA). All the measurements were done in duplicate.

2.3.6 Freeze-dried YM/BC microstructural analysis

The powders microstructure was analyzed by scanning electron microscopy (SEM) under high vacuum conditions with a FEI, Quanta 200 microscope (Netherlands). The samples attached to stubs using two-sided adhesive tape and coated with a layer of gold (40–50 nm) were examined with an acceleration voltage of 20 kV.

2.4 Chemical analysis

2.4.1 Antioxidant concentrations of the YM/BC powder

The total phenolic concentration (TP; mg GAE (gallic acid equivalents) (g d.b. (dry weigh basis)) ^–1 was assessed with the Folin-Ciocalteau method [17] with an UVmini-1240 UV-Vis Spectrophotometer (Shimadzu Scientific Instruments, Japan).

The monomeric anthocyanin (MAC) content of the YM/BC powder was determined with the pH differential method reported by Giusti and Wrolstad [18]. The monomeric anthocyanins were extracted with ethanol:HCl 0.1 N (85:15). After diluting the extracts with buffer to achieve an appropriate concentration range, the extract’s absorbances were measured at 520 (λ_max) and 700 nm with a spectrophotometer U-1900 (HITACHI, Japan). MAC content, calculated with equations (3) and (4), was expressed as mg CyGE (cyanidin-3-glucoside equivalents) (g d.b.) ^–1. $A = (A λ_{vis - \max} - A λ_{700 nm})_{pH 1.0} - (A λ_{vis - \max} - A λ_{700 nm})_{pH 4.5}$ (3) $MAC (mg L^{- 1}) = \frac{A * MW * DF * 1000}{\in * 1}$ (4)

MAC molecular weight (MW) = 449.2 g mol^–1; extinction coefficient (ɛ) = 26900 L cm^–1 mol^–1; DF = dilution factor [18].

To analyze the ascorbic acid (AA) content, the sample (0.5 g) was extracted with an aqueous solution 50 g L^–1 of metaphosphoric acid (Carlo Erba S.A, BCN, Spain) followed by centrifugation at 2000 rpm (Rolco CM 2036, Buenos Aires, Argentina). AA concentration was determined by high-performance liquid chromatography (Waters, model R-414, Milford, MA, USA). The method consisted of an isocratic elution procedure with UV-Visible detection at 245 nm using AA (Food grade, Parafarm) as external standard. Before injection, the extracts were filtered with a pre-filter and a 0.45μm millipore membrane. Separations were done on a 5 mm RP C18 column of 150 mm –4.6 mm (Symmetry, Waters, Dublin, Ireland) at 25°C. The mobile phase was a mixture of 5 g L^–1 metaphosphoric acid:acetonitrile (93:7) with a flow rate = 1 mL min^–1 [15, 19]. To prevent the loss of AA, standard solutions and extracted samples were protected from light. Quantitation was performed according to Orjuela-Palacio et al. [14].

2.4.2 Antioxidant activity

The antiradical activity (% I; Equation 5) of the reconstituted YM/BC beverage was analyzed with the DPPH• scavenging assay [20] as described by Valerga et al. [21]. $I % = [A_{b}^{0} - A_{s}^{t}) / A_{b}^{0}] \times 100$ (5)

A_b⁰ and A_s^t are the absorbances of the blank (b) and the sample (s) at time = 0 and at t = 120 min.

2.5 Simulated in vitro gastrointestinal digestion

The effect of gastrointestinal digestion on the bioactives’ concentrations and the antioxidant activity of the YM/BC powdered instant beverage was assessed as described by Chiang et al. [22]. The method comprised 2 sequential steps: a gastric digestion (pepsin/HCl pH = 2.5) followed by intestinal digestion (pancreatin/bile salts pH = 8) with active (G_act/GI_act) or heat-inactivated (G_inac/GI_inac) enzymes. In the latter, the enzymatic solutions were heated at 90°C for 5 min [22]. Before digestion, the freeze-dried powder was dissolved in milli-Q water (60 g L^–1) and homogenized for 5 min under constant agitation.

To determine the TP, MAC and the antiradical (DPPH•) capacity, fractions of the gastric and gastrointestinal stages were collected and centrifuged at 13000 rpm (Rolco CM 2036, Buenos Aires, Argentina) for 10 min at 25°C, the supernatant was filtered with a nylon membrane (0.45μm diameter pore) and kept at –80°C until analysis.

2.6 Dynamic sensory characterization

2.6.1 Consumers’ acceptance: Overall acceptance and time-intensity to liking

Seventy five consumers [33 males / 42 females; age = 18–65 years (M = 32 years SD = 10.6)], selected between the students and staff from the Faculty of Physical Sciences of the La Plata National University (UNLP; La Plata, Argentina), took part voluntarily in the experiment. The sensory evaluation was done in three stages.

After an initial instruction regarding the software use, the participants assessed the beverage's overall acceptance degree with a 9-point category hedonic scale anchored at: dislike very much (1), like very much (9), and a neutral point at 5 (neither like nor dislike).

In the second phase, the consumer panel evaluated the acceptance level's variation over time with the Time-Intensity (T-I) data acquisition module (SensoMaker v1.8; Federal University of Lavras, Brazil) as described by Rocha-Parra et al., [11] modifying the number of sips. In the current study the participants took 4 sips, one sip every 20 seconds (s) (0 (initial), 20, 40 and 60 s) and the total rating time was 80 s.

In the third step and after the dynamic evaluation, the overall acceptance degree was measured again following the same procedure described in (a).

2.6.2 Dynamic profile of the YM/BC beverage determined by the Temporal Dominance of Sensations (TDS) methodology

A panel of 20 participants, experienced in sensory descriptive evaluation, was trained during eight sessions to generate the attributes that described the main characteristics of the sample: Sweet, Acid, Bitter, Astringent aftertaste, Total aroma, Berries aroma (Retronasal (R)), Blackcurrant aroma (R) and Yerba mate (R)).

The participants were instructed to take 3 successive sips (15 mL each, 10°C) every 20 seconds (s) (0 (initial), 20 and 40 s) and to rate them during 60 s as described in Sec. 2.6.1. Immediately after sipping, the panelist had to identify and select the dominant attribute. TDS analysis was done according to the procedure described by Pineau et al. [12] in duplicate on different days using the SensoMaker v1.8 software.

2.7 Statistical analysis

All results of the physicochemical and sensory analysis were reported as mean±standard deviation (SD). Treatment effects were determined by analysis of variance followed by pairwise comparisons with the Tuckey test, statistical significance was set at p value < 0.05 (Infostat (v. 2013) Universidad Nacional de Cordoba, Argentina).

TDS curves show graphically the time dependence of the proportion of assessors identifying as dominant a specific attribute. The dominance rate (%) for each attribute at a given time was determined as the fraction of judgments (assessors*replicates) for which the given attribute was selected as dominant and is directly related with the agreement’s degree across the panelists.

The chance (P_o) level is the dominance rate that an attribute can obtain by chance [11] and was calculated with equation (6) where p is the number of attributes.

$P_{o} = \frac{1}{p}$ (6)

The significance level represents the minimum value this proportion should equal to be considered significantly greater than P_o and was calculated with the confidence interval of a binomial proportion based on a normal approximation [12]; (Equation 7).

$P_{S} = P_{o} + 1.645 \times \sqrt[2]{[P_{o} \times \frac{(1 - P_{o})}{n}]}$ (7)

Where, P_s is the lowest significant proportion value (α= 0.05) at any point in time for a TDS curve, n is the number of subject replication [12]. Preliminary TDS training sessions showed that 4 sips produced a fatigue effect in the participants therefore the number of sips was reduced to 3.

3 Results and discussion

3.1 Characterization of YM/BC freeze-dry powde

The powder’s humidity content (3.11±0.03 % wet weight basis (w.b.)) was 50 % lower than those reported by Yatsu et al. [23] and Franceschini et al. [24] for spray dried YM infusions or freeze-dried blackberry pulp.

YM/BC’s solubility in cold water was 89.93±1.20%, its color attributes were: 53.69±2.36 (L*), 21.83±0.75 (a*) and 0.98±0.05 (b*) whereas its hue angle (HA) and saturation index (SI) were 0.12±10^–3 and 21.85±2.36 respectively.

The powder’s glass transition temperatures (°C) were 55.64±13.9 (T_g (onset)), 60.59±13.54 (T_g (I)) and 66.45±14.31 (T_g (final). YM/BC T_g (I) value’s was similar to that reported by Ferrari et al., [25] for spray dried blackberry with 7 % MD. The high T_g (onset) values indicated that the YM/BC powders could be stored at 20–25 °C in air tight containers without losing its glassy state.

The water activity value (0.089) was within the range (a_w <0.2) recommended by Labuza and Altunakar [26] for preserving biochemical stability and microbiological safety. Although the YM/BC humidity levels (3.11 % w.b) were similar to those reported by Ferrari et al. [25] (3.25 %) and Franceschinis et al. [24] (3.7 %) for spray dried blackberry pulp with no sugar, its water activity was 191 % and 54 % lower respectively due in part to sucrose’s a_w reducing capacity [26].

UPS pharmacopeia convention (2012) reported that powders with repose angles >50° have flow problems during processing. Although the YM/BC’s repose angle (52.75°±2.19) was 5.5 % higher than that limit, the difference was not significant (P > 0.05) therefore their flow capacity was still acceptable for manufacturing purposes.

Several studies [27, 28] demonstrated that particle size, shape and surface characteristics affected powder’s fluidity. Compared with smooth round particles, those with smaller size, irregular shapes and rough surfaces have more points of contact between them and consequently a higher cohesiveness and a lower flowing capacity [29, 30]. Since YM/BC particles microstructure fulfilled this condition, their flow behavior could be partially explained by their morphology.

The powder’s microstructure was consistent with its low humidity (Fig. 1A) since the particles’ shape and size were highly irregular with sharp edges and porous wrinkled surfaces (broken glass) suggesting the absence of agglomeration. The sucrose crystal’s microstructure (Fig. 1B) corroborated these observations as it also presented well-defined edges and some freezedried particles on its surface indicative of the YM/BC sample’s low humidity. Since the sucrose was incorporated to the YM/BC/MD freeze-dried powder, it is expected that at the sample’s humidity content, the sucrose crystals would appear separated from the YM/BC/MD matrix as indicated by Fig. 1B.

Fig.1

Scanning electron micrographs of the freeze-dried YM/BC powder’s surface (A, magnification = 1.600 x); Sucrose crystal (B, magnification = 200x).

The reconstituted drink solution was stable at 10°C since only 10 % sedimented after centrifugation. The TP, MAC and AA concentrations of the YM/BC before freeze-drying were:

TP(YM/BC) _bfd = 228.63±6.21 mg GAE g^–1 d.b.

MAC(YM/BC) _bfd = 583.15±1.40 mg CyGE 100 g^–1 d.b

AA(YM/BC) _bfd = 175.16±7.35 mg AA 100 g^–1 d.b.

Comparison of the YM/BC bioactives levels with those from the YMI/BC before freeze-drying [(YM/BC) _bfd] indicated that this process caused a 43% and 58% reduction in the MAC and AA contents respectively whereas for TP, the loss was 96%.

Although citrus are the most popular ascorbic acid sources, there are other fruits like strawberries with a much higher content and also have the advantage of being naturally rich in polyphenols and anthocyanins with excellent antioxidant properties [31]. Based on the results reported by Da Silva et al., [32] 1 L (60 g) of the YM/BC beverage provided a TP amount similar to 163 g of strawberries or 426 g of oranges while the AA level was equivalent to those present in 100 g of strawberries or 233 g of fresh oranges. The MAC intake was equal to that provided by 32–100 g strawberries.

3.2 Influence of the in vitro gastrointestinal digestion on the Bioactives concentrations and antioxidant activity of YM/BC

Figure 2 A and B show the effect of the gastric and gastrointestinal digestion with active (G_act/GI_act) and heat inactivated enzymes (G_inac/GI_inac) on the total polyphenols (Fig. 2A) and monomeric anthocyanins (Fig. 2B) concentrations.

Fig.2

Effect of the gastric and gastrointestinal digestion with active (G_act/GI_act) and heat inactivated enzymes (G_inac/GI_inac) on the Total Polyphenols (A; TP); Monomeric Anthocyanins Concentrations (B; MAC) and YM/BC antiradical capacity (C; % I). Control = undigested YM/BC. Bars with different letters (a-c) are statistically different with Tukey test (α= 0.05).

Gastrointestinal digestion caused a 59–70 % loss (P < 0.05) in TP, higher than those reported by Correa et al. [7] for YMI digested with active enzymes (20–33 %). Olejnik et al., [8] indicated that in vitro GI digestion of blackcurrant reduced the level of hydroxycinnamic acids, flavanols and flavonols between 47–19.8%.

The ranking was: C > G_act ≈ G_Iact ≈ GI_inac > G_inac.

Comparison between G_act and G_inac indicated that the pepsin activity increased TP’s stability by 26 % possibly through the partial formation of products more resistant to the existing experimental conditions. Statistical analysis of the G_act and GI_act data showed that TP was not affected (P > 0.05) by the intestinal digestion, however, when samples were treated with inactive enzymes the GI_inac treatment enhanced TP by 28 % from 2.54 to 3.25 mg GAE g^–¹ d.b. This could be in part due to the neutral pH used to simulate intestinal conditions capacity to reduce MAC’s stability, favoring the formation of more unstable compounds like hemiketals, chalcones and quinonoids that may break down to produce phenolic compounds positive to the Folin-Ciocalteu reaction [1]. Hydroxybenzoic acids, presumably derived from the B ring of the aglycones have been identified after incubation of anthocyanins at neutral pH and room temperature [33]. Tsuda et al. [34] reported the presence of protocatechuic acid in the serum of rats fed cyanidin 3-O-glucoside.

Gastrointestinal digestion diminished the monomeric anthocyanins concentrations 92–95 % (Fig. 2B), these losses were greater than those reported by Olejnik et al. [8] (54.5–59.6 %) for blackcurrant extracts. In contrast with the TP results (Fig. 2A), active enzymes reduced MAC’s stability 15 % and 33 % respectively. Although alkaline pH is detrimental to anthocyanins stability [1], MAC’s content in the G_inac and GI_inac were statistically similar (P > 0.05).

The order of decreasing stability was: C > GI_inac ≈ G_inac > G_act ≈ GI_act.

McDougall et al. [35] reported that wine anthocyanins were stable in simulated gastric conditions but were degraded after the GI step forming breakdown products which may be bioavailable and bioactive by themselves. Studies done with Ribes magellanicum and R. punctatum [36] indicated that the polyphenols loss was significant in both gastric and intestinal steps.

Figure 2 C shows the effect of the simulated in vitro gastrointestinal digestion on the YM/BC antiradical capacity measured with the DPPH• test, the control C corresponded to the undigested sample. In spite of the large drop detected in total polyphenols and MAC levels, the loss of YM/BC antiradical capacity was much lower. Gastric digestion with active enzymes did not affect (P > 0.05) YM/BC's activity however, an 8 % loss (P < 0.05) was detected after the GI_act treatment, pancreatin’s activity diminished AOA from 85 to 80 % (P < 0.05).

Correa et al., [7] reported that gastrointestinal digestion reduced the antioxidant capacity of yerba mate infusions determined with the DPPH• and ORAC assays 28 % and 22 % respectively. Olejnik et al. [8] showed that the BC extracts, followed a similar trend, with a 42 % loss in ORAC antiradical capacity. Nevertheless, the digested products retained high levels of their original functional properties including antioxidant, antibacterial and antitumor activities as well as the capacity to prevent reactive oxygen species formation and oxidative DNA damage induced in intestinal cells [7, 8]. The difference detected in the antiradical activity between the digested YM/BC and its ingredients could be attributed to a positive interaction between the YM and the BC polyphenols that may in part compensate the negative effects of the GI digestion process. Cilla et al., [37] found that although gastrointestinal digestion reduced the polyphenol (36 %) and ascorbic acid (16 %) levels of a beverage prepared with a mix of apricot, grape and orange juices, its TEAC and ORAC values increased 20 and 59 % respectively.

3.3 Dynamic sensory characterization

3.3.1 Consumers’ acceptance

Statistical analysis indicated that the variation between initial and final evaluation of the YM/BC beverage’s overall acceptance after 80 s was not significant (P > 0.05), the values were 7.04±1.21 (initial) and 7.33±1.04 (80 s) (Fig. 3).

The T-I curves (Fig. 3) showed that during sip 1 the acceptance increased up to a value of 6.7, further intakes (sips 2–4) confirmed this behavior since after sip 4, the overall acceptance was 18.3 % higher (P < 0.05).

Fig.3

Initial and final overall acceptance determined by hedonic scale (▾) and time-intensity curves for the acceptance level of the YM/BC beverage during 80 s (4 sips).

Table 1 shows t_Imax (time to reach the I_max), I_max and AUC values for the four sips split by gender. The sip effect was highly significant (P < 0.05), I_max increased 20–23 % and AUC 75–77 %, however, the gender influence in both parameters was not significant (P > 0.05).

Table 1

Time to reach maximum intensity (t_Imax), Maximum intensity (I_max) and Area under the curve (AUC) of the T-I curves for the YM/BC’s acceptance across the four sips splitted by Gender (F = Female; M = Male)

Gender	Sip	t_Imax	I_max	AUC
F	1	7.84±3.85^a	6.77±1.54^a	91.11±27.44^a
	2	25.41±6.14^b	7.34±1.26^ab	133.05±25.00^b
	3	45.61±6.50^c	7.70±1.36^bc	140.75±25.72^b
	4	62.59±4.74^d	8.09±1.12^c	158.16±23.81^c
M	1	7.33±2.93^a	6.33±1.43^a	86.06±24.13^a
	2	23.48±5.17^b	6.79±1.29^ab	124.12±27.66^b
	3	45.00±9.37^c	7.30±1.21^bc	132.03±23.52^b
	4	64.24±5.69^d	7.73±1.33^c	150.91±23.25^c

Values within a column with different superscripts (^a - d) are statistically different by Tukey test (α= 0.05).

The results from both tests (overall acceptance, I_max and AUC) clearly correspond to the “like” zone of the hedonic scale (scores >5), indicating that the YM/BC beverage had a high probability of being accepted by consumers.

3.3.2 Temporal dominance of sensations (TDS)

Table 2 shows the TDS parameters calculated for an evaluation period of 60 s. DUR_max represents the maximum dominance rate for each attribute, t_max is the time (s) for DUR_max’s formation and t_max90 % the time interval corresponding to a dominance rate equal or greater than 90% of DUR_max. Berries (R) had the highest dominance rate and appeared 2nd after Sweet (27 s) while the Astringent, Acid and Bitter attributes achieved DUR_max practically at the end of the assay (t_max ≥58 s). Yerba mate’s DUR_max was the lowest and Blackcurrant was not detected.

Table 2
Temporal Dominance of Sensation parameters for YM/BC’s attributes

Attributes DUR_max t_max t_max90 %

Sweet 0.37 9.00 25.20

Acid 0.22 60.00 24.00

Bitter 0.15 60.00 0.00

Astringent 0.25 58.80 3.00

Total aroma (R) 0.17 18.60 36.00

Berries (R) 0.61 27.00 8.00

Blackcurrant (R) 0.00 0.00 0.00

Yerba mate (R) 0.11 28.20 27.60

Attributes	DUR_max	t_max	t_max90 %
Sweet	0.37	9.00	25.20
Acid	0.22	60.00	24.00
Bitter	0.15	60.00	0.00
Astringent	0.25	58.80	3.00
Total aroma (R)	0.17	18.60	36.00
Berries (R)	0.61	27.00	8.00
Blackcurrant (R)	0.00	0.00	0.00
Yerba mate (R)	0.11	28.20	27.60

DUR_max (maximum rate of the dominance for each attribute evaluated), t_max (time to the duration of DUR_max), t_max 90% (interval at which the rate of dominance is greater than 90% of DUR_max).

Figure 4 shows the mean TDS curves for the different sips, each curve represents the consensus variation across panelists for a certain dominant attribute [12]; the broken lines represent the chance (P_o = 0.125) and significance levels (P_s = 0.199) for the 8 attributes respectively.

Fig.4

Temporal Dominance of Sensations of YM/BC’s flavor attributes during 60 s (3 sips).

Only Berries (R) and Sweet were dominant across the three sips, the time periods (s) where these attributes were higher than P_s were: 80 s for the former and 5–16, 30–42 and 47–56 s for the latter. Astringent was dominant in the periods of 42–49 s and after 55 s (Fig. 4), when the Berry (R) aroma and Sweet decreased. Therefore, there was no evidence of the accumulative effect through the intake. The dominance of the attributes directly related to the main components of the Yerba mate were not significant.

Due to the complex nature of the sensory interactions between blackcurrant and yerba mate, it is difficult to determine the components ratio that maximizes the beverage’s sensory appeal. It is evident that the ingredients were mixed in optimum proportions, because it was possible to mask the flavor of the Yerba mate decreasing the negative effects of its astringency to levels where consumers’ acceptance was not affected.

In a previous study [38] using a beverage based on the same ingredients, we found that sweetness had a high influence on the perception of the other attributes and contributed to the balance among them. In the present paper, besides the sweet taste, the berry aroma also helped towards the formulation’s improvement.

4 Conclusions

The YM/BC water activity values were within the range recommended for preserving biochemical stability and microbiological safety. The high T_g (onset) values (55.64°C) indicated that the YM/BC powders could be stored at 20–25 °C in airtight containers without losing its glassy state. The powder’s solubility was high (90 %) and stable at 10°C since only 10 % sedimented after centrifugation.

Although freeze-drying reduced TP, MAC and AA contents 43–96 %, the beverage was a good source of highly active antioxidants. 1 L (60 g L^–¹) provided a TP amount similar to 163 g of strawberries or 426 g oranges while the MAC intake was equivalent to that supplied by 32–100 g strawberries. In addition, the ascorbic acid dose corresponded to 50 % of the daily intake recommended by FAO/WHO [39].

Although in vitro gastrointestinal digestion caused 59 to 70% reduction in total polyphenols and monomeric anthocyanins, the drop in antiradical activity of the YM/BC beverages was only 7%.

Overall acceptance and TI curves parameters (I_max and AUC) were in the range of the Like moderately to like very much zone. The TDS methodology showed that Berries and Sweet were the only dominant attributes and that Astringency accumulative effects were not detected. The combination of two dynamic methods, one to evaluate the acceptance and the other to determine the dominant sensations along a repeated intake was very effective in explaining consumers’ liking for the new beverage. Given the unusual ingredients combination, the high acceptance scores obtained by the YM/BC suggested that the product has a high probability of being accepted by the consumers.

Funding

The authors report no funding.

Conflict of interest

The authors declare no conflict of interest.

Footnotes

Acknowledgments

Generous financial support was provided by the Argentine National Institute of Yerba Mate (Instituto Nacional de la Yerba Mate, INYM). We thank Eng Chris Young and Dr R. Shorthose for assisting with the English translation and Claudio Reyes for the ascorbic acid analysis. J. M. Orjuela-Palacio was supported by the Argentinean Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET).

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