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Investigation of Stingless Bee Honey from West Sumatra as an Antihyperglycemic Food
1Department of Animal Products Technology and 2Department of Animal Production, Faculty of Animal Science, Universitas Andalas, Padang 25163, Indonesia
Correspondence to:This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Prev Nutr Food Sci 2024; 29(2): 170-177
Published June 30, 2024 https://doi.org/10.3746/pnf.2024.29.2.170
Copyright © The Korean Society of Food Science and Nutrition.
Abstract
Keywords
INTRODUCTION
Hyperglycemia, often called as diabetes mellitus, is a metabolic disorder characterized by elevated blood glucose levels and carbohydrate, lipid, and protein metabolism disruptions. This condition arises from insulin deficiency or resistance (Surya et al., 2014). Hyperglycemia is caused by lipid peroxidation and oxidative stress, which result from an increase in free radical compounds, leading to decreased pancreatic β-cell secretion, inflammation, and endothelial damage (Vijay and Vimukta, 2014).
Antioxidant and phenolic compounds play a substantial role in various biological activities, including acting as antidiabetic agents. Furthermore, the consumption of polyphenol-based antioxidant-rich foods helps in mitigating the formation of reactive free radicals and oxidative stress within the body. Consequently, this can protect pancreatic β-cells and promote their proliferation, thereby activating insulin signaling and secretion (Fatima et al., 2022). Additionally, polyphenolic compounds contribute to inhibiting the α-glucosidase enzyme, which represents another mechanism of antihyperglycemic action. This inhibition leads to reduced blood glucose levels postcarbohydrate consumption in individuals with diabetics as the enzyme aids in the breakdown of complex carbohydrates (Sansenya et al., 2023).
Stingless bee honey, locally known as Galo-galo honey in West Sumatra, is sourced from stingless bees (Apidae; Meliponini). In addition to being a beneficial nutritional source, honey is consumed for its health-promoting properties. Its primary constituents are carbohydrates, specifically fructose and sucrose (Bogdanov et al., 2008). Moreover, honey contains high levels of antioxidants and polyphenolic compounds. Honey from stingless bees exhibits higher antioxidant activity and total polyphenol content compared to honey produced by stinging bees of the genus
Traditionally, high-carbohydrate foods are believed to increase blood glucose levels and are not recommended for consumption by individuals with diabetes. However, recent research suggests that honey can act as an antidiabetic agent. The antihyperglycemic mechanism of honey involves the role of fructose and oligosaccharides in glycemic control (Erejuwa et al., 2012), contribution of flavonoid compounds and its antioxidant activity, which mitigate oxidative stress (Tadera et al., 2006; Garba et al., 2012).
The nutritional profile of honey, including antioxidants and flavonoids, plays a crucial role in its potential as an antihyperglycemic food. However, the composition of honey is influenced by factors such as its geographic origin, botanical nectar source, environmental conditions, climate, and processing techniques. Bee preferences for foraging, behavior, and body size can affect the honey composition, causing variations in the total and types of flavonoids present (Ismail et al., 2016). Despite these factors, the potential antihyperglycemic properties of stingless bee honey from West Sumatra, Indonesia, remain largely unexplored. Therefore, there is a pressing need for studies to investigate the potential of stingless bee honey as an antihyperglycemic food source.
MATERIALS AND METHODS
Materials
Raw honey samples were sourced from bee farms at the Faculty of Animal Science, Universitas Andalas. These farms encompass three varieties of stingless bees:
Experimental design
The study was designed to assess the potential of honey sourced from three varieties of stingless bees as antihyperglycemic food, juxtaposed with honey from the stinging bee,
Determination of α-glucosidase enzyme inhibition activity
The α-glucosidase inhibition activity was assessed following the method outlined by Sancheti et al. (2009). The reaction mixture comprised 50 μL of 0.1 M phosphate buffer (pH 7.0), 25 μL of 4-nitrophenyl a-D-glucopyranoside (dissolved in 0.1 M phosphate buffer, pH 7.0), 10 μL of the sample (10 mg dissolved in 1 mL of dimethyl sulfoxide and aquabides solvent), and 25 μL of α-glucosidase enzyme solution (0.04 units mL−1 in 0.1 M phosphate buffer, pH 7.0). The mixture was then incubated at 37°C for 30 min. Following incubation, the reaction was terminated by adding 100 μL of 0.2 M sodium carbonate solution. The extent of enzymatic hydrolysis of the substrate was determined by measuring the amount of
Determination of antioxidant activity (IC50)
The antioxidant activity was assessed following the method proposed by Salazar-Aranda et al. (2011). Initially, samples (100 g) underwent extraction using the maceration method with methanol (3×600 mL) for 2 h at 29°C. Subsequently, the obtained extracts were filtered and concentrated. For the assay, standard solution, blank, and samples (concentrations of 0.625, 1.25, 2.5, 5, 10, and 20 mg/mL) were added to microplate wells in aliquots of 100 μL. Following this, ethanol (100 μL) and a 1,1-diphenyl-2-picrylhydrazyl (DPPH) solution were added up to 300 μL. The mixture was then incubated for 30 min at 29°C. After incubation, the absorbance was measured using a microplate reader at a wavelength of 517 nm. The free radical scavenging capacity was calculated as the percentage inhibition of DPPH, representing the percentage of the scavenging effect.
Determination of total phenolic content
The analysis of total phenolic content followed the method proposed by Hodzic et al. (2009). Initially, a sample weighing 100 mg was added to 5 mL of 95% ethanol in a covered reaction tube, followed by vortexing. The tube containing the mixture was centrifuged at 1,409
Determination of total flavonoid content
The total flavonoid content was determined following the method described by Pontis et al. (2014), with slight modifications. Initially, 2.5 g sample was dissolved in 5 mL of ethanol, followed by sonication for 15 min and filtration. Subsequently, 1.5 mL of the sample solution was mixed with 0.1 mL of 10% AlCl3 and 0.1 mL of 1 M Na-acetate in 3.3 mL of ethanol. The resulting mixture was incubated for 30 min, after which the absorbance was measured using a UV-Vis spectrophotometer at a wavelength of 425 nm. Quantitative analysis was performed by constructing a standard curve of quercetin concentrations (6, 8, 10, 12, and 14 ppm). Subsequently, the total flavonoid content of the sample was determined using the quercetin standard curve.
FTIR analysis
Honey spectra were recorded following a method Gok et al. (2015) outlined, with slight modifications. For the measurement of honey spectra, 0.1 g of each sample was analyzed using an FTIR spectrometer (PerkinElmer Inc.) equipped with a zinc selenide crystal-fitted diamond single reflection attenuated total reflectance (ATR) accessory. Spectral acquisition was conducted using PerkinElmer Frontier C90704 Spectrum IR Version 10.6.1 software. Infrared spectra were collected within the 4,000∼600 cm−1 range with a 4 cm−1 spectral resolution.
Data analysis
Data analysis was performed utilizing the one-way ANOVA procedure in IBM SPSS version 22 (IBM Corp.). Upon detection of differences indicated by ANOVA, Duncan’s multiple range test was applied at a significance level of 5%.
RESULTS AND DISCUSSION
α-Glucosidase enzyme inhibition activity
The enzyme α-glucosidase plays a pivotal role in elevating blood glucose, and its inhibition can yield beneficial antidiabetic effects by reducing blood glucose levels. Table 1 presents the inhibitory activities of α-glucosidase enzyme in honey sourced from
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Table 1 . α-Glucosidase inhibition activity of stingless bee honey from West Sumatra
Sample % Inhibition Heterotrigona itama 6.79±0.37b Geniotrigona thoracica 2.81±0.27a Kelulut matahari 6.61±0.28b Apis cerana 9.56±0.31c Values are presented as mean±SD of three replication.
Different letters in a column (a-c) show a significant difference (
P <0.05).
The results indicated that the inhibitory activities against α-glucosidase enzymes varied among the different types of honey, ranging from 2.81% to 9.56%. Among the honey samples from stingless bees, no substantial difference (
The findings of this study aligned with those reported by Peláez-Acero et al. (2022), who observed that α-glucosidase enzyme inhibition activities of honey from Mexico ranged from 5.28% to 14.50%. Honey from stingless bees
Antioxidant activity (IC50)
The antioxidant activity in this study was assessed using IC50 values, which indicate the sample concentration required to reduce DPPH activity by 50%. Table 2 presents honey’s antioxidant activity values (IC50) from
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Table 2 . Antioxidant activity (IC50) of stingless bee honey from West Sumatra
Sample IC50 (mg/mL) Heterotrigona itama 52.08±0.87c Geniotrigona thoracica 64.19±0.82d Kelulut matahari 8.82±0.06a Apis cerana 23.56±0.04b Values are presented as mean±SD of three replication.
Different letters in a column (a-d) show a significant difference (
P <0.05).IC50, half maximal inhibitory concentration.
The results revealed considerable differences (
Regarding honey from
Antioxidant compounds play a crucial role in antihyperglycemic activity, with one mechanism involving the reduction of free radicals by these compounds present in honey. This process effectively diminishes oxidative stress and shields organs from oxidative damage (Erejuwa et al., 2009). Sahhugi et al. (2014) also demonstrated that honey sourced from the stingless bee Gelam could mitigate oxidative damage by reducing malondialdehyde levels in rats.
Total phenolic and flavonoid contents
The total phenolic and flavonoid contents in honey sourced from
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Table 3 . Total phenolic and flavonoid contents of stingless bee honey from West Sumatra
Sample Total phenolic content (mg GAE/g) Total flavonoid content (mg EQ/100 g) Heterotrigona itama 59.98±0.72b 1.51±0.01b Geniotrigona thoracica 39.98±0.28a 1.23±0.01a Kelulut matahari 75.80±1.20c 1.96±0.02c Apis cerana 81.02±0.54d 2.18±0.01d Values are presented as mean±SD of three replication.
Different letters in a column (a-d) show a significant difference (
P <0.05).GAE, gallic acid equivalent; EQ, quercetin equivalent.
The total flavonoid content mirrored the pattern observed in the total phenolic content across all four honey samples, indicating significant differences (
The total phenolic and flavonoid contents positively correlate with antioxidant activity, wherein higher levels of polyphenolic compounds are typically associated with higher antioxidant activity (Islam et al., 2017). However, in this study, although honey from
Phenolic and flavonoid compounds have been reported to inhibit α-glucosidase. The findings of this study elucidated a correlation between the total phenolic and flavonoid contents and α-glucosidase enzyme inhibition activity. Specifically, honey sourced from
FTIR spectra interpretations
The results of the FTIR analysis for the four honey samples in the wavenumbers 4,000∼650 cm−1 are depicted in Fig. 1. According to Huang et al. (2020), the region 1,500∼750 cm−1 corresponds to the primary absorption of fructose, glucose, and sucrose, while the peaks at 1,642 and 3,297 cm−1 are indicative of water absorption in honey. Overall, the FTIR spectra of all honey samples exhibited similar patterns. The sample from
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Table 4 . Prediction of Fourier-transform infrared spectrum peaks of stingless bee honey from West Sumatra
Apis cerana Heterotrigona itama Geniotrigona thoracica Kelulut matahari Functional group Reference 768 767 − − Anomeric region of carbohydrate Gallardo-Velázquez et al. (2009) 815 − 818,915 − C-H bending (carbohydrate) Gallardo-Velázquez et al. (2009) 1,030 1,029 1,026 1,028 C-O stretching Subari et al. (2012) 1,251 1,254 1,260 1,254 C-O stretching (C-OH group) in carbohydrate structure Tewari and Irudayaraj (2004); Gallardo-Velázquez et al. (2009) 1,351 1,353 − 1,352 O-H bending on the C-OH group Gallardo-Velázquez et al. (2009) 1,416 1,416 1,419 1,415 Combination of O-H bending from the C-OH group and C-H bending of alkenes Gallardo-Velázquez et al. (2009) 1,643 1,643 1,643 1,642 H-O-H bending vibration Nayik et al. (2019) 2,931 2,934 2,937 2,934 C-H stretching (carboxylic acids) and NH3 stretching band (free amino acids) Gallardo-Velázquez et al. (2009) 3,270 3,269 3,269 3,272 C-H stretching (carboxylic acids) and NH3 stretching band (free amino acids)
-
Figure 1. Fourier-transform infrared-attenuated total reflectance spectra of honey from stingless bee procured from West Sumatra.
Sucrose exhibits bands in the range of 928∼1,427 cm−1 with notable peaks at 994 and 1,049 cm−1, fructose demonstrates in the range of 923∼1,418 cm−1 with a prominent peak at 1,053 cm−1, glucose in the range of 902∼1,431 cm−1 with a key peak at 1,032 cm−1, and maltose in the range of 920∼1,353 cm−1 with a key peak at 1,032 cm−1 (Wang et al., 2010). In this study, dominant peaks were observed at 767∼1,643 cm−1 across all four honey samples, indicating a sugar profile consisting of sucrose, fructose, glucose, and maltose. Fructose and oligosaccharides present in honey, such as palatinose, turanose, raffinose, and isomaltose, have been identified as potential antidiabetic agents (Erejuwa et al., 2012). Fructose in honey can enhance hepatic glucose absorption, glycogen synthesis, and storage, thereby improving glycemic control in diabetes mellitus (Eteraf-Oskouei and Najafi, 2013).
Oligosaccharides found in honey have been associated with several roles as antihyperglycemic agents through various mechanisms, such as their involvement in the gut and gut microbiota as prebiotics, enhancement in glucose-stimulated insulin secretion, and reduction in the rate of glucose absorption (Erejuwa et al., 2011). Additionally, isomaltose, a compound detected in honey, exhibits inhibitory effects on the human incretin-degrading enzyme dipeptidyl peptidase IV (DPP-IV), which is responsible for deactivating glucose-regulating hormones such as glucagon-like peptide-1. Therefore, the inhibitory action of DPP-IV can alleviate diabetic conditions (Bharti et al., 2015; Belay et al., 2017).
Regarding phenolic compounds, peaks in the vibration region spanning 1,175∼940 cm−1 represent C-OH groups and stretching of C-C and C-O in carbohydrate moieties as well as C-O in phenol (Masek et al., 2014); in this study, these peaks are observed at 1,026, 1,028, 1,029, and 1,030 cm−1 in all four honey samples. The region at 1,540∼1,175 cm−1 represents deformations of O-H, C-O, C-H, and C=C corresponding to phenol and flavanol (Masek et al., 2014); in this study these peaks are observed at 1,419∼1,251 cm−1 in all four honey samples.
Thummajitsakul et al. (2023) reported that the abundance of peaks generated by FTIR indicates the high total phenolic content present in ethanol extracts of
Differences in bee species contribute to differences in honey composition, affecting α-glucosidase enzyme inhibition, antioxidant activity (IC50), and total phenolic and flavonoid contents.
The FTIR results revealed that all four honey samples mostly exhibited absorption in the region of 767∼1,643 cm−1, indicating dominant absorption by monosaccharides and disaccharides. These findings underscore the potential of stingless bee honey as an antihyperglycemic food, as evidenced by its inhibitory activity against the α-glucosidase enzyme, antioxidant activity, and comparatively high total phenolic content.
FUNDING
The funding for this study is provided by Faculty of Animal Science, Universitas Andalas (contract no. 001.e/UN. 16.06.D/PT.01/SPP.RDP/FATERNA/2023).
AUTHOR DISCLOSURE STATEMENT
The authors declare no conflict of interest.
AUTHOR CONTRIBUTIONS
Concept and design: RDS, SM, IJ. Analysis and interpretation: RDS, SM, IJ, R. Data collection: RDS, R. Writing the article: RDS. Critical revision of the article: RDS. Final approval of the article: all authors. Statistical analysis: RDS. Obtained funding: RDS, SM. Overall responsibility: RDS.
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Article
Original
Prev Nutr Food Sci 2024; 29(2): 170-177
Published online June 30, 2024 https://doi.org/10.3746/pnf.2024.29.2.170
Copyright © The Korean Society of Food Science and Nutrition.
Investigation of Stingless Bee Honey from West Sumatra as an Antihyperglycemic Food
Rizki Dwi Setiawan1 , Sri Melia1 , Indri Juliyarsi1 , Rusdimansyah2
1Department of Animal Products Technology and 2Department of Animal Production, Faculty of Animal Science, Universitas Andalas, Padang 25163, Indonesia
Correspondence to:Rizki Dwi Setiawan, E-mail: rizkidwi@ansci.unand.ac.id
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
This study aimed to investigate the potential in vitro antihyperglycemic activity of honey sourced from three different species of stingless bees (Heterotrigona itama, Geniotrigona thoracica, and Kelulut matahari) by assessing their α-glucosidase enzyme inhibition, antioxidant activity, and total phenolic and flavonoid contents in comparison with honey from Apis cerana, obtained from West Sumatra, Indonesia. The honey samples were obtained from stingless bee farms at the Faculty of Animal Science, Universitas Andalas. Variations were observed in α-glucosidase enzyme inhibition, antioxidant activity (half maximal inhibitory concentration, IC50), and total phenolic and flavonoid contents among the honey samples from H. itama, G. thoracica, K. matahari, and A. cerana. In terms of α-glucosidase inhibition, honey from the stinging bee A. cerana demonstrated higher inhibition than that from the other three stingless bees species. Honey derived from K. matahari exhibited the lowest IC50 value, indicating its superior antioxidant activity, followed by honey from A. cerana, H. itama, and G. thoracica. The highest total phenolic and flavonoid contents were found in honey from A. cerana, followed by honey from K. matahari, H. itama, and G. thoracica. Analysis using Fourier-transform infrared spectroscopy revealed that the predominant absorptions in all four honey samples were observed at 767∼1,643 cm−1, indicating that absorptions are primarily ascribed to monosaccharides and disaccharides. Additionally, some peaks implied the presence of phenolic and flavonoid compounds. Overall, honey from stingless bees shows promise as an antihyperglycemic food, as evidenced by its α-glucosidase enzyme inhibition activity, antioxidant activity, and relatively high total phenolic content.
Keywords: antihyperglycemic, functional food, honey, stingless bee
INTRODUCTION
Hyperglycemia, often called as diabetes mellitus, is a metabolic disorder characterized by elevated blood glucose levels and carbohydrate, lipid, and protein metabolism disruptions. This condition arises from insulin deficiency or resistance (Surya et al., 2014). Hyperglycemia is caused by lipid peroxidation and oxidative stress, which result from an increase in free radical compounds, leading to decreased pancreatic β-cell secretion, inflammation, and endothelial damage (Vijay and Vimukta, 2014).
Antioxidant and phenolic compounds play a substantial role in various biological activities, including acting as antidiabetic agents. Furthermore, the consumption of polyphenol-based antioxidant-rich foods helps in mitigating the formation of reactive free radicals and oxidative stress within the body. Consequently, this can protect pancreatic β-cells and promote their proliferation, thereby activating insulin signaling and secretion (Fatima et al., 2022). Additionally, polyphenolic compounds contribute to inhibiting the α-glucosidase enzyme, which represents another mechanism of antihyperglycemic action. This inhibition leads to reduced blood glucose levels postcarbohydrate consumption in individuals with diabetics as the enzyme aids in the breakdown of complex carbohydrates (Sansenya et al., 2023).
Stingless bee honey, locally known as Galo-galo honey in West Sumatra, is sourced from stingless bees (Apidae; Meliponini). In addition to being a beneficial nutritional source, honey is consumed for its health-promoting properties. Its primary constituents are carbohydrates, specifically fructose and sucrose (Bogdanov et al., 2008). Moreover, honey contains high levels of antioxidants and polyphenolic compounds. Honey from stingless bees exhibits higher antioxidant activity and total polyphenol content compared to honey produced by stinging bees of the genus
Traditionally, high-carbohydrate foods are believed to increase blood glucose levels and are not recommended for consumption by individuals with diabetes. However, recent research suggests that honey can act as an antidiabetic agent. The antihyperglycemic mechanism of honey involves the role of fructose and oligosaccharides in glycemic control (Erejuwa et al., 2012), contribution of flavonoid compounds and its antioxidant activity, which mitigate oxidative stress (Tadera et al., 2006; Garba et al., 2012).
The nutritional profile of honey, including antioxidants and flavonoids, plays a crucial role in its potential as an antihyperglycemic food. However, the composition of honey is influenced by factors such as its geographic origin, botanical nectar source, environmental conditions, climate, and processing techniques. Bee preferences for foraging, behavior, and body size can affect the honey composition, causing variations in the total and types of flavonoids present (Ismail et al., 2016). Despite these factors, the potential antihyperglycemic properties of stingless bee honey from West Sumatra, Indonesia, remain largely unexplored. Therefore, there is a pressing need for studies to investigate the potential of stingless bee honey as an antihyperglycemic food source.
MATERIALS AND METHODS
Materials
Raw honey samples were sourced from bee farms at the Faculty of Animal Science, Universitas Andalas. These farms encompass three varieties of stingless bees:
Experimental design
The study was designed to assess the potential of honey sourced from three varieties of stingless bees as antihyperglycemic food, juxtaposed with honey from the stinging bee,
Determination of α-glucosidase enzyme inhibition activity
The α-glucosidase inhibition activity was assessed following the method outlined by Sancheti et al. (2009). The reaction mixture comprised 50 μL of 0.1 M phosphate buffer (pH 7.0), 25 μL of 4-nitrophenyl a-D-glucopyranoside (dissolved in 0.1 M phosphate buffer, pH 7.0), 10 μL of the sample (10 mg dissolved in 1 mL of dimethyl sulfoxide and aquabides solvent), and 25 μL of α-glucosidase enzyme solution (0.04 units mL−1 in 0.1 M phosphate buffer, pH 7.0). The mixture was then incubated at 37°C for 30 min. Following incubation, the reaction was terminated by adding 100 μL of 0.2 M sodium carbonate solution. The extent of enzymatic hydrolysis of the substrate was determined by measuring the amount of
Determination of antioxidant activity (IC50)
The antioxidant activity was assessed following the method proposed by Salazar-Aranda et al. (2011). Initially, samples (100 g) underwent extraction using the maceration method with methanol (3×600 mL) for 2 h at 29°C. Subsequently, the obtained extracts were filtered and concentrated. For the assay, standard solution, blank, and samples (concentrations of 0.625, 1.25, 2.5, 5, 10, and 20 mg/mL) were added to microplate wells in aliquots of 100 μL. Following this, ethanol (100 μL) and a 1,1-diphenyl-2-picrylhydrazyl (DPPH) solution were added up to 300 μL. The mixture was then incubated for 30 min at 29°C. After incubation, the absorbance was measured using a microplate reader at a wavelength of 517 nm. The free radical scavenging capacity was calculated as the percentage inhibition of DPPH, representing the percentage of the scavenging effect.
Determination of total phenolic content
The analysis of total phenolic content followed the method proposed by Hodzic et al. (2009). Initially, a sample weighing 100 mg was added to 5 mL of 95% ethanol in a covered reaction tube, followed by vortexing. The tube containing the mixture was centrifuged at 1,409
Determination of total flavonoid content
The total flavonoid content was determined following the method described by Pontis et al. (2014), with slight modifications. Initially, 2.5 g sample was dissolved in 5 mL of ethanol, followed by sonication for 15 min and filtration. Subsequently, 1.5 mL of the sample solution was mixed with 0.1 mL of 10% AlCl3 and 0.1 mL of 1 M Na-acetate in 3.3 mL of ethanol. The resulting mixture was incubated for 30 min, after which the absorbance was measured using a UV-Vis spectrophotometer at a wavelength of 425 nm. Quantitative analysis was performed by constructing a standard curve of quercetin concentrations (6, 8, 10, 12, and 14 ppm). Subsequently, the total flavonoid content of the sample was determined using the quercetin standard curve.
FTIR analysis
Honey spectra were recorded following a method Gok et al. (2015) outlined, with slight modifications. For the measurement of honey spectra, 0.1 g of each sample was analyzed using an FTIR spectrometer (PerkinElmer Inc.) equipped with a zinc selenide crystal-fitted diamond single reflection attenuated total reflectance (ATR) accessory. Spectral acquisition was conducted using PerkinElmer Frontier C90704 Spectrum IR Version 10.6.1 software. Infrared spectra were collected within the 4,000∼600 cm−1 range with a 4 cm−1 spectral resolution.
Data analysis
Data analysis was performed utilizing the one-way ANOVA procedure in IBM SPSS version 22 (IBM Corp.). Upon detection of differences indicated by ANOVA, Duncan’s multiple range test was applied at a significance level of 5%.
RESULTS AND DISCUSSION
α-Glucosidase enzyme inhibition activity
The enzyme α-glucosidase plays a pivotal role in elevating blood glucose, and its inhibition can yield beneficial antidiabetic effects by reducing blood glucose levels. Table 1 presents the inhibitory activities of α-glucosidase enzyme in honey sourced from
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Table 1 . α-Glucosidase inhibition activity of stingless bee honey from West Sumatra.
Sample % Inhibition Heterotrigona itama 6.79±0.37b Geniotrigona thoracica 2.81±0.27a Kelulut matahari 6.61±0.28b Apis cerana 9.56±0.31c Values are presented as mean±SD of three replication..
Different letters in a column (a-c) show a significant difference (
P <0.05)..
The results indicated that the inhibitory activities against α-glucosidase enzymes varied among the different types of honey, ranging from 2.81% to 9.56%. Among the honey samples from stingless bees, no substantial difference (
The findings of this study aligned with those reported by Peláez-Acero et al. (2022), who observed that α-glucosidase enzyme inhibition activities of honey from Mexico ranged from 5.28% to 14.50%. Honey from stingless bees
Antioxidant activity (IC50)
The antioxidant activity in this study was assessed using IC50 values, which indicate the sample concentration required to reduce DPPH activity by 50%. Table 2 presents honey’s antioxidant activity values (IC50) from
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Table 2 . Antioxidant activity (IC50) of stingless bee honey from West Sumatra.
Sample IC50 (mg/mL) Heterotrigona itama 52.08±0.87c Geniotrigona thoracica 64.19±0.82d Kelulut matahari 8.82±0.06a Apis cerana 23.56±0.04b Values are presented as mean±SD of three replication..
Different letters in a column (a-d) show a significant difference (
P <0.05)..IC50, half maximal inhibitory concentration..
The results revealed considerable differences (
Regarding honey from
Antioxidant compounds play a crucial role in antihyperglycemic activity, with one mechanism involving the reduction of free radicals by these compounds present in honey. This process effectively diminishes oxidative stress and shields organs from oxidative damage (Erejuwa et al., 2009). Sahhugi et al. (2014) also demonstrated that honey sourced from the stingless bee Gelam could mitigate oxidative damage by reducing malondialdehyde levels in rats.
Total phenolic and flavonoid contents
The total phenolic and flavonoid contents in honey sourced from
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Table 3 . Total phenolic and flavonoid contents of stingless bee honey from West Sumatra.
Sample Total phenolic content (mg GAE/g) Total flavonoid content (mg EQ/100 g) Heterotrigona itama 59.98±0.72b 1.51±0.01b Geniotrigona thoracica 39.98±0.28a 1.23±0.01a Kelulut matahari 75.80±1.20c 1.96±0.02c Apis cerana 81.02±0.54d 2.18±0.01d Values are presented as mean±SD of three replication..
Different letters in a column (a-d) show a significant difference (
P <0.05)..GAE, gallic acid equivalent; EQ, quercetin equivalent..
The total flavonoid content mirrored the pattern observed in the total phenolic content across all four honey samples, indicating significant differences (
The total phenolic and flavonoid contents positively correlate with antioxidant activity, wherein higher levels of polyphenolic compounds are typically associated with higher antioxidant activity (Islam et al., 2017). However, in this study, although honey from
Phenolic and flavonoid compounds have been reported to inhibit α-glucosidase. The findings of this study elucidated a correlation between the total phenolic and flavonoid contents and α-glucosidase enzyme inhibition activity. Specifically, honey sourced from
FTIR spectra interpretations
The results of the FTIR analysis for the four honey samples in the wavenumbers 4,000∼650 cm−1 are depicted in Fig. 1. According to Huang et al. (2020), the region 1,500∼750 cm−1 corresponds to the primary absorption of fructose, glucose, and sucrose, while the peaks at 1,642 and 3,297 cm−1 are indicative of water absorption in honey. Overall, the FTIR spectra of all honey samples exhibited similar patterns. The sample from
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Table 4 . Prediction of Fourier-transform infrared spectrum peaks of stingless bee honey from West Sumatra.
Apis cerana Heterotrigona itama Geniotrigona thoracica Kelulut matahari Functional group Reference 768 767 − − Anomeric region of carbohydrate Gallardo-Velázquez et al. (2009) 815 − 818,915 − C-H bending (carbohydrate) Gallardo-Velázquez et al. (2009) 1,030 1,029 1,026 1,028 C-O stretching Subari et al. (2012) 1,251 1,254 1,260 1,254 C-O stretching (C-OH group) in carbohydrate structure Tewari and Irudayaraj (2004); Gallardo-Velázquez et al. (2009) 1,351 1,353 − 1,352 O-H bending on the C-OH group Gallardo-Velázquez et al. (2009) 1,416 1,416 1,419 1,415 Combination of O-H bending from the C-OH group and C-H bending of alkenes Gallardo-Velázquez et al. (2009) 1,643 1,643 1,643 1,642 H-O-H bending vibration Nayik et al. (2019) 2,931 2,934 2,937 2,934 C-H stretching (carboxylic acids) and NH3 stretching band (free amino acids) Gallardo-Velázquez et al. (2009) 3,270 3,269 3,269 3,272 C-H stretching (carboxylic acids) and NH3 stretching band (free amino acids)
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Figure 1. Fourier-transform infrared-attenuated total reflectance spectra of honey from stingless bee procured from West Sumatra.
Sucrose exhibits bands in the range of 928∼1,427 cm−1 with notable peaks at 994 and 1,049 cm−1, fructose demonstrates in the range of 923∼1,418 cm−1 with a prominent peak at 1,053 cm−1, glucose in the range of 902∼1,431 cm−1 with a key peak at 1,032 cm−1, and maltose in the range of 920∼1,353 cm−1 with a key peak at 1,032 cm−1 (Wang et al., 2010). In this study, dominant peaks were observed at 767∼1,643 cm−1 across all four honey samples, indicating a sugar profile consisting of sucrose, fructose, glucose, and maltose. Fructose and oligosaccharides present in honey, such as palatinose, turanose, raffinose, and isomaltose, have been identified as potential antidiabetic agents (Erejuwa et al., 2012). Fructose in honey can enhance hepatic glucose absorption, glycogen synthesis, and storage, thereby improving glycemic control in diabetes mellitus (Eteraf-Oskouei and Najafi, 2013).
Oligosaccharides found in honey have been associated with several roles as antihyperglycemic agents through various mechanisms, such as their involvement in the gut and gut microbiota as prebiotics, enhancement in glucose-stimulated insulin secretion, and reduction in the rate of glucose absorption (Erejuwa et al., 2011). Additionally, isomaltose, a compound detected in honey, exhibits inhibitory effects on the human incretin-degrading enzyme dipeptidyl peptidase IV (DPP-IV), which is responsible for deactivating glucose-regulating hormones such as glucagon-like peptide-1. Therefore, the inhibitory action of DPP-IV can alleviate diabetic conditions (Bharti et al., 2015; Belay et al., 2017).
Regarding phenolic compounds, peaks in the vibration region spanning 1,175∼940 cm−1 represent C-OH groups and stretching of C-C and C-O in carbohydrate moieties as well as C-O in phenol (Masek et al., 2014); in this study, these peaks are observed at 1,026, 1,028, 1,029, and 1,030 cm−1 in all four honey samples. The region at 1,540∼1,175 cm−1 represents deformations of O-H, C-O, C-H, and C=C corresponding to phenol and flavanol (Masek et al., 2014); in this study these peaks are observed at 1,419∼1,251 cm−1 in all four honey samples.
Thummajitsakul et al. (2023) reported that the abundance of peaks generated by FTIR indicates the high total phenolic content present in ethanol extracts of
Differences in bee species contribute to differences in honey composition, affecting α-glucosidase enzyme inhibition, antioxidant activity (IC50), and total phenolic and flavonoid contents.
The FTIR results revealed that all four honey samples mostly exhibited absorption in the region of 767∼1,643 cm−1, indicating dominant absorption by monosaccharides and disaccharides. These findings underscore the potential of stingless bee honey as an antihyperglycemic food, as evidenced by its inhibitory activity against the α-glucosidase enzyme, antioxidant activity, and comparatively high total phenolic content.
FUNDING
The funding for this study is provided by Faculty of Animal Science, Universitas Andalas (contract no. 001.e/UN. 16.06.D/PT.01/SPP.RDP/FATERNA/2023).
AUTHOR DISCLOSURE STATEMENT
The authors declare no conflict of interest.
AUTHOR CONTRIBUTIONS
Concept and design: RDS, SM, IJ. Analysis and interpretation: RDS, SM, IJ, R. Data collection: RDS, R. Writing the article: RDS. Critical revision of the article: RDS. Final approval of the article: all authors. Statistical analysis: RDS. Obtained funding: RDS, SM. Overall responsibility: RDS.
Fig 1.
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Table 1 . α-Glucosidase inhibition activity of stingless bee honey from West Sumatra
Sample % Inhibition Heterotrigona itama 6.79±0.37b Geniotrigona thoracica 2.81±0.27a Kelulut matahari 6.61±0.28b Apis cerana 9.56±0.31c Values are presented as mean±SD of three replication.
Different letters in a column (a-c) show a significant difference (
P <0.05).
-
Table 2 . Antioxidant activity (IC50) of stingless bee honey from West Sumatra
Sample IC50 (mg/mL) Heterotrigona itama 52.08±0.87c Geniotrigona thoracica 64.19±0.82d Kelulut matahari 8.82±0.06a Apis cerana 23.56±0.04b Values are presented as mean±SD of three replication.
Different letters in a column (a-d) show a significant difference (
P <0.05).IC50, half maximal inhibitory concentration.
-
Table 3 . Total phenolic and flavonoid contents of stingless bee honey from West Sumatra
Sample Total phenolic content (mg GAE/g) Total flavonoid content (mg EQ/100 g) Heterotrigona itama 59.98±0.72b 1.51±0.01b Geniotrigona thoracica 39.98±0.28a 1.23±0.01a Kelulut matahari 75.80±1.20c 1.96±0.02c Apis cerana 81.02±0.54d 2.18±0.01d Values are presented as mean±SD of three replication.
Different letters in a column (a-d) show a significant difference (
P <0.05).GAE, gallic acid equivalent; EQ, quercetin equivalent.
-
Table 4 . Prediction of Fourier-transform infrared spectrum peaks of stingless bee honey from West Sumatra
Apis cerana Heterotrigona itama Geniotrigona thoracica Kelulut matahari Functional group Reference 768 767 − − Anomeric region of carbohydrate Gallardo-Velázquez et al. (2009) 815 − 818,915 − C-H bending (carbohydrate) Gallardo-Velázquez et al. (2009) 1,030 1,029 1,026 1,028 C-O stretching Subari et al. (2012) 1,251 1,254 1,260 1,254 C-O stretching (C-OH group) in carbohydrate structure Tewari and Irudayaraj (2004); Gallardo-Velázquez et al. (2009) 1,351 1,353 − 1,352 O-H bending on the C-OH group Gallardo-Velázquez et al. (2009) 1,416 1,416 1,419 1,415 Combination of O-H bending from the C-OH group and C-H bending of alkenes Gallardo-Velázquez et al. (2009) 1,643 1,643 1,643 1,642 H-O-H bending vibration Nayik et al. (2019) 2,931 2,934 2,937 2,934 C-H stretching (carboxylic acids) and NH3 stretching band (free amino acids) Gallardo-Velázquez et al. (2009) 3,270 3,269 3,269 3,272 C-H stretching (carboxylic acids) and NH3 stretching band (free amino acids)
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