KR100405958B1 - Method for separation of pigment from Chitin by-product and crustacean and Coloring agent for Fish and Laying hen using that pigment - Google Patents

Abstract

The present invention relates to a method for separating a pigment from a crustacean or chitin by-product and a colorant composition for fish or laying hens containing the same as a main ingredient, and more specifically, a pigment solution generated by acid / alkali treatment of crustaceans including shrimps and crabs. Or a method for separating pigments which can improve egg yolk and fish color by utilizing wastewater generated during decalcification and deproteinization during chitosan manufacturing process, and a natural colorant having oxidative stability with the pigment as a main component. will be.

The method for separating the dye of the present invention comprises the steps of collecting the waste water discarded in the decalcification process and the deproteinization process during the acid-alkaline solution or chitin manufacturing process generated by the acid-alkali treatment of crustaceans as raw materials, and Mixing the wastewater; adjusting the solution mixed by an electric process to an appropriate pH to float the pigment protein in the form of particles; and precipitating and collecting the pigment protein from the mixed solution which has been subjected to the electrical process.

According to the present invention, natural pigments can be easily separated from the shells of crustaceans, which are unused resources, or by using wastewater which is a seawater environmental pollution source generated during the chitin manufacturing process. It is possible to provide economically.

Description

Method for separation of pigment from Chitin by-product and crustacean and Coloring agent for Fish and Laying hen using that pigment}

The present invention relates to a method for separating a pigment from a crustacean or chitin by-product and a colorant for fish or laying hens, the main ingredient of which is decalcification and decalcification by acid and alkali treatment of crustaceans or during chitosan production. The present invention relates to a method for separating pigments capable of improving egg yolk and fish meat color by using wastewater generated in protein as a raw material of a colorant, and to a natural colorant having an oxidative stability based on the pigment.

Pigments in food make the appearance of the product more prominent and play a decisive role in increasing consumer preference and product completion in the market. The importance of these colors is especially emphasized in livestock, fish (farming), and agricultural products that directly observe and distribute products. The freshness and nutritional value of these products are mainly evaluated by their appearance. Its value is highly appreciated.

The degree of pigmentation in livestock or farmed fish is directly related to the management status, growth and feed factors in the process of breeding or farming in the case of bright yellow-orange pigments. Especially in the case of feed factors, the color of egg yolk, farmed fish, broiler meat, shrimp shell, etc. depends on the content of flavonoids, β-carotene and xanthophylls in the feed, but these natural pigments in nature The content is very small or the rate of migration in the animal is low, so the color of the food desired by consumers is hardly seen. Therefore, most feed companies replenish the feed by adding synthetic pigments to solve these problems, and no pigments are consumed for feed, so there are no domestic pigment-related producers.

This phenomenon is because, in the past, the size of livestock and aquaculture fishery in our country was small and small, so it was managed to improve the productivity rather than the quality of the product. However, in recent years, when products are being upgraded and functionalized, and their size is also being enlarged and commercialized, the demand for these pigments is gradually increasing, and the amount of import is gradually increasing, and this situation is expected to continue to accelerate. Therefore, research on the development of such pigments in the domestic conditions is very urgent situation. In this regard, the patent contents related to the colorant registered up to now include a manufacturing method of artificial coloring agent for meat coloring of salmon using a shellfish pigment (patent application 92-0001015), poultry feed for producing vitamin fortified eggs (patent application 97-0063560) , A method for producing a functional egg containing an astaxanthin pigment (patent application 94-025235), but there is no direct relationship with the present content.

In particular, most of the pigments used as colorants are synthetic carotenoid pigments, but in fact, natural carotenoid pigments are a representative group of natural pigments of sulfur, light, red and purple which are widely distributed in the flora and fauna in nature. .

As such, a huge amount of carotenoids are known to biosynthesize these pigments, such as shells of aquatic organisms, plant leaves, zooplankton, algae, and some fish and poultry. In particular, through 30 years of epidemiological investigations and recent empirical experiments, not only some natural carotenoids were known to be carcinogenic substances, but they also exhibited higher order functionalities such as antioxidant activity, improved reproduction and growth rate of animals, inhibition of disease occurrence, and color improvement of fish meat. As it is known, its availability in various fields is increasing in the future. Therefore, if natural pigments are supplied through the feed of farmed fish or livestock animals instead of existing synthetic pigments, they can not only improve the appearance quality of the product but also additionally improve feed efficiency, fertility, disease occurrence suppression, and meat color improvement of these animals. I think that.

In particular, synthetic pigments, which are currently used by most domestic feed companies, have raised the issue of chemical stability as food additives.In some developed countries, artificial pigments are prohibited from adding artificial synthetic pigments to livestock and fish farming. Research is underway around the world, and in some countries, a synthetic colorant used to color egg yolk to yellow and salmon and trout to pink may deposit on the cornea of a growing child and affect vision. The ban is being considered. Japan currently uses 100% natural colorants, Sweden has banned the use of synthetic colorants since 1991, and the UK has banned the use of artificial colorants since November 1998. According to Article 11 of the Fair Standards (Feeding Law), which was proposed by the Ministry of Agriculture and Forestry and the Korean Feed Association in 1999, colorants should not be used except for animal feeds, and astaxanthin should only be used for bream, salmon and trout. The content in blended feed is limited to 100g or less per ton. However, the amount of pigments actually used in trout farms is about 300g (300ppm) per ton, so the remaining shortage should be replaced with natural pigments. In addition, since most of the dyes currently used are sold at high prices as synthetic dyes, it is thought that more research is needed to find and use economical natural color materials at the present time. In addition to the physiological functions of vitamin A precursors, egg yolk and flesh and epidermal pigmentation, carotenoids have other functional advantages over synthetic pigments, as well as physiological activities such as lipid oxidation prevention, hormonal function enhancement, and environmental adaptability. It is known.

The inventors found out that the wastewater discarded during the acid / alkali treatment or chitin preparation of crustaceans is rich in carotenoid pigments liberated from crustaceans or red crabs before the application of the invention.

For example, as a result of performing a pigment extraction experiment by collecting wastewater for each treatment process at the Chitosan manufacturing plant in Yeongdeok, it was shown that red pigment is discharged largely from the washing water once or twice out of eight washing processes after deproteinization. With 25 tons of fresh water, 2,400 tons are produced annually in one chitosan manufacturing plant, and the total amount of wastewater generated in Korea is estimated to be at least 24,000 tons.

In light of the above, the present inventors propose an economical method for producing natural carotenoid natural pigments by separating and purifying natural pigments from wastewater generated during the chitin manufacturing process without pretreatment of raw materials for pigment extraction. According to the conditions, we propose a method to produce natural carotenoid pigment through acid and alkali treatment directly to crustaceans including red crab and shrimp in the same process.

Accordingly, an object of the present invention is to extract pigment directly from crustaceans including shrimp, crab, etc., or to efficiently separate carotenoid pigments from wastewater generated during chitin manufacturing process, and oxidative stability of the separated pigments. It is to provide a method for producing a feed fish for coloring fish and laying hens.

Figure 1 is a process for separating the pigment from the shellfish or chitosan by-products of the present invention.

Figure 2 is a TLC result of the extracted pigment in the red crab shell and chitosan production by-products.

Solvent developed: Ethylacetate: Dichloromethane = 1: 4

Rf of each band (b)

Chitosan by-product (A): b1- 0.24, b2-0.46, b3-0.57, b4-0.90 b5-0.96

Red crab (B): b1-0.40, b2-0.52, b3-0.63, b4-0.73, b5-0.88, b6-0.94

-Pigment material estimated by each band

A (b2) and B (b1): β-carotene, A (b3) and B (b2): Lutein,

B (b3): zeaxanthin, A (b4) and B (b5): Astaxanthin

A (b5) and B (b6): Diaoxanthin

FIG. 3 is an HPLC chromatogram of total carotenoids in red crab shells and chitosan preparation byproducts.

4 is a comparison result of the rainbow trout growth rate by the pigment addition.

The present invention is a method for separating the raw material of the natural coloring agent added to the feed of livestock or fish by using the crustacean or chitin by-products, waste water (decalcium) generated during the acid treatment process on the red crab (or other shellfish) during the chitin manufacturing process Collecting the wastewater discarded in the step of sintering) and alkali treatment (deproteinization), mixing the wastewater, and adjusting the solution mixed by the electric process to an appropriate pH to float the pigment protein in the form of particles. And the step of precipitating and collecting the pigment protein from the mixed solution that has undergone the electrical process.

In the acid treatment (decalcification) process is usually a step of adding a 2-3% 40% hydrochloric acid solution (pH1-3) and reacting at 95 ℃ or more for 10 hours or more, alkali treatment process (deproteinization) The process) includes adding 2 to 3 times of 40% NaOH to the red sea crab that has been subjected to the acid treatment and reacting at 95 ° C. for 10 hours or more, whereby the pigments bound to the protein are reacted with a strong alkali solution. It is separated and removed with the wash water during the washing process. In general, carotenoid pigments are fat-soluble pigments, which are relatively stable to heat, and are mainly present as carotenoid-protein complexes, which are protein complexes. These states are more stable than the free state but highly unsaturated in structure. Therefore, since the complex is known to be unstable to oxygen, light, strong acid or strong heat, the red pigment present in the strong alkaline liquid generated during the alkali treatment of shellfish or the deproteinization process of the chitin manufacturing process is present in a relatively stable state. In particular, when the pH of the alkaline (deprotein) solution is about 13.6 on average, the pigment component dissolved in the solution is somewhat saponified by the strong alkali solution, so that it is possible to obtain a pigment having high purity according to the separation conditions.

In another aspect, the present invention discloses a colorant composition for fish or laying hens containing a suitable amount of excipients as a main component of the pigment separated by the above method.

Hereinafter, the separation process of the dye protein of the present invention will be described in more detail.

Figure 1 shows the separation process of the pigment protein of the present invention, first the wastewater (generally in the range of about pH 2.5 to 3.5) and crustaceans discarded during the decalcification process during the acid treatment solution or chitin production process of shellfish Collecting (2) the wastewater (generally in the range of pH 12.5-14) discarded during the treatment solution or deproteinization process, and mixing (2) the wastewater (pH 8-10) with each other. (3) adjusting the pH of the solution mixed to float the pigment protein in the form of particles in an appropriate range, and precipitating and collecting the pigment protein from the solution which has passed through step (3). ).

The proper pH of the mixed solution for floating the dye protein of step (3) in the form of particles is preferably set to pH 4 or less, which is the isoelectric point of the protein, more preferably in the range of pH 2-4.

The method of precipitating the dye protein of step (4) does not require any particular limitation, but the method of collecting only the precipitated pigment using a continuous decanter after being left for 3 to 24 hours, preferably 3 to 24 hours, Alternatively, the method of performing precipitation using 0.05 to 0.5% by weight of silicic acid or silicate based on the mixed solution in the solution passed through step (3) is suitable for the practice of the present invention. The silicic acid or silicate is a porous material that serves to adsorb the dye protein suspended in the solution in the porous portion to make it easy to precipitate. In this case, it is preferable to leave the silicic acid or silicate in the solution for 2 to 24 hours for adsorption.

The present invention also includes a method of further comprising an extraction purification step (5) for purity control in the precipitate obtained in the above process.

In this case, an organic extraction solvent is used for extraction and purification, and preferably chloroform using a small amount of dichloromethane solution or acetic acid is used.

In addition, the present invention includes a colorant composition for a fish or a laying hen based on a pigment (including a pigment protein) obtained by the above process. Pigment protein separated by the above process (4) can be provided in the colorant composition for fish or laying hens enough by itself, the pigment obtained by further comprising the extraction, purification process of step (5) for purity control It is also preferable in the practice of the present invention that all have as a main component.

The colorant composition according to the above method can be used to enhance the degree of coloring of farmed fish, egg yolk and broiler. Coloring is not a factor that affects the nutritional value of the product, but consumers generally prefer dark yellow color, and also because fish and fish eggs infected with digestive disorders or internal parasites usually do not absorb pigments. Egg yolk is important as a measure of health status for chickens and fish.

The coloration of fish, broiler and egg yolk are made by accumulating xanthophylls belonging to the carotenoid, a yellow pigment, in subcutaneous fat tissue or egg yolk. It is proportional to the health of the laying hens. In particular, in order for the flesh color of farmed salmon to be pink or red like natural salmon, pigments should be added to the feed. If no pigment is added, the value of the product rapidly decreases due to the color fading.

The colorant composition of the present invention includes a colorant composition containing an excipient other than the dye. The excipient may be added various sugars including gelatin and glucose, and antioxidants include α-tocopherol, BHT, BHA, Etoxqunone and the like.

The excipient preferably adds 80 to 99 wt% of various sugars including glucose and 0.5 to 5 wt% of gelatin, and 0.01 to 0.1 wt% of the antioxidant based on 100 wt% of the pigment.

The coloring agent composition of the present invention serves to enhance the body color of the yellowed egg yolk and cultured fish as a color source of cultured fish and laying hens by feeding a proper amount to the feed composition for farmed fish and laying hens. In addition, the pigment material contained in the colorant composition of the present invention is very suitable as a feed additive because it is rich in inorganic mineral elements such as calcium and phosphorus as well as carotenoid pigments as an active ingredient.

Hereinafter, the content of the present invention will be described in detail through examples. However, these examples are only for explaining the present invention, the scope of the present invention is not limited to these examples.

Example 1 Composition Analysis of Pigment Extracted from Chitin by-Products

In order to examine the pigment composition extracted from the chitin by-products, five bands were observed when the extracted and purified pigments were developed with a solvent of ethyl acetate: dichloromethane = 1: 4 on TLC coated with silica gel-paper (FIG. 2). HPLC analysis showed that 13 peaks (FIG. 3) were detected. Six bands could be detected in the pigment composition of chitin, the raw material of chitin, but only five bands were detected in the pigment composition of chitin by-products. Was observed.

<Example 2> Pigment separation experiment according to the pH change of the chitin by-product solution.

In this experiment, we used chitin by-product as waste colorant, that is, wastewater generated during the washing process after decalcification. The amount of dye used was about 0.02mg / 100ml, depending on the composition of the raw material.

Since the carotenoids present in the chitin by-product solution have polarity in the form of binding to the protein, the change of the chitin by-product (solution) was adjusted to examine the change, and the results are shown in Table 1. As a result, as the pH was lowered, a reddish substance was precipitated and the color of the supernatant liquid gradually became pale.As a result of measuring the chromaticity of the supernatant with a color difference meter, the a value (red value) decreased rapidly and b value (yellow value). The change of showed a gentle change. Particularly, the value of a was rapidly changed based on pH 4, and the amount of precipitate was the maximum based on pH 4 and 3, which is near the isoelectric point of protein, and decreased with increasing pH. It is estimated that the red pigment substance dissolved in the chitin by-product solution forms a complex with the protein.

<Table 1> Changes in chromaticity and precipitation of the filtrate by pH concentration

pH Color Characteristic Sedimentation amount (%) L a b △ E One 87.9 -2.59 22.1 25.3 Pale yellow transparent liquid 0.043 2 87.4 -2.92 20.9 24.6 0.043 3 83.7 -1.28 20.7 26.3 0.043 4 71.6 3.77 21.7 35.9 Lower pH fades red 0.031 5 57.3 16.4 25.1 52.1 0.031 6 61.0 14.9 27.3 49.8 0.031 7 64.3 13.4 28.9 47.8 0.031 8 64.9 15.1 30.5 48.8 0.013 9 66.0 15.7 32.0 49.2 0.009 10 66.7 15.7 32.8 49.3 Dark red - ^* 11 64.4 16.8 32.7 51.1 - 12 64.4 16.8 32.7 51.1 - 13 70.3 12.9 34.3 47.1 - Stock solution (13.6) 71.9 15.8 40.4 51.7 -

* Not detected

Example 3 Precipitation Experiment of Pigment in Chitin by-products

Pigment protein obtained by adjusting the pH of chitin by-products, ie, pigment-containing precipitates, required a high centrifugal force of more than 3,000 rpm even in the laboratory stage to collect in a suspended state with loosely cross-linked particles. When using a common filtration device, the bonds between the particles were broken and returned to the original red solution, that is, the pigment stock solution, and even when the membrane filter was used, it was recovered, but it required a lot of time and cost to process a large amount of the stock solution. This was required. In particular, the chitin by-products were adjusted to pH 3 and left to stand at room temperature for 3 hours or more, indicating that most of the pigments were precipitated. It showed a characteristic to go back.

Therefore, a strong method of inducing and binding bonds between pigment particles is to add casein as a protein seed, to extract protein, and to add ethanol and methanol and silicic acid to induce binding between particles in a porous form. The precipitation time and recovery degree between the precipitation methods by the method of adding silica) are shown in Table 2 below.

<Table 2> Effect of precipitants on chitosan by-product (pH3)

precipitant Settling time State of the supernatant Chitosan byproduct (after pH adjustment) Casein ^* Ethanol ^** Methanol ^** Silica ^* 24 hours <24 hours <18 hours <14 hours <30 minutes-2 hours Clear Yellow Solution Clear Yellow Solution Red Solution Red Solution Clear Yellow Solution

* About 0.01g <for 200ml of chitosan by-product

** Equivalently add 99% purity to 200ml of chitosan by-product

When the chitosan by-product (solution) was added to silica, ethanol, and methanol as a precipitant and allowed to stand for a predetermined time, the pigment protein suspended in the solution precipitated, resulting in a clear yellow or a slightly reddish supernatant. At this time, the precipitated pigment material could be collected by simple filtration, and because the chitin by-products were bound more tightly than the pigment material obtained by standing for 3 hours or more, the precipitated state showed a tendency to be maintained even in a simple filtration process.

Casein was found to have no precipitation effect in each pretreatment. In case of ethanol and methanol, the pigment was not completely precipitated, resulting in the supernatant reddish and precipitating only sugars or proteins in chitin by-products. In general, the smaller the amount of ethanol added, the more difficult it was to extract pigments and white precipitates were formed. The higher the amount of ethanol, the better the recovery of pigments. The extraction of pigments by ethanol treatment is considered difficult to use in reality.

When silicic acid (silica) was added, the pigment protein in the chitosan by-products precipitated rapidly (about 30 minutes to 2 hours), and all of the suspended pigment protein mass precipitated. Silicic acid is an inert substance used as an excipient in most animal feed and pharmaceutical products as a calcium fortifying agent, so it is not a problem even when added during the pigment manufacturing process.

Example 4 Collection of Pigments, Characteristics of Dry Pigments and Purification Methods

The precipitate (pigment) obtained in Example 2 was washed with distilled water 2-3 times, collected by filtration, and then added with acetone or dichloromethane of 1/10 to 2/10 to the precipitate (pigment). After drying within 30 minutes to 2 hours and examining the general components, it was confirmed that the total carotenoid pigment content is a composite material of about 5-8%. The precipitate was insoluble in water having a pH of 4 to 5, and the pH was adjusted to alkali with NaOH. As a result, the precipitate became dark red in aqueous solution. As it was neutralized, the pigments that were insoluble in the aqueous solution were presumed to have an increasingly darker color, and after adjusting to pH 7, the salinity was measured to be about 3%.

While the carotenoid pigment content, which is a fat-soluble pigment in the sediment, was 5-8%, the fat content was 14.37%, which is about twice that of the fat, and the protein and carbohydrate contents were 21.51% and 45.09%, respectively. ) Was also available as a feed additive formulation (Table 3).

<Table 3> Composition and minor components of the pigment extracted from the chitin produced by-products ^*

division content General ingredient (%) Moisture crude protein Crude ash NFE ^** 1.5321.5114.3717.5045.09 Mineral CaPMgKClSMnFeCuZnCdPb 0.09% 0.34% 0.02% 0.09% 14.80% 0.36% 9ppm1797ppm179ppm16ppm Not detected Not detected amino acid(%) LYSCYSTHRVARHISILEARG 0.721.47-1.480.270.51- Total Carotenoids (mg / 100g) 6,848

* Slight fluctuation depending on raw material condition

** Nitrogen Free Extract

The pigment material collected by adjusting the pH to 3 was collected in the sludge state having a large amount of water, and thus the extraction effect of each organic solvent was examined for purification. As a result, 10 times more ether, petroleum ether, nucleic acid, and chloroform were added to the pigment material and mixed. As a result, the color of the extraction solvent tended to become red as time passed (about 24 hours). It appeared to be extracted to some extent. On the other hand, chloroform and dichloromethane added with a small amount of acetic acid were mixed with the pigment material, and the color of the extraction solvent was dark red. It was observed that most pigments were extracted and released (Table 4).

<Table 4> Dissolution characteristics of precipitates (pigments) extracted from chitin by-products

Solvent ^1,2) Dissolution property ³⁾ Color L a b △ E Chitosan by-product (liquid, before pH adjustment) EtherPetroleum EtherHexanChloroformChloroform + acetic acid (small amount) -Not extracted ⁴⁾ Not extracted ⁴⁾ Not extracted ⁴⁾ Not extracted ⁴⁾ Extracted 71.976.871.671.575.166.066.7 15.84.03.83.84.315.715.7 40.430.521.720.630.332.032.8 51.738.335.934.239.349.249.3

1) 10 times addition to precipitate

2) more than 99% purity

3) After adding solvent → 2 ~ 3 times mixing → Politics → Supernatant color measurement

4) 24 hours <Extraction

Example 5 Coloring Effect of Extracted Pigment (Rainbow Trout)

In general, the coloration effect of trout and salmon is astaxanthin, and the commercial color should be used as a caro pill pink (Roche), but the caro pill is more effective than commercial caro pill pink. Red (Roche) is used. Therefore, the comparative treatment of this experiment also used the carophyll red, the main component of this pigment is known as Canxanthin (Canthxanthin). The result of comparing the rainbow trout growth rate by the extraction pigment with the commercial pigment is shown in FIG. 4, and the results of the body color and flesh color of the fish bred using the color difference meter are shown in Table 5.

The growth rate of the extracted pigment-added group was higher than that of the no-added group, which is presumed to be the result of natural pigments containing some fat-soluble vitamins such as vitamins A, E, and D or antioxidants. In particular, the commercial pigment addition group, which is a synthetic pigment, exhibited a lower growth rate than the non-added group, and it was thought that the growth inhibition effect by the synthetic pigment was also to some extent.

Visual observations over the entire breeding period showed that the body color began to redden mainly in the lateral line, pectoral fin and hind fin. At this time, the body color was visually confirmed from the first month compared to the control group, and in the case of meat color, there was no difference between treatments even one month after feeding the experimental feed.

<Table 5> Coloring Result of Rainbow Trout

Classification 1 week 4 Weeks 8 Weeks l a b △ E l a b △ E l a b △ E Control 47.2 4.03 11.4 48.7 47.2 4.03 11.4 48.7 37.72 -0.472 10.7 58.4 Commercial dye group 47.2 4.03 11.4 48.7 35.8 2.27 5.82 36.5 44.4 5.17 12.3 57.2 Extractive pigment addition group 47.2 4.03 11.4 48.7 44.0 4.60 11.04 49.2 42.3 3.81 12.1 59.1 Extraction Color + Commercial Color 47.2 4.03 11.4 48.7 45.1 5.25 11.63 48.3 38.7 2.92 10.4 62.3

In general, for the coloring of rainbow trout in the fish farms, it is known that the coloration effect is greatest when the feed containing the pigments is added to adults of 500 to 600 g or more 8 weeks before shipping. In this experiment, 30g of fry were tested and their coloration was low until 8 weeks. Initially, the pigmentation of the extracted pigment group was higher than that of the commercial pigment group (Carophyll red). The a value (Red value) of the extract group was also higher than that of the commercial group and the control group. However, as the feeding period elapsed, the addition of extract pigments tended to have a higher degree of coloration than the commercial pigment addition group (Carophyll red), whereas the commercial synthetic pigments were canthxathin, whereas the extract pigments were β-carotene and The result is because it is a complex pigment composed of 8-9 carotenoids including taraxanthin.

Overall, the weight of 50g to 300g showed a tendency of low color and flesh color effect in all pigmented groups. In particular, the pigmentation effect of the extracted pigments tended to be lower than those of commercially available pigments, making it difficult to measure them visually and mechanically. The shape of rainbow trout also showed different trends depending on the pigment additions. The commercial pigment additions retained the long and thin form of the fish, while the extraction pigment additions were round and the upper and lower widths were similar. The weight of the bulb tended to be about 5% higher than the non-added group, and about 10% higher than the commercial pigmented group. Since the extracted pigment is a natural pigment, it is estimated to be a result of containing some fat-soluble vitamins such as vitamins A, E, and D or antioxidants, but the growth rate of the extract pigment was more than 800ppm, indicating a slow growth rate. It was thought that trout caused a drastic reduction in feed intake due to the distinctive odor of extracted pigments. Table 6 shows the palatability and visual depiction of the meat color differences between rainbow trout after 12 weeks.

<Table 6> Preference and Visual Description of Rainbow Trout Meat 12 Weeks After Pigmentation

Treatment Control Commercial Colors Extraction Pigment Commercial + Extract Symbol diagram ¹⁾ 3.1 4..2 4.6 4.3 Visual description Strong yellow due to corn protein (jade gluten) added to the feed Yellowish yellow due to corn protein (jade gluten) added to the feed but slightly reddish Reddish yellowish, clear yellow meat of corn protein (jade gluten) added to feed Yellow and red color of corn protein (jade gluten) added to feed

1) 5-point scoring method

During the entire experimental period, the pigmentation effect was observed by adding the extracted pigment to the commercial feed fish feed, and it was found that jade gluten (corn embryo) added as a protein source in the commercial feed decreased the coloring effect of the pigment. Therefore, the use of protein additives other than corn gluten may be advantageous for the coloration of trout when the feed is prepared by the addition of extract pigments. In particular, the color of the extract pigments showed a tendency to be different from the other three treatments, which had a weak yellow color and a transparent red color, and showed a strong yellow color. The following is the result of the coloring effect of rainbow trout fry weighing about 100g with different concentrations of extracted pigments.

<Table 7> Changes in Meat Color of Rainbow Trout by Feeding with Different Pigments

Treatment District Chromaticity (2 weeks) Chromaticity (4 weeks) Symbol diagram ¹⁾ L ²⁾ a ³⁾ b ⁴⁾ Symbol L a b General fish farming group 1.5 48.12 3.90 3.42 3.1 44.54 4.06 2.80 Commercial pigment group (300ppm) ⁴⁾ 2.1 41.61 5.06 2.29 4.5 42.19 4.98 5.77 Commercial Colors + Extraction Colors ⁵⁾ 2.0 43.80 4.53 3.35 3.8 41.88 4.04 5.88 Extractive Pigment Group (300ppm) 2.2 39.40 6.43 5.69 4.2 42.60 3.55 2.88 Extractive Pigment Group (600ppm) 2.2 39.71 7.80 3.24 5.1 42.00 4.80 4.86 Extractive Pigment Group (900ppm) 2.4 40.52 6.65 3.44 5.6 44.68 4.95 6.10

¹⁾ 9 point scoring method, red intensity, ²⁾ L (lightness), ³⁾ a (redness / greenness), ⁴⁾ b (yellowness / blueness), ⁵⁾ Carophyll Red, ⁶⁾ Carophyll Red 150ppm + extract pigment 150 ppm

Visual observations of the body color changes of rainbow trout over the entire breeding period showed that the body color began to be colored mainly from the lateral line, pectoral fin and hind fin, and flesh color became red from the tail. In the second week of the experiment, the color difference between the control and the addition was slightly visible from the tail. In the results of color difference analysis, there was a clear difference between the treatments, resulting in decreasing lightness and increasing a (redness / green-ess), but not b (yellowness / blueness). From the fourth week of the experimental feed, the color was clearly visible to the naked eye, but the difference between the treatments was not different from the treatments in the color difference.

Example 6 Coloring Effect of Extracted Pigment (Egg yolk)

In order to investigate the effect of egg yolk coloring by adding red crab, oolongseng, and extract pigments, the pigmentation effect was confirmed by adding extract pigments to commercial laying hens. The amount of addition was about 5ppm and 3ppm of extracted pigments and commercially available pigments, respectively. The number of laying hens in each treatment group was used as the experimental group, and the experimental feed for each treatment was divided into two doses of 110g per day. The scattering rate and egg weight during the whole experiment are shown in Table 8.

<Table 8> Scattering Performance by Types of Pigment Additives

Treatment Spawn rate I'm in Control 90.55 ± 6.69 52.21 ± 1.67 Commercial Colors 92.85 ± 6.93 51.74 ± 1.44 Extraction Pigment 92.96 ± 7.97 51.85 ± 2.02

There was no statistically significant difference in egg production rate and egg weight in the commercial pigments, red crab, green crab, extract, and general treatment sections, so the addition of these pigment additives had no effect on the scattering performance. It was estimated to be large.

The pigmentation degree of the yolk by the experimental period of the commercial color, the extracted color, and the general treatment is shown in Table 9. Overall, the degree of coloring of the added group was higher than that of the non-added group, indicating that there was a coloring effect. However, since the extracted pigment is a complex carotenoid pigment, the color of the red color is estimated to be slightly lower than that of the commercial pigment.

<Table 9> Coloring effect of egg yolk according to the type of pigment additive (Roche color fan score)

Treatment 3 days 1 week 2 weeks 3 weeks Control 7.4 ± 0.9 7.9 ± 1.1 7.4 ± 0.8 7.3 ± 0.6 Commercial Colors 10.3 ± 1.2 11.4 ± 0.9 11.6 ± 0.6 11.7 ± 0.8 Extraction Pigment 10.3 ± 1.3 11.1 ± 0.8 11.0 ± 0.7 11.1 ± 0.8

In general, egg yolk is measured by the Roche color fan score. Egg yolks with no pigment are 6-7, and egg yolk of the most preferred egg has a Roche color fan score of 11-13. As a result of this experiment, commercial and extracted pigments showed Roche color fan score of 11-12, showing slightly lower fan score than target 11-13, which can be modified by adjusting the amount of colorant added. All treatments except no addition showed similar coloring effect on egg yolk after 7 days of the experiment.

Roche color fan score is evaluated by the visual ability of the sensory sources, so the results of measuring the yolk after 3 weeks by mechanical (color difference meter) are shown in Table 10. As a result of the Roche color fan score, the coloring effect of the added group was different from the non-added group. In particular, commercially available pigments tended to have high a (red), low L (light) and high △ E (browness) values, whereas extracted pigments tended to have high b value (yellow). Was high and ΔE was low. These results are thought to be the result of the extraction of the complex carotenoid pigments because of the high content of β-carotene with strong yellow color in the composition.

<Table 10> Result of coloring experiment of eggs (3 weeks)

Treatment Average weight (g) 3 days later L a b △ E Roche color fan score Control 52.21 52.57 11.60 26.27 49.16 7.3 Commercial Colors 51.74 51.66 16.75 26.30 51.35 11.7 Extraction Pigment 53.20 53.20 11.69 28.21 49.62 11.0

According to the present invention, natural pigments can be easily separated using shells of shellfish, which are environmental pollutants, and wastewater, which is a seawater environmental pollutant generated during the chitin manufacturing process. It is possible to provide as.

Claims (8)

In the method for separating the raw material of the natural coloring agent added to the feed of livestock or fish using the shell of shellfish and chitin by-products, Collecting wastewater from the decalcification process of the crustacean acid treatment or chitin manufacturing process, and wastewater discarded from the deproteinization process of the crustacean alkali treatment or chitin manufacturing process; Mixing the wastewaters; Adjusting the pH to 4 or less in order to float the pigmented protein in the solution mixed by the above-described process, Separation of pigments from chitin by-products comprising the step of collecting and collecting the pigment protein in the mixed solution that has undergone the electrical process delete 2. The method of claim 1, wherein the precipitation of the pigment protein in the process is carried out using a continuous decanter after standing for 3 to 24 hours. The method of claim 1, wherein the precipitation of the pigment protein in the process is carried out using 0.05 to 0.5% by weight of silicic acid or silicate. The pigment from any of the by-products of chitin preparation according to any one of claims 1 to 4, further comprising the step of extracting and purifying the precipitate collected through the above step using a predetermined organic extraction solvent. How to separate it. The method of claim 5, wherein the organic extraction solvent is a chloroform or dichloromethane with a small amount of acetic acid. A coloring composition for a fish or a laying hen, characterized in that it comprises a suitable amount of excipients as a main component of the pigment separated by any one of claims 1 to 6. According to claim 7, Coloring composition for fish or laying hens, characterized in that 0.5 to 5% by weight of gelatin, 80 to 99% by weight of glucose, 0.01 to 0.1% by weight of antioxidant is added based on 100% by weight of pigment protein. .