Enhancing quality and safety of raw buffalo meat using the bioactive peptides of pea and red kidney bean under refrigeration conditions

Figures & data

Figure 1. SDS-PAGE of red kidney bean protein isolate (RPI) and Alcalase-red kidney bean hydrolysate (RBAH) generated by Alcalase after hydrolysis time (0, 15, 30 and 60 min), M: Tris glycine Marker.

Figure 2. SDS-PAGE of protein isolate (PI) and 11S pea globulin (11SGP). (M: Tris glycine Marker; AS: acidic subunit; BS: basic subunit).

Figure 3. The pH-protein solubility curve of Alcalase-red kidney bean hydrolysate (RBAH) and 11S pea globulin (11SGP).

Figure 4. Development of red kidney bean protein isolate (RPI) hydrolysis with time at 37 °C and pH 6.

Figure 5. 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity of Alcalase-red kidney bean hydrolysate (RBAH) and 11S pea globulin (11SGP) concentration as compared to tertiary butylhydroquinone (TBHQ). Data are presented means ± SE, different lowercase and uppercase letters indicate significant differences p ≤ .05.

Figure 6. Diameter of inhibition zones (DIZ) in tested G + and G − bacteria, i.e. 1) Bacillus cereus; 2) Staphylococcus pyogenes; 3) Escherichia coli; 4) Acinetobacter baumannii affected by Alcalase-red kidney bean hydrolysate (RBAH) and 11S pea globulin (11SGP), A-F, Alcalase-red kidney bean hydrolysate (RBAH) and 11S pea globulin (11SGP) concentration (25, 50, 100, 200, 400 and 800 µg/mL).

Table 1. Antimicrobial activity of red kidney bean alcalase hydrolysate (RABH) and 11S pea globulin (11SGP) (n = 5).

Microorganism		RBAH						11SGP							MIC
	25	50	100	200	400	800	Mean	25	50	100	200	400	800	Mean	RBAH	11SGP
Bacteria
Bacillus cereus	0	0	21^d	25^cd	30^b	37^a	18.83^A	0	0	0	35^c	39^b	43^a	23.4^A	80^d	120^e
listeria monocytogenes	0	0	20^d	25^cd	28^bc	36^ab	18.16^A	0	0	0	31^d	32^cd	39^b	20.4^AB	75^e	140^d
Streptococcus pyogenes	0	0	19^e	26^c	29^bc	36^ab	18.33^A	0	0	0	30^d	35^c	39^b	20.8^AB	90^d	160^c
Escherichia coli	0	0	0	0	23^d	28^bc	8.50^C	0	0	0	25^e	27^de	30^d	16.4^B	250^b	180^b
Acinetobacter baumannii	0	0	0	0	26^c	30^b	9.33^B	0	0	0	27^de	28^de	32^cd	17.4^B	210^c	145^d
Pseudomonas aeruginosa	0	0	0	0	24^c	26^c	8.33^C	0	0	0	25^e	24^e	29^de	15.6^B	290^a	190^a
Mean	0.0	0.0	10.00^C	12.66^C	26.66^B	32.16^A		0.0	0.0	0.0	28.83^B	30.83^AB	35.33^A		165.83	155.83
ANOVA					p Value							p Value			p Value	p Value
Bacteria (B)					<.001							<.001			<.001	<.001
Concentration (C)					<.001							<.001			<.001	<.001
B × C					<.001							<.001			<.001	<.001
Fungi
Alternaria alternate	0	0	8.0^a	7.0^ab	3.5^c	1.0^e	3.25^A	0	0	7.0^ab	6.5^b	2.5^c	0.5^e	2.75^B	80^a	60^b
Aspergillus flavus	0	0	7.0^ab	6.7^b	3.0^c	0.5^e	2.86^B	0	0	7.0^ab	6.5^b	3.9^c	1.5^d	3.15^AB	55^b	55^b
Fusarium oxysporum	0	0	8.0^a	6.5^b	3.1^c	1.5^d	3.18^A	0	0	7.9^a	6.8^b	3.2^c	1.6^d	3.25^A	70^a	80^a
Monascus purpureus	0	0	7.5^ab	7.2^ab	2.9^c	1.8^d	3.23^A	0	0	7.8^a	7.2^ab	2.9^c	1.8^d	3.28^A	60^b	70^a
Mean	0.0	0.0	7.63^A	6.85^B	3.13^C	1.20^D		0.0	0.0	7.43^A	6.75^B	3.13^C	1.35^D		66.25	66.25
ANOVA					p Value							p Value			p Value	p Value
Fungi (F)					<.001							<.001			<.001	<.001
Concentration (C)					<.001							<.001			<.001	<.001
F × C					<.001							<.001			<.001	<.001

Means followed by different letters differ significantly by LSD (p ≤ .05).

Bold and italic uppercase letters belong to bacteria or fungi.

Bold and not italic uppercase letters belong to the concentrations of RBAH or 11SGP.

Not bold and not italic lowercase letters belong to the interaction effect.

Figure 7. The effect of Alcalase-red kidney bean hydrolysate (RBAH) (B) and 11S pea globulin (11SGP) (A) with levels 800 µg/mL on Alternaria alternate growth.

Figure 8. Growth curve of G + and G − bacteria in the presence of Alcalase-red kidney bean hydrolysate (RBAH) and 11S pea globulin (11SGP) peptides (60 µg/mL) concentration, different lowercase letters indicate significant differences p ≤ .05.

Table 2. Physicochemical changes during raw buffalo meat supplemented with red kidney bean alcalase hydrolysate (RABH and 11SGP) during cold preservation for 15 d (n = 5).

Peptides	Storage period	pH					Met myoglobin (%)					Inhibition in lipid oxidation (%)
		0	100	250	400	Mean	0	100	250	400	Mean	0	100	250	400	Mean
Alcalase-red kidney bean hydrolysate (RBAH)	0	6.00^cd	6.00^d	6.00^d	6.00^d	6.00^C	13.00^k	13.00^k	13.00^k	13.00^k	13.00^D	35.00^a	35.00^a	35.00^a	35.00^a	35.00^A
	5	6.50^abcd	6.33^bcd	6.20^cd	6.10^cd	6.28^CD	40.00^e	32.00^g	25.00^i	21.00^j	29.50^C	25.00^e	29.00^cd	31.00^bc	32.00^b	29.25^B
	10	6.93^ab	6.50^abcd	6.30^bcd	6.20^bcd	6.48^AB	60.00^b	40.00^e	35.00^f	29.00^h	41.00^B	21.00^f	26.00^e	29.00^d	30.00^bc	26.50^C
	15	7.20^a	6.81^abc	6.50^bcd	6.20^bcd	6.67^A	75.33^a	54.00^c	45.00^d	40.00^e	53.58^A	10.00^h	15.00^g	22.00^f	25.00^e	18.00^D
	Mean	6.66^A	6.41^AB	6.25^B	6.12^B		47.08^A	34.75^B	29.50^C	25.75^D		22.75^D	26.25^C	29.25^B	30.05^A
	ANOVA			p Value					p Value					p Value
	Storage (S)			.002					<.001					<.001
	Concentration (C)			.031					<.001					<.001
	S × C			.867					<.001					<.001
11S pea globulin (11SGP)	0	6.00^cd	6.0^d	6.0^d	6.0^d	6.00^C	13.00^j	13^j	13^j	13^j	13.00^D	35.00^a	35^a	35^a	35^a	35.00^A
	5	6.50^abcd	6.3^bcd	6.2^cd	6.1^cd	6.28^CD	40.00^e	29^g	23^h	19^i	27.75^C	25.00^e	30^cd	31^bc	33^b	29.75^B
	10	6.93^ab	6.4^bcd	6.3^bcd	6.2^bcd	6.46^B	60.00^b	37^e	33^f	24^h	38.50^B	21.00^f	25^e	29^d	32^b	26.75^C
	15	7.20^a	6.5^abcd	6.3^bcd	6.3^bcd	6.58^A	75.33^a	50^c	45^d	39^e	52.33^A	10.00^h	17^g	23^f	27^e	19.25^D
	Mean	6.66^A	6.3^AB	6.2^B	6.15^B		47.08^A	32.25^B	28.5^C	23.75^D		22.75^D	26.75^C	29.5^B	31.75^A
	ANOVA			p Value					p Value					p Value
	Storage (S)			.001					<.001					<.001
	Concentration (C)			.033					<.001					<.001
	S × C			.865					<.001					<.001

Means followed by different letters differ significantly by LSD (p ≤ .05).

Bold and italic uppercase letters belong to storage period.

Bold and not italic uppercase letters belong to the concentrations of RBAH or 11SGP.

Not bold and not italic lowercase letters belong to the interaction effect.

Table 3. Total bacterial count (TBC) and psychrophilic bacterial count (PBC) of raw buffalo meat during storage for 15 d at 4 °C (n = 5).

Peptides	Storage period	Total bacterial count (Log CFU)					Psychrophilic bacterial count (Log CFU)
		Concentration					Concentration
	Concentrations (µg/g)	0	100	250	400	Mean	0	100	250	400	Mean
Alcalase-red kidney bean hydrolysate (RBAH)	0	4.00^e	4.00^e	4.00^e	4.00^e	4.00^D	2.50^h	2.50^h	2.50^h	2.50^h	2.50^D
	5	5.80^c	5.50^d	5.20^d	4.80^e	5.32^C	4.00^ef	3.80^fg	3.20^gh	2.90^gh	3.47^C
	10	7.10^b	6.40^c	5.80^cd	5.20^d	6.12^B	6.50^b	5.80^c	5.00^cd	4.80^de	5.52^B
	15	9.50^a	8.00^b	7.3^b	6.00^c	7.70^A	8.40^a	7.00^b	6.80^b	6.80^b	7.25^A
	Mean	6.60^A	5.22^B	5.57^B	5.00^C		5.35^A	4.77^B	4.37^BC	4.25^C
	ANOVA			p Value					p Value
	Storage (S)			<.001					<.001
	Concentration (C)			<.001					<.001
	S × C			<.001					.0044
11S pea globulin (11SGP)	0	4.00^e	4.0^e	4.0^e	4.0^e	4.00^C	2.50^f	2.5^f	2.5^f	2.5^f	2.5^D
	5	5.83^c	4.6^e	4.5^e	4.2^de	4.78^C	4.00^d	2.8^e	2.7^e	2.6^e	3.03^C
	10	7.17^b	4.9^d	4.8^d	4.7^d	5.39^B	6.50^b	4.7^c	4.6^c	4.5^c	5.08^B
	15	9.50^a	7.2^b	6.5^b	5.8^c	7.25^A	8.40^a	6.1^b	6.0^b	5.9^b	6.60^A
	Mean	6.62^A	5.18^B	4.95^C	4.68^C		5.35^A	4.03^B	3.95^BC	3.88^C
	ANOVA			P Value					P Value
	Storage (S)			<.001					<.001
	Concentration (C)			<.001					<.001
	S × C			<.001					.0042

Means followed by different letters differ significantly by LSD (p ≤ .05).

Bold and italic uppercase letters belong to storage period.

Bold and not italic uppercase letters belong to the concentrations of RBAH or 11SGP.

Not bold and not italic lowercase letters belong to the interaction effect.

Table 4. The fluctuations in colour parameters during RBAH and 11SGP-supplemented meat during storage for 15 d at 4 °C. (n = 5).

Peptides	Concentration	L*			a*			b*
		Storage period			Storage period			Storage period
		0	15	Mean	0	15	Mean	0	15	Mean
Alcalase-red kidney bean hydrolysate (RBAH)	0	51.00	52.90	51.95	9.50^b	6.80^c	8.15^B	20.62^d	22.70^c	21.66^C
	100	50.90	51.00	50.95	9.70^b	6.85^c	8.27^AB	20.90^d	21.60^c	21.25^C
	250	52.00	53.50	52.80	9.80^ab	6.90^c	8.35^AB	21.89^c	22.75^b	22.32^B
	400	52.20	53.80	53.00	10.90^a	7.10^c	9.00^A	22.75^b	23.65^a	23.20^A
	Mean	51.53	52.80		9.98^A	6.91^B		21.54^B	22.68^A
	ANOVA		p Value			p Value			p Value
	Storage period (S)		.177			<.001			<.001
	Concentration (C)		.389			.043			<.001
	S × C		.898			.049			.015
	Concentration	0	15	Mean	0	15	Mean	0	15	Mean
11S pea globulin (11SGP)	0	51.00	52.90	51.95^A	9.50^a	6.80^c	8.15^B	20.62	22.70	21.66
	100	50.80	51.96	51.38^B	9.55^a	8.20^b	8.88^A	20.60	21.45	20.72
	250	49.00	49.77	49.38^BC	9.45^a	7.90^b	8.68^A	21.50	22.48	21.99
	400	49.13	50.17	49.65^C	9.30^a	6.20^d	7.75^C	21.45	22.41	21.93
	Mean	49.98	51.20		9.45^A	7.28^B		21.04^B	22.26^A
	ANOVA		p Value			p Value			p Value
	Storage period (S)		.063			<.001			.014
	Concentration (C)		.022			<.001			.406
	S × C		.915			<.001			.719

Means followed by different letters differ significantly by LSD (p ≤ .05).

Bold and italic uppercase letters belong to storage period.

Bold and not italic uppercase letters belong to the concentrations of RBAH or 11SGP.

Not bold and not italic lowercase letters belong to the interaction effect.

L* (white-black), a* (redness-greenness) and b* (yellow-blueness).

Table 5. Sensory properties of RBAH and 11SGP-supplemented buffalo meat during the storage period (n = 5).

Peptides	Concentration	Colour			Odour			Appearance			Overall acceptability
		Storage period			Storage period			Storage period			Storage period
		0	15	Mean	0	15	Mean	0	15	Mean	0	15	Mean
Alcalase-red kidney bean hydrolysate (RBAH)	0	8.00^a	1.50^d	4.75^C	7.00^b	1.00^e	4.00^B	6.50^b	1.00^f	3.75^C	6.70^c	1.20^e	3.95^B
	100	7.20^a	3.00^c	5.10^BC	7.90^ab	4.00^d	5.95^A	7.80^a	3.00^e	5.40^B	7.60^bc	4.30^d	5.95^A
	250	7.50^a	4.00^b	5.75^A	8.00^a	4.50^cd	6.25^A	7.50^a	4.00^d	5.75^B	8.00^a	4.50^d	6.25^A
	400	7.60^a	3.50^bc	5.55^AB	7.80^ab	5.00^c	6.40^A	7.60^a	5.00^c	6.30^A	7.70^ab	4.80^d	6.25^A
	Mean	7.57^A	3.00^B		7.68^A	3.63^B		7.35^A	3.25^B		7.50^A	3.7^B
	ANOVA	p Value			p Value			p Value			p Value
	Storage period (S)	<.001			<.001			<.001			<.001
	Concentration (C)	.009			<.001			<.001			<.001
	S × C	<.001			.001			<.001			<.001
	Concentration	0	15	Mean	0	15	Mean	0	15	Mean	0	15	Mean
11S pea globulin (11SGP)	0	8.00^a	1.50^d	4.75^C	7.00^a	1.00^d	4.00^C	6.50^d	1.00^h	3.75^D	6.73^a	1.17^d	3.95^B
	100	7.10^a	3.50^c	5.30^BC	6.90^a	4.00^c	5.45^B	6.90^c	2.00^g	4.45^C	7.10^a	4.00^c	5.55^A
	250	7.15^a	4.20^bc	5.68^AB	7.20^a	5.00^b	6.10^A	7.50^b	3.00^f	5.25^B	6.80^a	5.00^b	5.90^A
	400	7.30^a	5.00^b	6.15^A	6.80^a	5.10^b	5.95^A	8.00^a	4.00^e	6.00^A	6.90^a	5.20^b	6.05^A
	Mean	7.38^A	3.55^B		6.98^A	3.78^B		7.25^A	2.50^B		6.88^A	3.84^B
	ANOVA	p Value			p Value			p Value			p Value
	Storage period (S)	<.001			<.001			<.001			<.001
	Concentration (C)	.002			<.001			<.001			<.001
	S × C	<.001			<.001			<.001			<.001

Means followed by different letters differ significantly by LSD (p ≤ .05).

Bold and italic uppercase letters belong to storage period.

Bold and not italic uppercase letters belong to the concentrations of RBAH or 11SGP.

Not bold and not italic lowercase letters belong to the interaction effect.