Physicochemical characterization of Galician Honeys

Abstract

The characterization of the north-west Spanish honeys was carried out on the basis of different parameters: density, viscosity, refractive index, glass transition temperature, pH, electrical conductivity, ash, sugars, and water activity. The influence of certain characteristics and honey composition (based on water content) on different physico-chemical properties was analysed.

Keywords:

• Honey
• Composition
• Viscosity
• pH
• Conductivity
• Sugars

INTRODUCTION

Honey is a concentrated solution of several sugars, a viscous and aromatic liquid, produced by bees from the nectar collected from flowers (or honeydew). Honey contains fructose and glucose as predominant monosaccharides, maltose and sucrose as the most important disaccharides, melezitose as the main trisaccharide, and other low-molecular weight oligosaccharides.[1] The composition of honey (sugars and moisture content) is considered responsible for its physicochemical properties, like density and viscosity. Most of the honeys are supersaturated solutions of glucose, which have a tendency to crystallize spontaneously at room temperature into the form of a glucose monohydrate.[2]

Water content is one of the most important factors that influence the preservation of the honey's quality or its storage. Sensory and physicochemical properties are very important parameters to determine the quality and acceptability of honey, and many studies have been devoted to explore such determinants of the product quality.[3,4] Composition and properties of honey change depending on the floral and honeydew sources used by honeybees, as well as on regional and climatic conditions. Some physicochemical parameters have been studied for their use in the identification of botanical and geographical origin of honey.[5] Physical and chemical properties of different kinds of honey have been reported by many scientists.[6] Electrical conductivity, ash content, and pH are widely used to discriminate different types of honeys. Electrical conductivity has been found as a good criterion of the botanical origin of honey. In the last decade, an important number of studies has analysed the influence of composition and temperature upon viscosity due to its importance upon different processing operation.[7,8] Certain discrepancy exists between the results obtained for different authors on the classification of the honeys between Newtonian and non-Newtonian fluids. Previous studies of our group have showed a slight non-Newtonian behaviour when the shear rate applied is low.[8] For this reason, the viscosimetric behaviour of honey samples is commonly analyzed on the basis of a Newtonian behaviour. Also, recent studies analysed other properties, such as total oxidant status, total antioxidant capacity, and total phenols, in order to certify honey characteristics.[9] Other studies[10,11] analysed other characteristics, such as the presence of pesticide residues, in the kind of honey employed in present work. The aim of this study was to determine several physico-chemical properties (density, viscosity, water content, water activity, glass transition temperature, pH, electrical conductivity, sugar composition, and ash content) for Galicia honeys, and to compare with the corresponding values obtained for honeys from different geographical origins.

MATERIALS AND METHODS

Eleven samples of honeys produced in various regions of the north-west of Spain (Galicia) were collected from beekeepers and obtained from local markets. The samples were stored in a refrigerator in glass containers until their analysis. Before being used, they were warmed up to 55°C to dissolve any crystals and kept in flasks at 30°C to remove air bubbles until its use.

Refractive Index and Density

Refractive index was determined using an Atago RX-5000 refractometer (Atago Co., Tokyo, Japan). Before doing the measurements, the refractometer was calibrated using bidistilled-deionized water in accordance to the instrument's instructions. Water was circulated into the apparatus through a thermostatically controlled bath, kept constant at 20°C. The refractive index measurements were done after the honey sample reached the refractometer's constant temperature. A pycnometer technique has been employed for density determination. Density was obtained by determining the weight of a bottle (10 mL) filled with each honey sample. The difference between the weight of the bottle with and without honey has been used to calculate the density value. The pycnometer was thermostatizated at 20°C.

Viscosity and Glass Transition Temperature

An Anton Paar DV-1P digital thermostated rotational viscometer (Anton Paar GmbH, Fraz, Austria) with two coaxial cylinders was used to carry out the rheological measurements. The viscometer was connected to a thermostat-cryostat and the temperature was maintained constant at 20°C. The glass transition temperature was measured with a differential scanning calorimeter (Mettler-Toledo). For the differential scanning calorimetry (DSC) studies, honey samples (40–50 mg aprox) were rapidly cooled with liquid nitrogen to values near −100°C, held for 5 min before heating them to 30°C at 20°C min⁻¹. The glass transition temperature was calculated using experimental data obtained in the DSC equipment, defined as the inflection point of the heat capacity change.

Water Content and Water Activity

Moisture in honey was determined using the value of refractive index that was obtained from the experimental procedure previously described. The water content has been calculated using the corresponding values of refractive index of each honey and the relation employed by Abu-Jdayil et al.[12] based on Chataway data.[13] The samples water activity (a_w ) was measured at 20°C using an Aqualab CX-2 water activity meter (Decagon Devices Inc., Pullman, WA, USA). The determination of water activity values was performed twice: before and after the heating of the samples at 55°C for 1 h. This heat treatment was carried out to dissolve crystals or nuclei, which might be present in honey and could influence the system water activity.

pH and Electrical Conductivity

The pH was measured with a pH meter Crison 2000 (Crison Instruments, Barcelona, Spain) in a 10% (w/v) solution of honey in distilled water.[14] Electrical conductivity was measured in a 20% (w/v) solution of honey in deionised water[15] with the Metler-Toledo SevenMulti conductimeter (Mettler-Toledo S.A.E., Barcelona, Spain). Both physical properties have been determined at 20°C.[16]

Sugars and Ash

Glucose, fructose, sucrose, and maltose were determined by employing the Boehringer-Mannheim enzymatic test[17] with a Kontron 922 Uvikon double beam spectrophotometer (NorthStart Scientific, Leeds, UK). All samples were analysed in duplicate. Ash percentage was measured by means of a calcinations process in a muffle furnace, for one night in a furnace at 550°C, until reached to a constant mass.[18]

RESULTS AND DISCUSSION

The results of physico-chemicals analysis of honey from different samples included in the guarantee of origin and quality “miel de Galicia” (Galician honey) are summarized in Tables 1 and 2. Table 1 shows the values of refractive index, density, viscosity, and glass transition temperature of the different samples of “miel de Galicia” honeys. The values corresponding to refractive index, density, and the glass transition temperature take similar values for all honey samples analysed in the present work. However, important changes have been observed for viscosity, since these viscosity values varied from 7.77 Pa s until 12.63 Pa s. There are great differences between the viscosity value obtained in previous studies and the present one, being an example the Brazilian honeys, with a maximum value for viscosity of 5.09 Pa s.[19] In other studies, the viscosity value for different honeys was similar or higher (maximum values around 20 Pa s) than the values obtained in the present work.[20] On the other hand, the most important difference was found when Galician honeys were compared with Greek ones,[2] that show an important number of honeys with similar viscosity, even though other samples show high viscosity values near to 200 Pa s. These great changes in viscosity were assigned to differences on honey composition and water content. One of the most important influences that has great important on viscosity is the water content that produces a clear decrease in the viscosity value.

Table 1 Physico-chemical properties (refractive index, density, viscosity, and glass transition temperature) of Galician honey samples

Sample	nD (-)	ρ (g·cm^−3)	η (Pa·s)	Tg (°C)
A	1.49317	1.2684	8.95	−43.04
B	1.49437	1.2690	14.80	−41.24
C	1.49375	1.2629	12.63	−42.92
D	1.49360	1.2695	10.46	−42.17
E	1.49391	1.2669	10.26	−42.66
F	1.49312	1.2648	10.30	−43.01
G	1.49334	1.2671	10.48	−42.85
H	1.49475	1.2680	10.50	−42.34
I	1.49337	1.2642	9.70	−43.31
J	1.49241	1.2663	7.77	−43.17
K	1.49403	1.2697	12.55	−43.47

Table 2 Physico-chemical properties (water content, water activity, pH, electrical conductivity, and ash content) of Galician honey samples

Sample	Moisture (g/100 g honey)	aw	pH(-)	κ (mS cm^−1)	Ash (g/100 g honey)
A	17.37	0.593	3.816	0.734	0.2
B	16.89	0.579	4.493	1.389	0.4
C	17.14	0.592	4.361	0.997	0.4
D	17.20	0.586	4.136	0.920	0.5
E	17.07	0.588	4.534	1.337	0.5
F	17.39	0.596	3.921	0.763	0.4
G	17.30	0.592	4.142	0.816	0.5
H	17.14	0.587	4.208	0.825	0.5
I	17.29	0.599	4.267	0.748	0.5
J	17.67	0.597	4.003	0.754	0.3
K	17.03	0.602	4.498	1.219	0.5

The previously commented physico-chemical properties, such as refractive index, density, and glass transition temperatures, didn't show important differences in honey samples analysed, and only slight changes were observed. Regarding density, the influence of honey composition was considered negligible and the comparison of the present data with other studies[21] shows that Galician honeys show a slight lower value (1.27 g cm⁻³) than other honeys, like Algerian honeys with a medium density value of 1.41 g cm⁻³.

In relation to the refractive index value and the glass transition temperature, a similar influence than previously commented one for viscosity was observed, because an increase in the refractive index (due to a decrease in water content) corresponds with an increase in viscosity too. As it was above mentioned, the variations found in the refractive index value and the glass transition temperatures were slightly varied. This behaviour is related to the presence of water in honey samples, since a decrease in water content produces an increase in both variables. A similar value of refractive index indicates that the water content remains nearly constant in honeys analysed in this work. A comparison between present and previous studies shows that the refractive index takes similar values for different honey samples.[21,22]

Figure 1 shows the experimental data employed to determine the glass transition temperature, as well as the methodology employed to calculate this parameter. The calculated data for glass transition temperature show slight variations between different samples of honey and these values are similar to the previous results obtained for other honeys, with variations from −34.6°C to −47.15°C.[2]

Figure 1 Glass transition temperature determination using Differential Scanning Calorimetry (DSC) for Galician honeys. (○) Honey C. (•) Honey D.

The great part of the physicochemical properties is widely related to honey composition,[15] and the conclusions of previous studies indicate that the presence of water in food plays an important role in the final value of these properties. Taking into account the influence of water, Table 2 summarises the value of other properties, like water content, water activity, pH, electrical conductivity, and ash content. The water content is very interesting, based on the previous discussion about the influence of this parameter on the value of viscosity, refractive index, and glass transition temperature. On the basis of the refractive index value for each honey and the equation obtained from the experimental data of Chataway,[13] the water content was calculated. The European regulation requires a maximum of 21% of moisture in honey for a safety against fermentation, but the specific regulation “miel de Galicia” requires a maximum of 18%.[23] In Table 2, we can observe that the water content for all samples does not reach the value of 18%.

The effect of water content on viscosity and glass transition temperature is shown in Fig. 2. The obtained behaviours for viscosity and glass transition temperature show that an increase in water content produces a decrease in both properties, with a linear trend for viscosity. For the glass transition temperature, the observed trend was potential. On the other hand, in relation to the value of water activity, similar values for all samples were obtained. Previous studies have concluded that there is a linear trend between water activity and solid content or water content,[24] and our experimental data confirms this behaviour for Galician honeys.

Figure 2 Influence of water content upon viscosity (○) and glass transition temperature (•) upon Galician honeys.

The value of pH and electrical conductivity has also been determined in the present work for all honey's samples, and these values are listed in Table 2. pH values are close to four for all honeys, and these values are in agreement with previous studies,[16,25,26] though for Brazilian honeys the pH values were more acidic than the previous commented results,[19] obtaining values lower that the country normative. In relation to the electrical conductivity values for Galician honeys, it is similar to previous studies for honeys from other countries, even for honeys with different pH values.[16,25,26] Several honey samples studied in present work reached values higher than 0.8 mS·cm⁻¹ and these values indicated that the presence of chestnut and eucalyptus pollen in the honeys.[27]

The pH and the electrical conductivity measurements depend on the ions concentration and the kind of ions present in honeys. In the present work, an increase in the pH corresponds to another increase in the electrical conductivity value (Fig. 3), and this behaviour indicates that the presence of H⁺ ions have a great effect on the electrical conductivity. This behaviour agrees with a previous one[16] for the great part of the analysed samples, though for certain samples the behaviour obtained was the opposite.

Figure 3 Relation between electrical conductivity and pH of Galicia honeys.

The ash content of the studied honey samples ranges from 0.2 to 0.5%. This range is not wide being compared to previous studies that obtained ash contents near to 0.02%.[16] Ash values are below 0.6% in all cases, as expected for nectar honeys (European Union standards). The ash content differences are dependent on the type of soil where the original nectar bearing plant is located. The obtained values for honeys samples in the present work are within the ranges for other studies.[16,21,25]

Reducing sugars, mainly glucose and fructose, represent the largest portion of honey composition, but small quantities of other sugars are also present, such as sucrose and maltose (see Table 3). Adding the percentages of fructose, glucose, and maltose was above 75%. This value was higher than previously obtained percentages, when honeys from other origins were analysed,[25] but other studies agree with the values obtained in this work.[28] In relation to the sucrose content, it is important to indicate that low values (less than 3%) were obtained for all samples, indicating that Galician honeys fulfil the European guideline 74/409/EC (maximum sucrose content of 5%). The ratio fructose/glucose takes higher values than others obtained in previous studies.[16,25,28] For different samples employed in the present work, these results are related to the botanical origin of honey, but for other samples (A, B, J, and K), an excess of ripening process could be produced.

Table 3 Analysis of sugar composition in Galician honey samples

Sample	Glucose (g/100 g honey)	Fructose (g/100 g honey)	Sucrose (g/100 g honey)	Maltose (g/100 g honey)	Ratio fructose/glucose
A	27.7	39.4	1.5	10.6	1.43
B	23.7	39.4	1.6	15.2	1.67
C	24.3	38.3	1.5	11.9	1.59
D	26.7	38.7	1.0	10.9	1.45
E	23.4	38.3	2.3	10.7	1.64
F	23.1	45.3	0.5	7.2	1.96
G	23.4	37.7	2.9	11.8	1.61
H	25.2	39.9	1.0	11.2	1.59
I	23.9	37.5	2.8	13.7	1.56
J	28.5	39.1	1.0	7.2	1.37
K	21.9	39.0	1.2	13.4	1.79

CONCLUSIONS

The present study describes the variability of some physicochemical characteristics of 11 honeys from the north-west of Spain (Galicia) and the experimental results indicated that these samples agree with the requirements of Galician, Spanish, and European requirements. The different physicochemical properties were compared to other honeys from different geographical origins and certain differences have been found. For viscosity, medium values were obtained, lejos de los extremos observados for honeys from other origins. In relation with other properties as pH, ash content, density, refractive index, and glass transition temperature, similar values were obtained for all samples and similar than other honeys. Higher values for electrical conductivity than 0.8 mS·cm⁻¹ were obtained for several samples probably due to the presence of chesnut and eucalyptus pollen.