Nanoencapsulation of Jania rubens seaweeds' antioxidants for food applications

Abstract

An additive for food products to extend shelf-life is provided where the additive is nanoparticles having extracted phytochemicals or anti-oxidants from Jania Rubens nano-encapsulated with chitosan-tripolyphosphate. Shelf-life was extended in contrast to the synthetic preservatives which are typically used in the food industry. The cost of the natural preservative is much less than that of the synthetic preservatives. Extension of shelf-life by a natural source is nowadays more desirable by the consumers due to the modem trends of consumption of food with no chemical preservatives.

Description

FIELD OF THE INVENTION

The invention relates to methods for extending shelf-life of food products.

BACKGROUND OF THE INVENTION

Lipid oxidation is a major degradative reaction limiting the shelf-life and deteriorating the quality of lipid containing food products. The oxidative deterioration of food products has a negative impact in the food industry in addition to the generation of potentially toxic products. Consequently, the inclusion of additives to slow down or stop the propagation of oxidation reactions is warranted, especially for prolonged storage durations. The present invention is directed towards technology to extend shelf-life of food products.

SUMMARY OF THE INVENTION

The development of natural antioxidants that can mitigate oxidation reactions in food products is on the rise. Several antioxidants have been developed from natural terrestrial plants, with less emphasis on marine seaweeds. Rancidity is a major degradative reaction limiting the shelf-life and deteriorating the quality of lipid containing food products. In this invention, the inventors focused on Jania Rubens algal extract encapsulated by chitosan-tripolyphosphate in retarding lipid oxidation reactions in vegetable oils as a food model system. Phytochemicals were extracted from the seaweeds' matrices by means of an organic solvent.

The antioxidant efficacy of the algal extract was evaluated by means of many assays. Bioactive compounds were further identified using gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry. To enhance the Jania Rubens' extract's efficacy, the phytochemicals were nanoencapsulated into chitosan-tripolyphosphate using the ionic gelation techniques. The optimum nanoformulation was characterized by transmission electron microscopy. It had a particle size of 161 nm, zeta potential of 31.2 my, polydispersity index of 0.211 and an entrapment efficiency of 99.7%.

An in-vitro phytochemicals' release study of the nanoencapsulated extract versus raw extract was performed by means of the dialysis bag diffusion method. This assay was carried out in two stimulation release media to mimic the intestinal and the gastric conditions. In addition, the ability of the optimum formula to extend the shelf-life of vegetable oils, corn, sunflower, soybean and palm oils, was based on peroxide value and thiobarbituric acid assays. Besides, headspace solid-phase microextraction was applied to detect the oils' volatiles as secondary markers of rancidity. The results revealed that the nanoencapsulated Jania Rubens' extract considerably reduced the rate of formation of the primary and secondary oxidation products in the oils.

In one embodiment the invention can be characterized as an additive for food products to extend shelf-life, where the additive are nanoparticles having extracted phytochemicals or anti-oxidants from Jania Rubens nano-encapsulated with chitosan-tripolyphosphate.

In another embodiment the invention can be characterized as a method of extending shelf-life of food products where the method distinguishes having nanoparticles having extracted phytochemicals or anti-oxidants from Jania Rubens nano-encapsulated with chitosan-tripolyphosphate, and adding the nanoparticles to a food product. An example of food products is corn oil, sunflower oil, soybean oil or palm oil.

Significant advantages are provided. Shelf-life was extended of vegetable oils by means of a natural additive, in contrast to the synthetic preservatives which are typically used in the food industry. The cost of the natural preservative is much less than that of the synthetic preservatives. Besides, the inventors were able to extend the shelf-life by a natural source which is nowadays becoming more desirable by the consumers due to the modern trends of consumption of food with no chemical preservatives.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows according to an exemplary embodiment of the invention a schematic presentation showing the cross-linking between the chitosan polymer and the Algal extract. The schematic presentation on the right shows the chitosan-tripolyphosphate (TPP) shell encapsulating the Jania Rubens algal extract. JCNP stands for Jania Rubens chitosan nanoparticles. 110=chitosan; 120=chitosan amino groups; 130=chitosan functional groups; 140=Jania Rubens algal extract; 150=Mixing step (mixing of the chitosan and algal extract solutions by agitation with a magnetic stirrer); 160=solution of chitosan and Jania Rubens algal extract; 170=sodium tripolyphosphate; 180=Homogenization (by means of a high pressure homogenizer to produce monodisperse small nanoparticles); 190=Chitosan-tripolyphosphate encapsulating the Jania Rubens algal extract.

FIGS. 2A-D show according to an exemplary embodiment of the invention extension of the shelf-life of oils expressed as primary oxidation products (peroxide value) versus time of storage in sunflower (FIG. 2A), corn (FIG. 2B), soybean (FIG. 2C) and palm oil (FIG. 2D), respectively. ‘Orange’ 210—oil with synthetic antioxidant, ‘green’ 220—oil with chitosan nanoparticles encapsulation the Jania Rubens extract, ‘grey’ 230—oil with raw extract, ‘blue’ 240—oil with pristine chitosan nanoparticles, ‘yellow’ 250—oil with no additives.

FIGS. 3A-D show according to an exemplary embodiment of the invention extension of the shelf life of oils expressed as secondary oxidation products (thiobarbituric acid value) versus time of storage in sunflower (FIG. 3A), corn (FIG. 3B), soybean (FIG. 3C) and palm oil (FIG. 3D), respectively. ‘Orange’ 310—oil with synthetic antioxidant, ‘green’ 320—oil with chitosan nanoparticles encapsulation the Jania Rubens extract, ‘grey’ 330—oil with raw extract, blue’ 340—oil with pristine chitosan nanoparticles, ‘yellow’ 350—oil with no additives.

DETAILED DESCRIPTION

Jania Rubens

Jania Rubens is available throughout the different seasons in the shores by e.g. Alexandria, Egypt. Jania Rubens is rich in many vital bioactive compounds including flavonoids, diterpenes, carotenoids, vitamins, fatty acids, tannins, phytol, and many more secondary metabolites. The major classes of polyphenols have been found to be flavonoids and tannins, which are known to process an antioxidant activity. To date, a minute amount of research has been done on this specific seaweed.

Antioxidants Extracted from Jania Rubens

The antioxidant efficacy of the Jania Rubens algal extract has been shown to be high by means of two antioxidants assays, i.e., 2,2-diphenyl-1-picrylhydrazyl, ferric reducing antioxidant power. Total phenolic content and total flavonoid content assays were carried out and both assays showed that the algal extract is rich in polyphenols and flavonoids, which are potent antioxidants. Diverse phytochemicals which possess an antioxidant activity were isolated from Jania Rubens by using gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry. The Jania Rubens' extract revealed abundance of fatty acids, e.g. syringic acid, benzene dicarboxylic acid, myristic acid, palmitelaidic acid, palmitic acid, heptadecanoic acid, dodecanoic acid, oleic acid, stearic acid, octanoic acid, itaconic acid, adipic acid, glycolic acid and arachidic acid, which exhibit an antioxidant activity. In addition to fatty acids, some alcohols were detected that exhibit an antioxidant activity e.g. 1,4-butanediol, diethylene glycol and glycerol. Several nitrogenous compounds were detected most of which exhibit an antioxidant effect including uracil, 1-propionylproline, pyrrolidine, 2-butyl-1-methyl, 3-pyridinol and 3-hydroxypicolinic acid. Detected sugars, which have an antioxidant effect, included galactopyranose, 2-O-glycerol-α-d-galactopyranoside and D-glucose. Other lypophilic metabolites identified with a potential antioxidant effect included phytol and neophytadiene. Table 1 below shows the algal phytochemicals detected by gas chromatography-mass spectrometry. Table 2 below shows the algal phytochemicals detected by liquid chromatography-mass spectrometry.

Examples of phytochemicals or anti-oxidants from Jania Rubens useful to be nano-encapsulated with chitosan-tripolyphosphate are, for example, polyphenols, flavonoids, phytol, neophytadiene, syringic acid, benzene dicarboxylic acid, myristic acid, palmitelaidic acid, palmitic acid, heptadecanoic acid, dodecanoic acid, oleic acid, stearic acid, octanoic acid, itaconic acid, adipic acid, glycolic acid, arachidic acid, 1,4-butanediol, diethylene glycol, glycerol, uracil, 1-propionylproline, pyrrolidine, 2-butyl-1-methyl, 3-pyridinol, 3-hydroxypicolinic acid, galactopyranose, 2-O-glycerol-α-d-galactopyranoside, or D-glucose, or any combination thereof. In one embodiment, polyphenols and flavonoids are recognized as the key phytochemicals or anti-oxidants from Jania Rubens useful to be nano-encapsulated with chitosan-tripolyphosphate. In another embodiment, at least polyphenols and flavonoids are recognized as the phytochemicals or anti-oxidants from Jania Rubens useful to be nano-encapsulated with chitosan-tripolyphosphate.

Encapsulation of Characterized Antioxidants

To further enhance the Jania Rubens' extract's efficacy, the algal extracts (i.e. antioxidants) were nanoencapsulated into chitosan-tripolyphosphate nanoparticles using an ionic gelation technique via high pressure homogenization. The optimum nanoformulation was characterized by scanning electron microscopy and transmission electron microscopy. The nanoformulation had a particle size of 161 nm, zeta potential of 31.2 my, polydispersity index of 0.211 and an entrapment efficiency of 99.7%. FIG. 1 shows a schematic presentation showing the cross-linking between the chitosan polymer and the Algal extract. The schematic presentation on the right shows the chitosan-tripolyphosphate shell encapsulating the Jania Rubens algal extract.

Shelf-Life

The nanoencapsulated Jania Rubens extracted antioxidant were added to oils to extend their shelf-life. The ability of the nanoparticle to extend the shelf-life of vegetable oils, corn, sunflower, soybean, and palm oils, was based on peroxide value and thiobarbituric acid assays. Additionally, headspace solid-phase microextraction was applied to detect the oils' volatiles as secondary markers of rancidity. The results revealed that the nanoencapsulated Jania Rubens' extract considerably reduced the rate of formation of the primary and secondary oxidation products in the oils. In other words, the nanoencapsulated Jania Rubens' extract extended the shelf life of the oils to a big extent, besides that its activity was comparable to that of a widely used synthetic antioxidant butylated hydroxytoluene.

FIGS. 2A-D show the extension of the shelf-life of oils expressed as primary oxidation products (peroxide value) versus time of storage in sunflower (FIG. 2A), corn (FIG. 2B), soybean (FIG. 2C) and palm oil (FIG. 2D), respectively. ‘Orange’ 210—oil with synthetic antioxidant, ‘green’ 220—oil with chitosan nanoparticles encapsulation the Jania Rubens extract, ‘grey’ 230—oil with raw extract, ‘blue’ 240—oil with pristine chitosan nanoparticles, ‘yellow’ 250—oil with no additives.

FIGS. 3A-D show the extension of the shelf life of oils expressed as secondary oxidation products (thiobarbituric acid value) versus time of storage in sunflower (FIG. 3A), corn (FIG. 3B), soybean (FIG. 3C) and palm oil (FIG. 3D), respectively. ‘Orange’ 310—oil with synthetic antioxidant, ‘green’ 320—oil with chitosan nanoparticles encapsulation the Jania Rubens extract, ‘grey’ 330—oil with raw extract, blue’ 340—oil with pristine chitosan nanoparticles, ‘yellow’ 350—oil with no additives.

TABLE 1
The algal phytochemicals detected by gas chromatography-mass spectrometry
	Retention				Molecular
Peak #	time (min)	KI	Area	Compound name	formula
1	7.05	1084.7	1152	Glycolic acid, 2TMS	C2H4O3
2	8.61	1173.2	874	Acetic acid	CH3COOH
3	9.76	1242.2	1306	4-Hydroxybutanoic acid	C4H8O3
4	10.17	1268.8	1424	Octanoic acid, TMS ester	C8H16O2
5	11.57	1360.2	1782	Itaconic acid, 2TMS	C5H6O4
6	13.68	1512.6	897	Adipic acid, 2TMS	C6H10O4
7	14.27	1557.8	692	Cinnamic acid	C9H8O2
8	15.48	1654.6	714	Dodecanoic acid	C12H24O2
9	8.55	1168.8	3084	1,4-Butanediol	C4H10O2
10	9.91	1252.2	1084	Diethylene glycol, 2TMS	C4H10O3
11	10.42	1285.6	7270	Glycerol, 3TMS	C3H8O3
12	7.47	1108.2	1182	3,4,5-Trimethylheptane	C10H22
13	16.02	1699.8	3232	Heptadecane	C17H36
14	15.41	1648.4	961	4-Acetamido-1-phenylpyrazole	C11H11N3O
15	17.72	1849.1	1591	Myristic acid, TMS	C14H28O2
16	18.77	1947.7	1334	Myristic acid, TMS	C14H28O2
17	19.58	2026.7	17204	Palmitelaidic acid-TMS	C18H34O2
18	19.77	2045.9	21646	Palmitic Acid-TMS	C16H32O2
19	20.73	2144.8	452	Heptadecanoic acid, TMS	C17H34O2
20	21.43	2218.9	2416	Oleic Acid-TMS	C18H34O2
21	21.49	2225.3	1744	Oleic Acid-TMS	C18H34O2
22	21.64	2242.8	2637	Stearic acid-TMS	C18H36O2
23	23.37	2441.9	544	Arachidic acid-TMS	C20H40O2
24	10.90	1316.3	1353	Unknown nitrogenous	—
				compound
25	11.33	1344.7	4306	Unknown nitrogenous	—
				compound
26	11.41	1349.8	1069	Uracil, 2TMS	C4H4N2O2
27	12.52	1425	1451	Unknown nitrogenous	—
				compound
28	12.74	1442.1	3221	1-Propionylproline, TMS	C8H13NO3
				derivative
29	13.55	1502.9	1156	Unknown nitrogenous	—
				compound
30	23.76	2491.6	498	Unknown nitrogenous	—
				compound
31	7.60	115.6	962	2-Butyl-1-methylpyrrolidine	C9H19N
32	8.12	1144.6	3372	3-Pyridinol, TMS	C5H5NO
33	8.77	1181.0	1996	3-Hydroxypicolinic acid, 2TMS	C6H5NO3
34	9.82	1246.4	1157	Urea, 2TMS	CH4N2O
35	12.33	1411.2	757	Phloroglucinol, O,O′-	C6H6O3
				bis(trimethylsilyl)
36	14.56	1579.3	1004	Unknown steroid, TMS	—
37	21.95	2276.9	1209	Unknown sterol, TMS	—
38	29.22	3182.4	3529	Cholesterol, TMS	C27H46O
39	19.34	2001.3	559	Galactopyranose, 5TMS	C6H12O6
40	21.88	2269.2	11343	O-Glycerol-α-galactopyranoside	C27H66O8
41	16.32	1726.1	566	Levoglucosan, 3TMS	C6H10O5
42	18.44	1916.8	470	D-Glucose, 6 TMS	C6H12O6
43	17.62	1839.6	670	Neophytadiene	C20H38
44	17.90	1865.2	506	Neophytadiene	C20H38
45	21.03	2176.3	6862	Phytol, TMS derivative	C20H40O

TABLE 2
The algal phytochemicals detected by liquid chromatography-mass spectrometry
Peak	Rt				Molecular	Error
No	(min)	[M − H]−	Metabolite	MSn ions (m/z)	Formula	(ppm)	Class
1.	0.51	200.96	Dihydroxyphenyl	183, 157, 110,	C9H11O5—	3.76	Phenolics
			glycerol	89
2.	0.51	272.96	Dihydroxycoumarin	255, 237, 228,	C9H5O8S—	5.78	Coumarin
			sulfate	214, 200, 187
3.	0.58	197.81	Syringic acid	170, 168, 153,	C9H9O5—	2.76	Phenolics
				135
4.	10.54	187.10	Laminine	169, 160, 142,	C9H19N2O2—	0.57	Betaine
				125
5.	11.12	165.95	Benzenedicarboxylic	133, 122	C8H5O4—	3.15	Aromatic
			acid				acid
6.	11.86	277.91	Syringic acid sulfate	197,165, 137	C9H9O8S—	0.26	Phenolics
7.	13.38	242.18	Pentadecanoic acid	225, 198, 181	C15H29O2—	−1.63	Fatty acid
8.	14.89	323.22	Hydroxyeicosadienoic	305, 279, 197,	C20H35O3—	2.81	Fatty acid
			acid	183
9.	16.57	265.15	Heptadecadienoic acid	239, 221, 98	C17H29O2—	1.92	Fatty acid
10.	17.04	297.15	Nonadecanoic acid	279, 253, 197,	C19H37O2—	0.62	Fatty acid
				183
11.	17.53	311.17	Arachidic acid	293, 267, 197,	C20H39O2—	1.73	Fatty acid
				183
12.	18.61	325.18	Arachidic acid methyl	296, 267,	C21H41O2—	2.04	Fatty acid
			ester	225, 197, 183
13.	19.25	339.20	Arachidic acid ethyl	311, 295, 239,	C22H43O2—	−0.90	Fatty acid
			ester	183

Claims

1. An additive for food products to extend shelf-life, comprising: nanoparticles having extracted phytochemicals or anti-oxidants from Jania Rubens nano-encapsulated with chitosan-tripolyphosphate.

2. The additive as set forth in claim 1, wherein the phytochemicals or anti-oxidants are polyphenols and flavonoids.

3. A method of extending shelf-life of food products, comprising: (a) having nanoparticles having extracted phytochemicals or anti-oxidants from Jania Rubens nano-encapsulated with chitosan-tripolyphosphate; and(b) adding the nanoparticles to a food product.

4. The method as set forth in claim 1, wherein the food products are corn oil, sunflower oil, soybean oil or palm oil.

5. The method as set forth in claim 1, wherein the phytochemicals or anti-oxidants are polyphenols and flavonoids.