**4. Honey as prebiotic**

The most well‐known properties of honey are its antioxidant and antimicrobial contents. Different types of honey contain different characteristics and properties. Hence, the different sources of honey reflect its content and characteristics.

Prebiotics are substances that beneficially affect the host by selectively stimulating the growth and/or activity of one or a limited number of bacteria in the colon, the probiotic bacteria. Traditionally, prebiotics were related to nondigestible oligosaccharides and poly‐ saccharides substances, which beneficially affect the host by selectively stimulating the growth and/or activity of one or a limited number of bacteria in the intestinal colon [18, 19]. However, this concept should be expanded to other substances, present in honey, which selectively benefit probiotic bacteria by stimulating is growth or activity. Most of the anti‐ oxidant compounds present in honey affect the viability of a series of undesirable microor‐ ganisms but does not affect probiotic bacteria or, in many cases, even stimulate their growth or activity [20–22].

Honey oligosaccharides had a potential prebiotic activity. These compounds selectively stimulate the growth of beneficial microorganisms, such as *Lactobacillus* and *Bifidobacterium* [23, 24]. Sanz et al. [24] conducted a study on how honey oligosaccharide affects the bacteria population in human gut intestinal track (GIT) and found honey that contain higher amount of oligosaccharide resulted in large amount of beneficial bacteria's growth.

The main oligosaccharides found in honeys surveyed in Brazil were the disaccharides, turanose, nigerose, melibiose, sucrose, isomaltose and four trisaccharides, maltotriose, panose, melezitose and raffinose [25]. Sanz et al. [24] found the highest amounts of maltulose and turanose (0.66–3.52 and 0.72–2.87 g/100 g of honey, respectively) in samples of honey from different regions of Spain and commercially available nectar and honeydew honeys. The trisaccharides, melezitose and panose, were the most abundant oligosaccharides from New Zealand honeys [26]. The fructooligosaccharides (FOS) quantified from wild Malaysian honeys were inulobiose, kestose and nystose [23].

Both lactobacilli and bifidobacteria are benefited in environments with low redox potential, and the presence of antioxidant compounds in honey is important in this regard. Flavonoids, amino acids and phenolic acids are the main antioxidant compounds in honey. Most valu‐ able and superior antioxidant compounds of honey such as some phenolic compounds and glutathione are unstable over time and thermolabile. Thus, its final quality is compromised when raw honey goes through conventional thermal processing.

Olofsson et al. [3] reported that 13 lactic acid bacteria symbionts from the honey stomach of honeybees (*Apis mellifera*) were also found in large concentrations in fresh honey as well as having a wide spectrum of antimicrobial activity against various honeybee pathogens and bacteria and yeasts from flowers. According to these authors, many of the unknown healing and antimicrobial properties of honey are linked with these LAB symbionts. Every single member of the LAB microbiota of honeybees produces different bioactive metabo‐ lites. Organic acids were produced by all tested strains but in different amounts. Lactic, formic and acetic acids were produced as well as a wide variety of other interesting metab‐ olites such as benzene and 2‐heptanone and also putative lactic acid bacteria proteins in different honey types, suggesting their importance in honey production and antimicrobial

The most well‐known properties of honey are its antioxidant and antimicrobial contents. Different types of honey contain different characteristics and properties. Hence, the different

Prebiotics are substances that beneficially affect the host by selectively stimulating the growth and/or activity of one or a limited number of bacteria in the colon, the probiotic bacteria. Traditionally, prebiotics were related to nondigestible oligosaccharides and poly‐ saccharides substances, which beneficially affect the host by selectively stimulating the growth and/or activity of one or a limited number of bacteria in the intestinal colon [18, 19]. However, this concept should be expanded to other substances, present in honey, which selectively benefit probiotic bacteria by stimulating is growth or activity. Most of the anti‐ oxidant compounds present in honey affect the viability of a series of undesirable microor‐ ganisms but does not affect probiotic bacteria or, in many cases, even stimulate their growth

Honey oligosaccharides had a potential prebiotic activity. These compounds selectively stimulate the growth of beneficial microorganisms, such as *Lactobacillus* and *Bifidobacterium* [23, 24]. Sanz et al. [24] conducted a study on how honey oligosaccharide affects the bacteria population in human gut intestinal track (GIT) and found honey that contain higher amount

The main oligosaccharides found in honeys surveyed in Brazil were the disaccharides, turanose, nigerose, melibiose, sucrose, isomaltose and four trisaccharides, maltotriose, panose, melezitose and raffinose [25]. Sanz et al. [24] found the highest amounts of maltulose and turanose (0.66–3.52 and 0.72–2.87 g/100 g of honey, respectively) in samples of honey from different regions of Spain and commercially available nectar and honeydew honeys. The trisaccharides, melezitose and panose, were the most abundant oligosaccharides from New Zealand honeys [26]. The fructooligosaccharides (FOS) quantified from wild Malaysian

of oligosaccharide resulted in large amount of beneficial bacteria's growth.

activity.

290 Honey Analysis

**4. Honey as prebiotic**

or activity [20–22].

sources of honey reflect its content and characteristics.

honeys were inulobiose, kestose and nystose [23].

The main criteria for selection of probiotics are resistance to gastrointestinal conditions [14, 27]; characterization of genus, species, strain and its origin [27]; antimicrobial activity, adhesion to the intestinal epithelium, interaction between probiotics and intestinal microbiota of the host; absence of history of pathogenicity and infectivity; metabolic activity of bile salts; lack of hemolytic activity; absence of genes that convey resistance to antibiotics [28]; potential for reducing biofilm formation by pathogenic microorganisms and resistance to lysozyme besides technological properties [29]. As safety criteria, besides being nonpathogenic, the cul‐ tures must have no history of disease, do not deconjugate bile salts or produce toxins, shall not adduce antibiotic resistance genes and do not translocate or induce them, and preferably to be of human origin [27].

We studied the effect of adding 5% of honey to fermented milks on the survival of *Lactobacillus paracasei* and *Lactobacillus rhamnosus* of human origin (isolated from fecal samples of infants), after simulated gastrointestinal tract conditions. The resistance of the examined strains under conditions simulating the gastrointestinal tract was tested as previously described [30] and modified by adding lysozyme (100 μg/mL) to intestinal juice. The production process is shown in **Figure 1**.

Honey did not affect the survival of *L. paracasei* but avoid the reduction of *L. rhamnosus* num‐ ber. Adding honey (5% w/v) in fermented milk positively affects the survival of *L. rhamnosus* during simulated gastrointestinal conditions. In the presence of honey, the population of *L. rhamnosus* after simulated intestinal condition was more than one log cycle higher than control without honey (**Figure 2**).

Similar response was observed with the commercial *L. casei‐*01 (Christian Hansen), which was not affected by the presence of honey, differently of *Bifidobacterium* strains [20].

*Bifidobacterium* are more sensitive to acids than *Lactobacillus* genus. In fermented milk, *Bifidobacterium longum* was more sensitive than *Bifidobacterium brevi* during storage at 10, 20 and 30°C for 10 days. The same was observed with the pH reduction in smoothie yogurt, *B. brevi* was not affected, whereas *Bifidobacterium longum* lost viability during pH reduction from 6.5 to 3.8 [31].

Indeed, the honey has prebiotic effect by stimulating the growth and activity of probiotic bacteria. Besides, because of osmotic constitution and composition of the honey, it acts as protectant to the passage of probiotic bacteria throughout gastrointestinal tract. In fact honey has three functions related to probiotics aspects: it may contain probiotic microorganisms itself, prebiotic substances and protective function to probiotics during the transit by gastro‐ intestinal conditions.

**Figure 1.** Production of probiotic fermented milk added with 5% of honey.

**Figure 2.** Survival of *Lactobacillus rhamnosus and Lactobacillus paracasei* after gastric and internal condition.

Favarin et al. [30] found that suspending free cells of two *Bifidobacterium* strains in honey solu‐ tions resulted in a protective effect, equivalent to the plain microencapsulation with sodium alginate 3% and concluded that microencapsulation and the addition of honey improved the ability of *Bifidobacterium* to tolerate gastrointestinal conditions *in vitro*.
