Influence of origins and bee species on physicochemical, antioxidant properties and botanical discrimination of stingless bee honey

Figures & data

Figure 1. Stingless bee. Heterotrigona itama (a); Geniotrigona thoracica (b)

Table 1. Description of the source of stingless bee honey used in this study

Species	Location	Botanical origin	Scientific name
Heterotrigona itama Geniotrigona thoracica	Terengganu Johor Pahang	Gelam Acacia Starfruit	Meleleuca cajaputi Acacia mangium Averrhoa carambola L
Apis mellifera	Johor	Acacia	Acacia mangium

Figure 2. Malaysian map indicating the location of honey sampling

Table 2. Physicochemical parameters of stingless bee honey from different floral origins

Parameters	Geniotrigona thoracica			Apis mellifera	Heterotrigona itama			Apis mellifera
	Acacia	Starfruit	Gelam		Acacia	Starfruit	Gelam
Moisture content (g/100g honey)	33.93^Aa ± 1.05	21.32^Ba ± 1.78	22.01^Ba ± 1.37	14.74^C ± 0.11	26.28^Ab ± 0.47	19.49^Cb ± 0.70	22.53^Ba ± 1.22	14.74^D ± 0.11
Water activity (aw)	0.87^Aa ± 0.00	0.78^Ba ± 0.02	0.79^Ba ± 0.01	0.67^C ± 0.01	0.82^Ab ± 0.00	0.76^Ba ± 0.01	0.77^Ba ± 0.01	0.67^C ± 0.01
pH	3.23^Cc ± 0.02	3.17^Cb ± 0.00	3.40^Bb ± 0.07	3.55^A ± 0.03	3.35^Bb ± 0.01	3.19^Cb ± 0.04	3.38^Bb ± 0.02	3.55^A ± 0.03
Free acidity (meq/kg honey)	136.50^Ba ± 6.36	170.50 ^Ab ± 4.00	101.83^Ca ± 9.19	38.17^D ± 0.23	64.50^Cb ± 4.00	207.67^Aa ± 3.30	103.00^Ba ± 2.83	38.17^D ± 0.23
Total soluble solid (Brix)	60.85^Cc ± 0.07	71.20^Bb ± 1.56	70.15^Bb ± 0.63	76.70^A ± 0.57	66.25^Cb ± 0.35	72.25^Bb ± 0.92	70.30^Bb ± 1.27	76.70^A ± 0.57
Ash (g/100g honey)	0.24^Aa ± 0.01	0.07^Cb ± 0.007	0.13^Bb ± 0.007	0.25^A ± 0.007	0.22^Ba ± 0.00	0.09^Cb ± 0.007	0.23^Aa ± 0.007	0.25^A ± 0.007
HMF (mg/kg honey)	ND^Aa	ND^Ab	ND^Ab	ND^A	ND^Ba	0.14^Aa ± 0.08	0.11^Aa ± 0.04	ND^B

ND: Not detected

Means with a different capital letters in the same row are significantly difference (p < 0.05) between honeys from different floral origins produced by the same bee species. Different small letters in the same row is significantly difference (p < 0.05) between honeys produced by different bee species from the same floral origin.

Table 3. Colour characteristics of stingless bee honey from different floral origins

	Geniotrigona thorocica				Heterotrigon aitama
Units	Acacia	Starfruit	Gelam	Apis mellifera	Acacia	Starfruit	Gelam	Apis mellifera
L*	92.14^Aa ± 1.88	92.57^Aa ± 3.06	85.06^Ba ± 2.35	84.66^B ± 2.18	92.49^Aa ± 0.34	86.60^Ba ± 1.34	73.98^Cb ± 1.34	84.66^B ± 2.18
a*	−2.08^Bb ± 0.08	−2.35^Bb ± 0.52	0.01^Bb ± 1.49	6.25^A ± 2.17	−1.46^Cb ± 0.18	−0.47^Cb ± 1.06	16.65^Aa ± 0.76	6.25^B ± 2.17
b*	24.62^Cb ± 4.75	39.06^Ba ± 3.60	62.01^Ab ± 0.07	41.34^B ± 5.69	29.42^Cab ± 0.77	52.70^Ba ± 7.69	88.66^Aa ± 3.88	41.34^BCa ± 5.69

a* indicates the degree of red (+a*) or green (-a*). The b* indicate the degree of yellow (+b*) or blue (-b*). The L* indicates the degree of lightness (0 for black until 100 for white). Means with a different capital letters in the same row are significantly different (p < 0.05) between honeys from different floral origins produced by the same bee species. Different small letters in the same row is significantly different (p < 0.05) between honeys produced by different bee species from the same floral origin.

Table 4. Sugar content (g/100 g honey), fructose/glucose (F/G) and glucose/water (G/W) of stingless bee honey from different floral origins

Parameter	Geniotrigona Thoracica			Apis Mellifera	Heterotrigona Itama			Apis Mellifera
	Acacia	Starfruit	Gelam		Acacia	Starfruit	Gelam
Fructose	7.89^Cb ± 0.36	9.34^Ba ± 0.06	8.43^BCb ± 0.11	24.87^A ± 0.56	13.00^Ba ± 0.21	9.68^Ca ± 0.72	13.58^Ba ± 0.33	24.87^A ± 0.56
Glucose	8.12^Cb ± 0.95	11.21^Bb ± 0.10	6.95^Cb ± 0.34	26.17^A ± 0.55	11.69^Ba ± 0.56	8.14^Ca ± 0.47	10.53^Ba ± 0.22	26.17^A ± 0.55
Sucrose	ND^Ba	ND^Ba	ND^Ba	3.86^A ± 1.45	ND^Ba ± 0.13	ND^Ba ± 0.11	ND^Ba ± 0.12	3.86^A ± 1.45
Maltose	28.96^Ba ± 1.17	40.82^Aa ± 0.79	43.15^Aa ± 2.36	ND^C	22.56^Cb ± 1.22	37.35^Ab ± 0.58	31.50^Bb ± 2.11	ND^D
Total Sugar	44.98^Ca ± 1.75	61.37^Aa ± 0.63	58.52^Aba ±2.82	54.90^B ±1.43	47.25^Ba ± 0.87	55.17^Ab ±0.83	55.61^Aa ± 2.00	54.90^A ± 1.43
F/G	1.13^Aba ± 0.05	0.83^Bb ± 0.002	1.37^Aa ± 0.26	0.95^B ± 0.04	1.14^Ba ± 0.04	1.08^BCa ± 0.01	1.56^Aa ± 0.07	0.95^C ± 0.04
G/W	0.24^Db ± 0.03	0.53^Ba ± 0.01	0.32^Cb ± 0.02	1.78^A ± 0.04	0.45^Ba ± 0.02	0.42^Bb ± 0.02	0.47^Ba ± 0.01	1.78^A ± 0.04

ND = Not detected; F/G = Fructose/glucose; G/W = Glucose/water

Means with different capital letters in the same row are significantly different (p < 0.05) between honeys from different floral origins produced by the same bee species. Different small letters in the same row is significantly different (p < 0.05) between honeys produced by different bee species from the same floral origin.

Table 5. Organic acids concentration (g/kg of honey) of stingless bee honey from different floral origins

Parameters	Geniotrigona thoracica			Apis mellifera	Heterotrigona itama			Apis mellifera
	Acacia	Starfruit	Gelam		Acacia	Starfruit	Gelam
Gluconic	0.48^Cb ± 0.005	<LOQ	0.55^Bb ± 0.02	0.68^A ± 0.03	0.90^Ba ± 0.02	0.39^Ca ± 0.08	1.48^Aa ± 0.22	0.68^BC ± 0.03
Tartaric	0.04^Ba ± 0.007	ND	<LOD	0.15^A ± 0.01	0.06^Ba ± 0.006	0.03^Ca ± 0.01	<LOD	0.15^A ± 0.00
Formic	ND	ND	ND	ND	<LOD	ND	ND	ND
D-malic	ND	ND	ND	0.48^A ± 0.03	0.03^B ± 0.01	ND	0.03^B ± 0.01	0.48^A ± 0.03
Lactic	0.20^Aa ± 0.08	0.03^Ba ± 0.01	0.17^Aa ± 0.05	ND	0.15^Aa ± 0.05	0.18^Aa ± 0.01	0.18^Aa ± 0.02	ND
Acetic	0.09^Bb ± 0.002	0.08^Ba ± 0.02	0.06^Ba ± 0.02	0.39 ^A ± 0.03	0.30^Aa ± 0.06	0.06^Ba ± 0.02	0.01^Ba ± 0.001	0.39^A ± 0.03
Citric	0.04^Ba ± 0.002	0.05^ABa ± 0.004	0.03^Bb ± 0.003	0.42^A ± 0.27	0.04^Ba ± 0.002	0.06^ABa ± 0.005	0.09^ABa ± 0.001	0.42^A ± 0.27
Succinic	0.52^Aa ± 0.08	0.07^Bb ± 0.10	ND	ND	0.32^Ab ± 0.03	0.38^Aa ± 0.10	0.34^Aa ± 0.01	ND

ND: Not detected; LOQ: Below limit of quantitation; LOD: Below limit of detection.

Means with a different capital letters in the same row are significantly different (p < 0.05) between honeys from different floral origins produced by the same bee species. Different small letters in the same row is significantly different (p < 0.05) between honeys produced by different bee species from the same floral origin.

Figure 3. The HPLC Chromatogram of (a) organic acid standard mixture; (b) Apis mellifera honey; (c) Acacia honey (HT); (d) Acacia honey (HI); (e) Starfruit honey (HT); (f) Starfruit honey (HI); (g) Gelam honey (HT); (h) Gelam honey (HI). HI = Heterotrigona itama; HT = Geniotrigona thoracica. Peak identification: 1 = Gluconic; 2 = Tartaric; 3 = Formic; 4 = D-malic; 5 = Lactic; 6 = Acetic; 7 = Citric; 8 = Succinic; 9 = Fumaric

Table 6. Total phenolic acid contents (TPC) (mg GAE/100g), total flavonoids content (TFC) (mg QE/100 g), antiradical scavenging activity (IC₅₀) (mg/mL) and ferric reducing antioxidant power (FRAP) (µmol FeSO₄.7H₂O/100 g)

Parameters	Geniotrigona thoracica			Apis mellifera	Heterotrigona itama			Apis mellifera
	Acacia	Starfruit	Gelam		Acacia	Starfruit	Gelam
Total phenolic contents (TPC)	55.86^Aa ± 2.40	39.38^Ba ± 1.33	27.33^Cb ± 0.83	29.15^C ± 2.73	33.12^Cb ± 0.91	40.44^Ba ± 2.15	52.25^Aa ± 0.67	29.15^C ± 2.73
Total flavonoids content (TFC)	5.81^Ba ± 0.20	8.46^Ab ± 0.10	2.38^Cb ± 0.20	1.40^D ± 0.03	2.96^Cb ± 0.20	9.31^Aa ± 0.10	6.53^Ba ± 0.20	1.40^C ± 0.03
Antiradical activity (IC50)	97.24^Aa ± 2.53	85.04^Ab ± 3.04	76.04^Aa ± 0.21	83.28^A ± 8.41	76.27^Bb ± 0.88	105.53^Ab ± 10.78	32.58^Cb ± 3.03	83.28^B ± 8.41
Ferric reducing antioxidant power (FRAP)	53.80^Cb ± 2.69	93.40^Aa ± 0.42	83.10^Bb ± 2.55	47.30^C ± 3.68	62.40^Ca ± 0.42	80.70^Bb ± 3.68	163.80^Aa ± 1.84	47.30^D ± 3.68

Means with a different capital letters in the same row are significantly different (p < 0.05) between honeys from different floral origins produced by the same bee species. Different small letters in the same row is significantly different (p < 0.05) between honeys produced by different bee species from the same floral origin.

Figure 4. The IC₅₀ graph of stingless bee and Apis honeys. HI = Heterotrigona itama; HT = Geniotrigona thoracica; RSA = Radical scavenging activity

Table 7. Pearson’s Correlation coefficients (r) analysis among total phenolic contents, total flavonoids content, antiradical scavenging activity (IC₅₀), ferric reducing antioxidant power (FRAP), colour, ash and HMF content of stingless bee honey

Parameters	TPC	TFC	IC50	FRAP	Ash	5-HMF
Colour L*	−0.144	0.036	0.754*	−0.749*	0.253	−0.583
TPC	1.000	0.581	−0.154	0.385	0.241	0.351
TFC		1.000	0.176	0.408	−0.592	0.619
IC50			1.000	−0.800**	−0.367	−0.174
FRAP				1.000	−0.109	0.583
Ash					1.000	−0.213
5-HMF						1.000

Colour L* = degree of lightness, TPC = Total phenolic contents, TFC = Total flavonoids content, IC₅₀ = antiradical scavenging activity, FRAP = ferric reducing antioxidant power, 5-HMF = 5-hydroxymethylfurfural. (r) near to +1 or −1 indicate strong relationship and near to 0 indicate weak/no relationship.

* Correlation is significant at the level of 0.05; ** correlation is significant at the level of 0.03

Figure 5. Score plot for PC1 versus PC2 (a); loading plot for PC1 versus PC2 (b); biplot for PC1 versus PC2 (c). = Heterotrigona itama (HI); = Geniotrigona thoracica (HT); = Apis mellifera.