KR20200056035A - Composition for preservation of a source of livestock feed comprising metabisulfite and preparation method of livestock feed using the same - Google Patents

Abstract

The present invention relates to a composition for the preservation of a feed raw material including at least one selected from metabisulfite and organic acids, and a feed manufacturing method using the same. More specifically, the present invention relates to novel uses of metabisulfite and organic acids capable of preserving fruit and vegetable discards (FVD) used as a feed raw material for a long time, and to a silage manufacturing method using the same. According to a result for processing the at least one selected from the metabisulfite and the organic acids in the FVD, microbiological stability and freshness of the FVD are maintained longer in an aerobic environment, and preservation of nutrients and an antioxidant function is very effective. A silage feed manufacturing method including a preprocessing step using at least one selected from the metabisulfite and the organic acids in the aerobic environment can improve a preservation feature for an anaerobic preservation step. Specifically, co-ensiling of the FVD used with almond shells and corn gluten has exhibited the best preservation feature.

Description

Composition for preservation of a source of livestock feed comprising metabisulfite and preparation method of livestock feed using the same}

The present invention relates to a composition for preservation of feedstock containing metabisulfite and a method for producing feed using the same, more specifically useful for long-term preservation of fruit and vegetable wastes (FVD) used as feedstock. It relates to a novel use of metabisulfite and a method for manufacturing silage using the same.

When human nutrients that cannot be consumed are used as feed for livestock, it can contribute to expanding the food supply by expanding the feed source and alleviating the instability of the feed. It is estimated that about a third of all processed fruits and vegetables are lost in various parts of the food supply chain (FAO, 2011. The State of Food Insecurity of the World: How Does International Price Volatility Affect Domestic Economies and Food Security? FAO, WFP, IFAD, Rome, Italy). These substances are still low-utility potential nutrients and bioactive compounds, but are self-decomposed in landfills and stockpiles, causing major environmental problems.

According to the results of a field survey conducted by the present inventors in a recent commercial packing house, fruit and vegetable discards (FVDs) are usually accumulated in a warehouse facility for one week until they are moved from the source to the recycling center. there was. However, during this period, especially during the hot summer months, FVD tends to decay rapidly due to accelerated decay and microbial growth due to high humidity and high content of simple carbohydrates. Since such decaying FVD causes environmental problems, it is necessary to immediately treat an effective preservative to the FVD source in order to prevent or delay the decomposition of FVD by microorganisms.

Sulfiting agents are one of the preservatives widely used in the food industry due to their high cost-performance ratio. Among the sulfites, sodium metabisulfite (SMB) is widely used in the feed and food industry, and the silage treated with SMB is effectively preserved due to the oxygen-scavenging activity of SMB that inhibits the growth of oxygen-dependent microorganisms. Can be. Recently, we conducted a series of preservation experiments on a laboratory and pilot scale confirming the successful preservation capacity of SMB. According to this, SMB showed the effect of delaying the deterioration of FVD quality, increasing microbiological stability, and maintaining nutrients during the period of exposure to the aerobic environment. Against this background, the present inventors conducted subsequent studies to further extend the storage stability of SMB-treated FVD under anaerobic conditions and completed the present invention.

Compared to the drying process, which is an energy-intensive and unsustainable method of conservation, ensiling is a long-term preservation of biomass with high moisture content under anaerobic conditions in which bacteria produce lactic acid that drops the pH of the silage down to about 4 It can be a simple yet effective method for. This acidic environment inhibits the growth of harmful parasitic microorganisms during the ensylating fermentation process. However, transportation problems such as high transportation costs and significant loss of dry matter (DM) during ensylation have been pointed out as a major problem associated with the ensilling of high moisture biomass such as FVD. These issues are important in terms of the environment and the economics of the preservation process. To cope with these problems, a silage management strategy that minimizes effluent loss and reduces the costs associated with drying and / or transporting feedstocks with high moisture content. A method has been proposed to achieve an adequate moisture content by adding DM ingredients. However, there have been no reports on the FVD's ensling process where SMB has been processed and exposed to an aerobic environment.

Republic of Korea Patent Publication No. 10-2016-0058241 International Patent Publication WO2018-081550

An object of the present invention is to provide a composition for preserving feed raw materials comprising at least one selected from metabisulfite and organic acids as an active ingredient.

Another object of the present invention is to provide a method for producing feed using fruit and vegetable waste as a raw material by using at least one selected from metabisulfite and organic acids.

In order to achieve the object of the present invention, the present invention provides a composition for preserving feed raw materials comprising at least one selected from metabisulfite and organic acids as an active ingredient. The feedstock is preferably fruit and vegetable wastes (FVD).

In one embodiment of the present invention, the metabisulfite is sodium metabisulfite (SMB).

In one embodiment of the present invention, the metabisulfite is preferably added at a rate of 6 to 8g per 1kg of fruit and vegetable waste, but is not limited thereto.

In one embodiment of the present invention, the organic acid may be one or more selected from sodium benzoate, potassium sorbate, and sodium nitrite.

In one embodiment of the present invention, the composition may further include almond shell and corn gluten.

In addition, the present invention provides a method for producing a feed using at least one selected from metabisulfite and organic acids. The method for preparing the feed includes a pretreatment step of adding and mixing one or more selected from metabisulfite and organic acid to fruit and vegetable wastes in an aerobic environment, and preserving them. It is a manufacturing method. In one embodiment of the present invention, the metabisulfite used in the pretreatment step may be sodium metabisulfite (SMB). In addition, the organic acid may be one or more selected from sodium benzoate, potassium sorbate, and sodium nitrite.

In one embodiment of the present invention, the metabisulfite is preferably added at a rate of 4 to 8g per 1kg of fruit and vegetable waste, but is not limited thereto.

In one embodiment of the present invention, the metabisulfite is added at a rate of 6 to 8 g per 1 kg of fruit and vegetable waste, and the preservation may be 6 to 9 days, but is not limited thereto.

In one embodiment of the present invention, the feed is silage. Silage generally refers to a succulent feed that has been fermented with lactic acid by storing the crop with high water content in a storage container called a silo in vacuum.

In one embodiment of the present invention, the method for producing the silage feed may include an ensiling step of storing and sealing fruit and vegetable wastes that have been subjected to the pre-treatment step in a silo container. .

In one embodiment of the present invention, the ensylating step may be performed at room temperature of 20 to 25 ° C for 7 to 90 days, but is not limited thereto.

In one embodiment of the present invention, the method for preparing the silage feed may be a method of co-ensiling almond husk and corn gluten together with fruit and vegetable waste that has been subjected to the pretreatment step. have. That is, the method of manufacturing the silage feed may include mixing almond shells and corn gluten with fruit and vegetable waste that has been subjected to the pre-treatment step, sealing the silo container, and storing the mixture.

In one embodiment of the present invention, the metabisulfite used in the pretreatment step of the method for preparing silage feed may be sodium metabisulfite (SMB), in which case the sodium metabisulfite is 1 kg of biomass. It is preferable to include in the silo container at a rate of 2.4 to 4 g of sugar, but is not limited thereto.

In one embodiment of the present invention, the method for producing silage feed is mixed by adding one or more selected from metabisulfite and organic acid to fruit and vegetable waste in an aerobic environment at a rate of 6 g per 1 kg of fruit and vegetable waste. Pre-treatment step of preserving for 7 days after prescribing; Crushing the fruit and vegetable waste that has been subjected to the pretreatment step; Mixing almond shells and corn gluten in the pulverized material, wherein the pulverized materials are mixed at a ratio of 39.8 to 66.6% by weight, and the almond shells and corn gluten are mixed at the same weight percentage; And ensylating step of putting the mixture in a silo container and sealing it before storing.

According to the present invention, as a result of treating at least one selected from metabisulfite and organic acids in fruit and vegetable waste (FVD), the microbiological stability and freshness of FVD in aerobic environment was maintained longer, and the nutritional and antioxidant functions of It was also confirmed that it was very effective for preservation. In addition, the method for preparing silage feed comprising a pre-treatment step of treating at least one selected from metabisulfite and organic acids in an aerobic environment according to the present invention has an effect of improving storage characteristics during an anaerobic storage process, especially almonds. The co-ensiling of FVD using shell and corn gluten together showed the best preservation properties.

FIG. 1 shows the chemical composition and proportion (% DM, unless otherwise stated) of the fruit and vegetable wastes used in Experiment 1 and Experiment 2 of Example 1 (mean ± SD, n = 5).
Figure 2 shows the mold appearance (moldy appearance), decayed odor (putrid odor), pH, and water-soluble carbohydrate (WSC) content of FVD with SMB throughput and time (Experiment 1 of [Example 1]).
Figure 3 shows the total number of bacteria (A), fungi (B), and yeast (C) as a function of SMB throughput and time (Experiment 1 of [Example 1]). The error bar for each point represents SE. Values are expressed as the average of five observations. The number of microorganisms is the logarithm number of colony-forming units / g wet biomass. The detection limit was 2.70.
Figure 4 shows the chemical composition of FVD as a function of SMB throughput and time (Experiment 1 of [Example 1]).
5 shows the pH, WSC and ammonia content (NH ₃ -N) of FVD at different locations affected by SMB treatment.
6 shows the results of microbial analysis at other locations affected by SMB treatment (Experiment 2 of [Example 1]). A) after 6 days; B) After 9 days. The total number of bacteria, yeast, and fungi on day 0 relative to the control group was 5.97, 3.86, and 4.03 log cfu / g biomass, respectively. The total number of bacteria, yeast and fungi on day 0 for the SMB treated group was 4.26, <2.70, and 2.83 log cfu / g biomass, respectively. Values are expressed as the average of 5 observations. The number of microorganisms is the logarithm number of colony-forming units / g wet biomass. The detection limit was 2.70. The error bar for each point represents SE.
FIG. 7 shows the nutrient profile of FVD at different locations affected by SMB treatment (Experiment 2 of [Example 1]; mean ± SD).
Figure 8 shows the microbiological and physicochemical properties and antioxidant capacity of liquid FVD affected by SMB treatment after 6 and 9 days.
Figure 9 shows the chemical composition and proportion of the total mixed waste used in the production of silage (mean ± SD, n = 4).
Figure 10 shows the composition of the components of the silage (Experiment 2 of [Example 2])
Figure 11 shows the nutritional composition and microbiological properties of FVD with or without SMB after storage for 0 and 7 days in aerobic conditions (mean ± SD, n = 4).
Figure 12 shows the microbiological and physicochemical properties of FVD with or without SMB treatment according to the anaerobic storage period (mean ± SD, n = 4). The number of microorganisms is the logarithm number of colony-forming units / g fresh matter. The detection limit was 2.8 log cfu per g fresh matter. Ensailing for 0 days corresponds to the end of the retention period (7 days; see FIG. 11).
FIG. 13 shows the change in total DM loss during the 90-day ensylation process. a) Silage without moisture control with or without metabisulfite (Experiment 1 of [Example 2]). b) Moisture-controlled silage with different SMB throughput (Experiment 2 of [Example 2]). Total DM loss was calculated from the deviation between weight and DM content. SMB0 = control silage without metabisulfite added. The percentage of FVD in control silage was 66.6%. Error bars at each point represent SD. SMB = sodium metabisulfite.
14 shows the lapse of time in the content of free sulfite content. a) Silage without moisture control with or without metabisulfite (Experiment 1 of [Example 2]). b) Moisture-controlled silage with different SMB throughput (Experiment 2 of [Example 2]). Day 0 corresponds to the end of the aerobic preservation (day 7). The detection limit was 2 mg / kg DM. Error bars at each point represent SD. SMB = sodium metabisulfite.
15 shows the composition of nutrients in silage having different SMB throughput during each storage period.
16 shows the microbiological and physicochemical properties of silage with different SMB throughput during each storage period.
17 shows the results of chemical and microbiological analysis for the evaluation of aerobic stability of silage after 90 days of ensylation and 5 days of aerobic exposure.

The present inventors conducted two experiments using sodium metabisulfite (SMB) to investigate the effect of metabisulfite on the preservation of fruit and vegetable waste (FVD) in an aerobic environment. One). First, in the first experiment, metabisulfite was treated for 0, 3, 6, 9, and 12 days in amounts of 0, 2, 4, 6, and 8 g / kg FVD, resulting in 6 and 8 g / kg FVD. It was confirmed that the metabisulfite treated with sheep was most effective in preserving nutrients and suppressing spoilage for 6 and 9 days, respectively. The second experiment was a pilot-scale experiment where SMBs of 0 and 8 g / kg FVDs were treated for 0, 6, and 9 days in a 600-L bucket in an outdoor environment. The SMB treatment was very effective in maintaining the integrity and freshness of FVD, inhibiting microbial growth, and preserving nutrients. According to the embodiment of the present invention, SMB can effectively preserve FVD in an aerobic environment, and accordingly, the FVD can be used as a feed through long-term recycling.

Furthermore, the present inventors conducted two experiments for evaluating the storage stability of FVD through ensiling based on the experimental results (Example 2). In this experiment, SMB was not treated with FVD, or SMB of 6 g SMB / kg FVD was treated and pre-treated in aerobic environment for 7 days. In the first experiment, SMB-treated FVD with high moisture content biomass was ensylated alone. SMB-treated silage had a low microbial population and negligible loss of nutrition. In the second experiment, the SMB-treated FVD was co-ensiling in various proportions with almond shell and corn gluten, in which case the calculated SMB loads were 4, 3.2, 2.4 and 1.6. g / kg biomass. On the other hand, as a result of experiments with microbiological analysis, nutritional characteristics, and sensory evaluation, it was evaluated that SMB throughput of 3.2 g / kg biomass showed the most desirable storage characteristics.

Furthermore, the present inventors pretreated to treat SMB and organic acids based on the above experimental results and put them in an aerobic environment for 7 days. As a result, the treatment showed minimal loss of building and water-soluble carbohydrates, and the combined use of SMB and organic acid promoted the growth of useful microorganisms, inhibited the growth of harmful bacteria, and was effective in preserving nutrients.

Therefore, the present invention can provide a use as a composition for preserving feed materials of metabisulfite, which can improve the storage properties of fruit and vegetable wastes that can be used as feed materials, based on which fruits and It is possible to provide a method for producing a recycled feed for fruit and vegetable waste, particularly silage feed, with improved storage characteristics, including pre-treating metabisulfite in vegetable waste.

Hereinafter, the present invention will be described in more detail by examples. The following examples are only intended to illustrate the present invention in more detail, and it will be apparent to those skilled in the art that the scope of the present invention is not limited to these examples.

[Example 1] Effect of SMB (sodium metabisulfite) treatment on FVD (fruit and vegetable discards) preservation

Experiment method

1-1. Sample preparation and experimental design

The first experiment (Experiment 1) consists of 25 combinations. Briefly, FVD (about 600 kg) was obtained for 3 consecutive days in a commercial packing house (E-mart Fresh Center, Icheon, Korea). The wastes were crushed into approximately 30-40 mm pieces and thoroughly mixed and then randomly divided into 5 sub-samples. Metabisulfite powder was not added or was added in an amount of 2, 4, 6 or 8 g / kg wet FVD. The ingredients in each bucket were thoroughly mixed with SMB for 10 minutes using a feed mixer (DDK-801 M, Daedong Tech, Korea), and randomized into 5 replicate buckets for each treatment group. The bucket was divided so that 4 kg of FVD was included in each bucket. Accordingly, a total of 125 buckets (10L) were prepared. The bucket was placed for 0, 3, 6, 9 and 12 days in an aerobic environment such as ambient temperature. At the above time points the total ingredients contained in each bucket were thoroughly mixed and representative sub-samples were obtained at random locations for chemical and microbiological analysis.

In the second experiment (Experiment 2), FVD (about 2500kg) was obtained for 3 consecutive days in the same place as above. Metabisulfite powder was not added or was added in an amount of 8 g / kg wet FVD. After crushing and thoroughly mixing the FVD in the same manner as above, each treatment group was randomly distributed into 4 replicate buckets to include 300 kg of FVD in each bucket. To minimize sampling errors, FVDs were placed in mesh sacks (280 mm wide x 560 mm long, 1 mm void) for every 2 kg of FVDs. The sacks were placed in three different locations in each bucket: surface (0-5 cm deep), middle (25-40 cm deep) and bottom (55-70 cm deep). The bucket (600-L capacity; 102 cm in height; 97 cm in width) was placed under aerobic exposure conditions outdoors for 0, 6 and 9 days. The maximum, minimum and average temperatures during the experiment were 28.4, 22.5, and 25.1 ° C., respectively. At the specified time for each treatment group, all contents in each bag were collected and mixed thoroughly, and representative samples were randomly selected for analysis.

1-2. Visual, microbiological, and physicochemical characterization

The evaluation of mold appearance was scored as 0 if the mold was not clearly visible and 1 if it was clearly visible (Kwak, WS, et al., 2008. Broiler litter supplementation improves storage and feed-nutritional value of sawdust-based spent mushroom substrate.Bioresource Technology 99, 2947-2955). The absence and presence of decaying odors were scored as 0 and 1, respectively (Kwak et al., 2008). Three panels performed this visual and sensory evaluation.

To investigate the metabolites produced during the storage period, 20 g of each sample was mixed with 80 mL of deionized sterile water in a sterile plastic bottle to obtain a juice extract. The suspension was shaken for 10 minutes and then filtered through two layers of cheesecloth. The pH of the extract was measured with a portable electrode (HI9321, Hanna Instrument, Portugal), and the microorganisms were counted. Total bacteria and lactic acid bacteria (LAB) are spread-plated on plate count agar and MRS agar (Difco Laboratories Inc., Detroit, MI, USA), respectively. Were counted using. Plates were incubated at 36 ± 1 ° C. for 48 hours (HK-IB157, Korea Total Instrument, Korea). Yeast and mold were visually identified and counted by spread-plating method in yeast extract glucose chloramphenicol agar (Difco Laboratories Inc.). The plates were incubated at 25 ± 1 ° C. for 3 days for yeast and 5 days for mold. The detection limit was 2.7 log cfu / g. Using the S-1100 UV spectrophotometer (Scinco, South Korea), water-soluble carbohydrates (WSC) are phenol-sulfuric acid procedure (Dubois, et al., 1956.Colorimetric method for determination of sugars and related substances. Analytical Chemistry 28, 350-356), NH3-N was quantified by phenol-hypochlorite procedure (Chaney and Marbach, 1962. Modified reagents for determination of urea and ammonia. Clinical Chemistry 8, 130-132). DM (Analysis No. 930.15), ether extract (Analysis No. 2003.05), crude protein (N × 6.25; Assay No. 990.03), ash (ash) (according to the standard analytical method of The Association of Official Analytical Chemicals (2012) Assay No. 942.05), NDF (neutral detergent fiber) (containing heat-resistant α-amylase and sodium sulfite; Assay No. 2002.04), and acid-detergent fiber (ADF) (Assay No. 973.18) were measured. Non-fibrous carbohydrates (NFC) were calculated by the formula 100-[crude protein + NDF + ether extract + ash].

One- 3. 2 ,2- Diphenyl -One- picrylhydrazyl Assay

2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical-erasing assay was performed on the liquid phase formed in the process of leaking juices and sugars from the cut FVD tissue of Experiment 2 above. Experimental methods are known methods (Wootton-Beard et al., 2011. Stability of the total antioxidant capacity and total polyphenol content of 23 commercially available vegetable juices before and after in vitro digestion measured by FRAP, DPPH, ABTS and Folin-Ciocalteu methods Food Research International 44, 217-224). Samples were centrifuged at 4000 × g for 5 minutes at room temperature, and the supernatant was collected. A methanol solution of DPPH • (0.1 mM, 3.9 mL) was added to a 50-μL aliquot of the supernatant. The mixture was left at room temperature for 30 minutes under dark conditions. Absorbance was measured at 517 nm with an S-1100 UV spectrophotometer (Scinco, Korea). The percent inhibition of DPPH radicals was calculated for the initial DPPH absorbance (without samples).

1-4. Data analysis

The data of Experiment 1 was analyzed using PROC MIXED with a completely randomized design. The treatment groups were arranged in a 5 × 5 factorial design, making a total of 25 combinations. Separation of least-square means was performed using Tukey's range test. The data in Experiment 2 were verified using ANOVA and the deviation between the mean of the treatment group using Student's t -test. The replicate bucket was considered an experimental unit. Microbial counts were converted to logarithm number of colony-forming units prior to statistical analysis. P <0.05 was considered as a significant value.

Experiment result

1-1. Initial characterization

The components and chemical composition of FVD used in Experiment 1 and Experiment 2 are shown in FIG. 1. The proportion of each component of the total waste was calculated as the average monthly production of waste at the end of the experiment. The above main ingredients accounted for more than 95% of the total waste each month. In experiments 1 and 2, oranges and plums each accounted for the largest proportion of the total waste mixture. Among the analyzed ingredients, sweet pumpkin and potato showed the highest DM content, and tomato and onion showed the lowest DM content. Crude protein content was highest in green onion stems and lemons, and grapes and apples had the lowest ash content. The ether extract was negligible in potatoes and showed the highest content in sweet pumpkin and chive stems. The NDF content was highest in sweet pumpkin, followed by paprika and chive stems, and lowest in grapes and potatoes.

1-2. Experiment 1: Visual, microbiological and chemical characterization

As a result of examining the appearance of FVD with or without SMB treatment during the experimental period, it was confirmed that the FVD treated with 6 and 8 g SMB / kg biomass maintained freshness without damage even after 9 days. In general, FVDs that do not process anything gradually undergo browning. However, as the SMB load increased, the color change decreased, because the SMB inhibits the browning reaction. Sulfites readily react with carbonyl intermediates, blocking the formation of brown pigments during the non-enzymatic browning process. In addition, sulfite inhibits many enzyme-catalyzed reactions, especially the activity of polyphenol oxidase, a catalyst that oxidizes phenol compounds to quinones. Quinones play a key role in the production of brown pigment in damaged tissue during the enzymatic browning process. In addition, sulfite treatment served to maintain the integrity of the tissue. In the case of FVD without SMB treatment, the cell wall is decomposed by enzymes secreted from the damaged tissue, so it softens rapidly.

The mold appearance and decaying odor of FVD with SMB throughput and time are shown in FIG. 2. Three days later, mold appeared on the surface of the waste with a decaying odor in the control sample, and in the sample treated with the smallest SMB (2 g / kg biomass), it appeared to be less than the control. This shows that the waste biomass decays very quickly. In the sample treated with the most SMB, there was no decaying odor until day 9, which means that the microorganisms causing decay were suppressed. In addition, sulfite has an effect of inhibiting the oxidation of carotenoids and essential oils, which are associated with the generation of undesirable odors, and thus partially explains the phenomenon of the occurrence of malodor in SMB-treated FVD. can do.

In the samples treated with the most SMB, mold formation and decay were effectively delayed for 9 days, but after 12 days, there was a slight decaying odor, which seems to be due to the progressive loss or inactivation of SMB during the retention period. Metabisulfite is a strong reducing agent that oxidizes when exposed to oxygen, reducing its preservative effect. Overall, treatment of SMB of 6 and 8 g / kg biomass was shown to effectively prevent decay and mold formation of FVD until 6 and 9 days, respectively.

3A-C show the total number of bacteria, mold and yeast over SMB load and time during aerobic storage of FVD. Compared to the number of the microorganisms detected on day 0 (before SMB treatment), the number of the microorganisms in the sample treated with the most SMB (8 g / kg FVD) was significantly reduced, which indicates that the microbial inhibitory effect of SMB Because it appeared immediately. Microbial populations detected on day 0 represent microorganisms that occur and reside during the harvest and distribution, transport, storage and packaging of fruits and vegetables.

In the control sample, total bacteria rapidly increased, and after 3 days, viable bacteria reached 7.79 log cfu / g. However, SMB treatment of 6 and 8 g / kg biomass was very effective in inhibiting the growth and proliferation of total bacterial individuals even after 9 days. Similarly, in the control sample, yeast and mold individuals increased significantly as the experimental period was extended (after 6 days). However, the growth of yeast and mold was very effectively inhibited in the samples treated with the most SMB. Although the mechanism of inhibiting microbial activity has not been identified, it is thought that the sulfite agent is toxic to enzymes and inhibits enzymes essential for microbial growth. It is also believed that the strong reducing power of SMB was toxic to aerobic organisms and some enzyme systems.

The pH and WSC content of FVD over SMB throughput and time are as described in FIG. 2 above. During the experimental period, the pH of the SV-treated (especially treated with SMB of 6 and 8 g SMB / kg biomass) remained relatively constant, but decreased by about 0.6 units after 9 days in the control sample. Did. This shows that FVD is effectively preserved by SMB processing. In the FVD sample without SMB treatment, the WSC content gradually decreased (63.5 g / 100 g DM on day 0 to 12.7 g / 100 g DM on day 12). However, in the samples treated with SMB of 4-8 g SMB / kg FVD, the WSC content decreased during the experiment period to a negligible level.

The decrease in pH and WSC according to the progress of decay in the control sample may be because WSC is used according to the rapid increase of microorganisms (FIG. 3). In general, pH is an indicator of acid production and decay during the aerobic decay process. When the acid production rate exceeds the decay rate, a decrease in pH is usually observed during the aerobic decay process.

1-3. Experiment 2: chemical composition analysis

The nutrient composition profile of FVD over SMB throughput and time is shown in FIG. 4. As the SMB throughput increased, the ash content increased from 4.50% to 7.61%, which is related to the high ash content of SMB (71.7%, FIG. 1). As the experiment progressed, the nutritional component profile of FVD treated with or without SMB or 2 g SMB gradually changed compared to other treatment groups. Compared to day 0, the NFC content of the control sample decreased by 30.2% after 12 days, which means the accumulation of NDF, protein, ether extract and ash. This is evidenced by the increased NDF and ADF content of aerobic unstable corn silage from other studies, and can be explained by the depletion (use) of WSC and the resistance of the fiber fraction to aerobic decay. However, the NFC content of the SMB-treated samples (4-8 g) was not significantly affected during the storage period, suggesting the efficacy of SMB to maintain components that are easily oxidized in FVD. The high content of NFC in FVD means that it can be considered as a partial replacement for energy-dense feed.

The DM loss in the control sample increased with a longer storage period (22% decrease after 12 days), which may be due to the conversion of carbohydrates to water and carbon dioxide during the aerobic decomposition process. Silage's studies on aerobic decay indicate that high populations of yeast contribute to the rapid conversion of soluble carbohydrates to ethanol, carbon dioxide and water, resulting in significant DM loss. A large number of yeasts in the control sample can contribute to rapid WSC depletion and DM loss (Figure 3C).

Crude protein content was not different in FVD treated with 4-8 g SMB / kg biomass. However, in the control sample and FVD treated with 2 g SMB / kg biomass, the above components increased during the experimental period. Conversely, the DM content of FVD treated with 4-8 g SMB increased during the experiment period, which may be related to its surface dehydration. However, despite the continued increase of microorganisms and accelerated decay, the loss of nutrient composition in FVD treated with 4 g SMB was negligible even after 12 days. Overall, SMB treatment of 6 and 8 g / kg biomass was very effective in preserving the nutrients of FVD even after 12 days of aerobic storage. Based on microbiological and sensory evaluation, SMB treatment of 6 and 8 g / kg biomass during aerobic exposure provides effective preservation for at least 6 and 9 days, respectively.

1-4. Experiment 3: Microbiological characterization

In this pilot scale experiment, we obtained sub-samples at different locations of the top surface, center and bottom portions of each bucket. The physicochemical and microbiological properties of the waste samples obtained at these different locations were found to be very diverse. Therefore, the data of this experiment did not combine with each other between the positions. Also, a few days after the FVD was transferred to the bucket, the materials began to flow out into the liquid. Therefore, the present inventors used separated samples for liquid phase microbial analysis.

The microbial organisms of FVDs at various locations affected by SMB treatment are shown in FIG. 6. Metabisulfite led to a significant inhibitory effect on the microbial profile of FVD. Similar to Experiment 1, SMB treatment significantly inhibited yeast and fungal individuals. Compared to the control group, FVD treated with SMB (8 g) showed a fairly small number of total bacteria, yeast and fungi even after 9 days. Fungal growth was not evident in the SMB-treated samples from the center and bottom. Both control and SMB-treated samples showed a greater number of total bacteria, yeast and fungi in the upper part than in the central and bottom positions. Compared to the 6th day of storage, a logarithmic reduction of 0.36 and 0.61 log cfu / g was found in the total number of bacteria and yeast in the upper part of the control sample after 9 days.

The changes in pH, WSC and ammonia concentrations of FVD at these various locations affected by SMB treatment are shown in FIG. 5. Similar to Experiment 1, the pH on day 0 of FVD was acidic and slightly increased (0.18 units) after SMB treatment. Yeast and fungi were believed to be the main causes of microbiological spoilage of FVD due to their ability to grow and dominate in low pH fruits. In general, metabisulfite ions in undissociated form are toxic because they can penetrate into microbial cells. The increased concentration of hydrogen ions (H +) will inhibit dissociation of the sulfite molecule when the maximum concentration in undissociated form is between pH 3-4. In both experiments, the acidic pH of the initial FVD indicates that most of the SMB was not dissociated and was present in a more sterile form.

Although the pH values at different locations of the SMB-treated biomass were slightly different, the pH value was significantly increased with the longer experimental period in the upper sample of the control sample (3.67 at day 0 to 6.61 after 9 days) . This increase in pH usually occurs in aerobic decaying silage, because the organic acid is oxidized to carbon dioxide and water, and at the same time the production of acidic end products due to metabolism of aerobic microorganisms decreases.

Compared to the control group, the WSC content was reduced to a much lower level in the SMB-treated FVD, which means that the SMB treatment effectively inhibited microorganisms related to the rapid use of carbohydrates. However, as the experiment progressed, the decrease in WSC content in the control sample was marked. For example, the WSC content on the top surface decreased from the initial content of 60.1 g / 100 g DM to 2.47 g / 100 g DM on day 9. The reduction can be explained by the active growth and metabolic activity of aerobic microorganisms that can easily use carbohydrates on the upper surface (FIG. 6). In addition, the downward movement of the soluble component is partly related to the reduction of the upper WSC.

The ammonia content increased as the experiment progressed in both the control and SMB-treated samples. However, in the SMB-treated samples, the increased amount was less, indicating the inhibitory effect of SMB on microorganisms involved in proteolysis and ammonia production. After 6 days of aerobic storage, the ammonia content on the upper surface of the control sample increased from the initial value of 4.09 to 24.3 μg / g DM and subsequently decreased to 12.3 μg / g DM after 9 days. This loss can be explained by the conversion of ammonia nitrogen into nitrate nitrogen and then nitrite nitrogen and nitrous oxide under an aerobic environment.

1-5. Experiment 4: Analysis of nutrient composition

7 shows the composition of FVD nutrient composition at different locations affected by SMB treatment. The same pattern as the nutrient composition pattern of wastes observed in Experiment 1 was found in a pilot-scale experiment. Similar to Experiment 1, SMB treatment was very effective in preserving the nutrient composition of the waste after 9 days. In general, various changes in chemical components were observed at different positions of the control sample, and the surface of the upper layer had a significant change in chemical components. For example, after 9 days, the NFC content on the upper surface decreased to 8.57% from the initial value of 68.6%, and to 32.5% in the bottom region. This reduction appears to be due to the rapid depletion of the components that are easily oxidized, and the relative proportions of carbohydrates (NDF and ADF), crude proteins, ether extracts and ash in more complex structures increased.

The accumulation of fibrous fraction can be explained by its chemical complexity, which makes it resistant to aerobic decay. At the same time, readily available carbohydrates can be easily consumed by microorganisms. The fat component of the upper part of the control sample also increased significantly (1.65% at day 0 to 6.84% after 6 days of aerobic preservation), indicating the resistance of the fat component to aerobic decay.

1-6. Experiment 5: Liquid Fraction  Microbial and antioxidant properties analysis

The microbiological profiles of liquid FVDs affected by SMB treatment after 6 and 9 days are shown in FIG. 8. The control sample showed a high number of viable LAB, fungi, and yeast, but the microorganisms were not found at the detection level of 500 cfu / g biomass in the SMB-treated sample.

After 6 and 9 days, the WSC content in the SMB-treated biomass was significantly higher than in the control group, and this phenomenon can be explained by a significant reduction in the microbial population that readily consumes WSC in the SMB-treated sample. Similar to the data on the ammonia content of FVD at other locations, a much lower concentration of ammonia was found in the SMB-treated samples compared to the control 9 days after storage (31.1 vs. 63.3 μg / mL).

The SMB-treated liquid obtained after 6 and 9 days scavenged 26% and 31% more DPPH radicals, respectively, compared to the control, meaning that SMB treatment preserved the antioxidant capacity of FVD. The reducing ability of the liquid fraction to DPPH radicals decreased slightly in both the control and SMB-treated samples as the experimental period increased from 6 days to 9 days.

In conclusion, based on the visual, microbiological and chemical evaluations, metabisulfite treatment of 6 and 8 g / kg biomass with aerobic exposure revealed stability and freshness of discarded fruits and vegetables, 6 and 9 days, respectively. It can be seen that it can be maintained for a day. Pilot-scale experiments have demonstrated that metabisulfite treatment for 9 days in an aerobic environment is effective in retarding spoilage, improving microbiological stability, and preserving nutrients and antioxidant function.

[Example 2] Long-term preservation of FVD through SMB treatment

Experiment method

2-1. Sample preparation and preservation studies

Detailed procedures regarding the sample preparation and preservation procedures were followed by our previous research methods. Briefly, FVD (about 600 kg) was obtained for 3 consecutive days in a commercial packing house (E-mart Fresh Center, Icheon, Korea). Information on the amount of individual ingredients to be discarded each month was investigated through input / output data provided by the Chief of Staff. In the month of the experiment, major wastes, which account for more than 90% of the total wastes, were obtained from investigations at the shipyard facility. The chemical composition of the individual components is shown in Figure 9.

The wastes were cut into approximately 30-40 mm pieces, mixed thoroughly and randomly divided into 7 samples. Metabisulfite powder was added in an amount of 0 or 6 g / kg FVD. Using a feed mixer (DDK-801 M, Daedong Tech, Korea), the samples were thoroughly mixed for 10 minutes under the condition of adding or not adding SMB, and then transferred to a 200 L bucket. To simulate the actual situation of FVD accumulation at the source, the bucket was placed for 7 days in an aerobic environment such as ambient temperature. Representative sub-samples were obtained at any location for chemical and microbiological analysis.

2-2. Silage manufacturing

After 7 days of exposure to the aerobic environment, SMB-treated FVDs were stored in silos with high humidity biomass alone or with almond shells and corn gluten supplied at different rates. The percentage of FVD in silage according to this example was 66.6%, 53.2%, 39.8% and 26.6%, which is calculated to provide 4, 3.2, 2.4 and 1.6 g of SMB per kg of biomass, respectively (Figure 10). .

Silage with a content of 66.6% FVD without SMB was used as a control. To minimize the difference in particle size, the FVD was ground using an electronic grinder. FVD ground without a water absorbent was ensiled in a 3L glass bottle silo weighed in advance. Four glass bottles were used for each treatment.

The rest of the ground FVD was mixed with different proportions of almond shell and corn gluten feed using a feed mixer (DDK-801 M, Daedong Tech, Korea). FVD biomass is characterized by a high content of simple carbohydrates, a high moisture content, and a low protein content. Therefore, considering not only the nutritional balance of the ensiling mixture, but also water retention and economic usefulness, an almond shell is used as a byproduct of water absorbent rich in fiber, and corn gluten as a byproduct of water absorbent rich in energy and protein is ensylated with FVD. It was selected as an ensling companion. The mixture (about 4 kg) was packaged in a polyethylene bag (45 cm wide x 75 cm high) and tightly sealed before ensiling at room temperature (20-25 ° C). Silos were weighed on preselected opening days (7, 14, 30, 60 or 90 days) and representative subsamples were randomly obtained for nutritional and microbiological evaluation. Silage was visually and sensoryly assessed by three experienced appraisers.

2-3. Microbiological evaluation and analysis method

The fermentation metabolites and microbial populations were investigated from the silage extract obtained by mixing the silage sample (20 g) with 120 mL of autoclaved distilled water. The suspension was shaken for 5 minutes and then filtered through two layers of medical gauze. The pH of the silage extract was immediately measured with a portable electrode (HI9321, Hanna Instrument, Portugal). Water-soluble carbohydrates (WSC; glucose equivalent); Dubois et al., 1956) and NH3-N (Chaney and Marbach, 1962) for centrifugation of another portion of the filtrate at 4 ° C. and 10,000 g for 10 minutes Did.

Ammonia-N is given on an N equivalent basis. After appropriate serial dilution of the silage extract, the microbiological composition of the silage sample was analyzed using a spread-plating method. The total number of viable bacteria was counted in plate count agar (Difco Laboratories Inc., Detroit, MI, USA). Viable colonies of lactic acid bacteria (LAB) were counted in MRS agar (Difco Laboratories Inc., Detroit, MI, USA). Plates were incubated at 36 ° C ± 1 ° C for 48 hours (HK-IB157, Korea Total Instrument, Korea). Yeast and mold were counted by yeast extract glucose chloramphenicol agar (Difco Laboratories Inc.). For counting, the plates were incubated at 25 ° C ± 1 ° C for 3 days for yeast and 5 days for mold. Differentiation of yeast and fungi was achieved by visual evaluation of colony appearance.

For nutritional component analysis, a fresh sample (about 200 g) was dried in a forced-air oven (72 hours at 65 ° C.) and then ground to a particle size of 1 mm. According to the standard analysis method of AOAC (The Association of Official Analytical Chemicals, 2012), DM (Method No. 930.15), ether extract (Method No. 2003.05), crude protein (Method No. 990.03), crude ash (Method No. 942.05), neutral detergent fiber (NDF) (containing heat-resistant α-amylase and sodium sulfite; method No. 2002.04), and acid-detergent fiber (ADF) (method No. 973.18) were measured. Non-fibrous carbohydrates (NFC) were calculated by the formula 100-[crude protein + NDF + ether extract + ash].

The time course of total DM loss was calculated from the difference between the initial weight (day 0) and the ensiling 7, 14, 30, 60 and 90 days, based on the DM content of the silage mass. The free sulfite content of the SMB-treated sample was quantified according to an enzymatic colorimetric method using a commercial kit (K-SULPH kit, Megazyme International Ireland Ltd., Bray, Co., Wicklow, Ireland). The aerobic stability of the final silage (90 days) was determined using a bottle system that measures the amount of carbon dioxide produced during the aerobic deterioration process (Ashbell et al., 1991). The silage was exposed to aerobicosis for 5 days at room temperature (25 ° C). The amount of carbon dioxide produced after aerobic decay, pH value, total bacteria, yeast and mold count were used as indicators of aerobic spoilage.

2-4. Statistical analysis

Microbial data were converted to log ₁₀ prior to statistical analysis. The preservation study of Experimental Method 1 and statistical analysis of silage were performed using a general linear model of SAS (SAS, 2004). When the F value was significant, the deviation between least-square means was confirmed using the t -test at the 0.05 probability level. The rest of the data was analyzed using PROC MIXED with a completely randomized design in SAS (SAS, 2004). The model below was used for analysis.

Y _ij = μ + T _i + ε _ij

Here, Yij is the observation j in treatment i, μ is the average, Ti is the storage period or the fixed effect of the treatment (aerobic stability test), εij is the residual error. Polybag silos were considered experimental units. When a significant treatment effect was observed (P <0.05), separation of least-square means was performed using Tukey's test. The linear response to the storage period from 0 to 90 days was estimated using polynomial contrasts (SAS, 2004).

Experiment result

2-1. Aerobic preservation

The nutritional composition and microbiological properties of FVD with or without SMB after storage for 7 days under aerobic conditions are shown in FIG. 11. Overall, SMB was very effective in minimizing nutrient loss and reducing the number of bacteria, yeast and fungi. LAB was found to be very vulnerable to SMB and was not found at a detection level of 2.8 log cfu / g in SMB-treated samples after storage for 7 days. LAB produces various amounts of hydrogen peroxide, which are known to react strongly with sulfite to produce sulfur trioxide anion radicals. These radicals are known to strongly inhibit the cell growth of LAB. In addition, the results of the unpublished preservation studies by the present inventors showed a strong inhibitory effect of SMB on LAB, yeast, and fungi.

After storage for 7 days under aerobic conditions, the WSC content was significantly decreased in the control sample, but crude protein (CP), ash, and NDF accumulated. This increase can be explained by the reduction of organic matter during the preservation process in aerobic conditions of FVD, thereby converting carbohydrates into carbon dioxide and water. As in Example 1 above, the present inventors performed a series of preservation experiments with SMBs of various loadings on a laboratory and pilot scale, resulting in 6 and 8 g / kg wet mass under aerobic exposure environment. It has been found that SMB keeps FVD relatively stable and can prevent loss of nutrients for up to 6 and 9 days, respectively. Therefore, the purpose of the experiments according to this embodiment was to extend the shelf life of FVDs for a longer period of time through ensiling.

2-2. Experiment 1: Anaerobic storage without moisture control

The microbiological and physicochemical properties of FVD with or without SMB according to the anaerobic storage period are shown in FIG. 12. The development of viable LAB reached a peak in 7.67 log ₁₀ cfu / g biomass after 14 days of ensiling, decreased over time in control silage, and 5.96 log cfu after 90 days of fermentation. / g The biomass has been reached. This can be seen as a general result as previous studies have reported that the number of LABs decreases as the duration of ensiling progresses. However, no LAB was found in SMB silage (detection limit = 2.8 log cfu / g). For efficient anaerobic conservation, lactic acid fermentation must be done quickly. LAB is known to serve as a major consumer of WSC during ensiling by converting WSC to lactic acid and promoting a sharp decrease in silage pH.

It was confirmed that the rapid development of LAB in this example immediately reduced the pH of the control silage (FIG. 11). However, as the number of unwanted microorganisms (ie, yeast and mold) in the silage mass increased, a decaying odor was detected. According to the results of previous studies, depending on the microflora of the pre-ensiling biomass, even if the number of LABs increases rapidly, it suppresses undesirable microbial growth or prevents DM loss during ensiling. You may not be able to. Essentially, lactic acid is a weak fungicidal compound, so if you have a large number of undesirable microorganisms, it will not be strong enough to stop their growth. Although the LAB proliferated rapidly in the embodiment of the present invention, the growth of undesirable microorganisms during ensiling was not inhibited, so the above phenomenon may also occur in this embodiment. Unlike control silage, LAB was reduced below detectable levels in SMB-treated silage over a 90-day storage period, but the loss of nutrition was negligible with respect to the small number of undesirable microorganisms. These results suggest that SMB is more effective as a silage fermentation inhibitor than hydrogen ions when FVD is ensylated without absorbents.

During the 90-day storage period in SMB-treated silage, mold counts were below the detection limit, but yeast was low during the early stages of ensiling (less than 4 log cfu / g) 60 SMB silage was not detected after days and 90 days of storage. Part of this observation may be related to the provision of strict anaerobiosis by SMB in the early stages of storage.

When SMB is used as a silage additive, it decomposes into sulfur dioxide, a strong antifungal compound that consumes a significant amount of oxygen during ensiling. This reduces aerobic stress during the early stages of the ensiling process, thus providing effective inhibition against aerobic microorganisms. Although the antimicrobial mechanism of sulfiting agents is not clearly understood, it appears that sulfites react easily with biomolecules through substitution reactions at electrophilic sites because of their strong nucleophilicity. It initiates a series of reactions that can damage key enzymes involved in the metabolism of microbial cells.

The NH ₃ -N / N content in both the control and SMB silage for 90 days of ensiling was lower than the total N threshold of 70 g / kg, estimated to be an indicator of successful silage fermentation. The low ammonia content of the control silage may be associated with a rapid decrease in pH, which is known to minimize proteolysis.

In addition, the total DM loss during the storage process is shown in FIG. 13. A major DM loss was seen in the control silage and a 21.1% loss during the 7 day ensiling period. A similar trend was observed in the WSC content, which decreased from 29.7% on day 0 to 16.6% after 7 days of ensiling. However, after 30 days of ensiling, the silage fermentation appeared to have stabilized. Unlike control silage, little loss of DM and WSC was observed in SMB-treated silage (DM decreased from 15.3% on day 0 to 14.3% after 90 days).

The strong microbial population present in the control silage mass prior to ensiling contributed to the complete oxidation of organic matter, such as WSC and fermentation products, into carbon dioxide and water, which is associated with excessive loss of DM and nutrients. This represents a potential loss of feed nutrients available to the animal, which is an undesirable waste of energy to avoid. In general, preservation through ensiling is usually within the typical range of 15% -20%, and is mainly related to the energy cost for DM loss (known as 'reducing silage') resulting from carbon dioxide production, This is of environmental and economic importance. This DM loss increases even more when the biomass with a high sugar content and a low DM content is ensiling. Consequently, factors contributing to excessive loss of nutrients in the control silage are probably the following: 1) aerobic exposure of FVD prior to ensiling to increase the number of decaying microorganisms; 2) high water content of FVD; And 3) high WSC content of FVD.

Regarding the storage period, the time course of the sulfite concentration freed from SMB silage is shown in FIG. 14. Compared to day 0 of storage, about 46% of the free sulfite was lost after 90 days of storage. This may help partially explain the slight increase in microbial populations as ensiling progresses in SMB-treated silage. In principle, when sulfite is used as a preservative, it will exist in a free form and in a combined form, reaching an equilibrium between the two forms. Of these, only the liberated form exhibits activity and functions as a preservative.

Overall, although rapid development of LAB contributed to a rapid decrease in pH, the control silage exhibited poor preservation properties with regard to undesirable microbial profiles and excessive DM loss, which is a silage from FVD that does not treat effective preservatives. It is a factor that hinders the possibility of commercialization of production. However, it has been demonstrated in this example that FVD can be treated with SMB and stored satisfactorily under anaerobic conditions. Metabisulfite alone can provide protection against undesirable microflora, which minimizes the loss of DM and nutrients, thereby providing a cleaner way to use silage more safely over long periods of time. The on-site anaerobic storage of soil FVD after treatment with SMB in an oxygen-limited storage tank can facilitate the practical use of this preservation technique for continuous use as a livestock feed source. One of the main difficulties in standardizing FVD, a component that is steadily used as a livestock feed, is that the composition of ingredients and nutrients varies. Where possible, anaerobic storage of relatively large amounts of FVDs, less than one month in the field, can minimize this variability and better manage this waste as an ingredient for inclusion in animal feed.

2-3. Experiment 2: moisture controlled anaerobic storage

Prior to describing the results of this example, the inventors attempted to make silage by adding almond shells and oat shells to SMB-treated FVDs in early experiments (66% FVD, 17% almond shells, 17% oat shells). ; 42% DM). After 90 days of experimentation, silage began to decay after ensiling began. This was accompanied by a drastic increase in DM and nutrients and a number of yeast and mold. A slight decaying odor was detected along with a widespread alcoholic odor during the ensiling. Similar to Experiment 1 above, LAB was below the detection limit at all sampling times. It is speculated that the increased porosity due to the low packing density and the addition of oat shells resulted in a rapid increase in aerobic microorganisms, especially yeast and mold. It is possible that the hollow structure of the oat shell increased the amount of air in the silage mass and thus increased the activity of aerobic microorganisms in the early stages of ensylation. It also appeared that the SMB load of the silage product was not sufficient to adequately suppress yeast and mold development. This quickly converted readily available carbohydrates to carbon dioxide and water, which is associated with inefficient anaerobic conservation. Based on these assumptions, we provided good compression with minimal air volume in the silage mass by selecting almond shell and corn gluten feed with small particle size in subsequent experiments.

Changes in the chemical composition and conservation properties of silage with different SMB loads are shown in FIGS. 15 and 16 according to storage period (0, 7, 14, 30, 60 or 90 days). The prolonged ensiling increased the ash content slightly in the SMB silage and increased more in the control silage. As expected, the NFC content was significantly reduced in the control silage and decreased to a lesser extent in the SMB silage, indicating an increase in fermentation, oxidation of oxidized components, and increased recalcitrant components such as fibers in the silage. It is an indicator. During normal silage production, the delay in sealing (aerobic exposure) is generally associated with a significant loss of nutrients and DM during ensilling. FVD's on-site accumulation is a potential vector of many undesirable microorganisms. Therefore, the undesirable microbial profile of FVD prior to ensylation (termination of aerobic preservation) could contribute to the development of potent metabolic processes during anaerobic storage in control silage, and thus appears to increase DM and nutrient loss.

In contrast, the nutritional loss of silage with higher SMB throughput showed a slight difference during 90 days of anaerobic storage. For example, in the silage of 3.2 g SMB / kg biomass, the WSC concentration was 21.8% before ensylation, and after 90 days ensylation, it decreased to 19.0% DM. As far as the inventors know, there have been no studies that have previously studied SMB and investigated the anaerobic storage dynamics of aerodynamically challenged FVD.

Ammonia levels increased with increasing storage time in SMB silage, with the highest values observed between 60 and 90 days of ensylation. This indicates that a long ensylation time promotes protein degradation. During the ensylation period, the protein can be decomposed into ammonia, which indicates the clostridial fermentation of the protein. In general, an indicator of successful silage fermentation is an NH ₃ -N content below the 70 g / kg total nitrogen threshold, while an amount greater than 100 g / kg total nitrogen indicates poor fermentation of the silage. As can be seen in Figure 16, it can be seen that there is no SMB silage having a NH3-N / N content higher than a threshold value indicating high quality silage.

Similar to Experiment 1 above, the LAB number was very low or below the detection limit at any SMB silage during the storage period. In general, total bacteria, yeast and fungi showed the lowest population in silage with the highest FVD ratio (higher SMB throughput). LAB proliferated rapidly and dominated the silage fermentation (FIG. 16), but the pH of the control silage did not change during the 90-day ensylation process. Decayed or moldy silage is believed to prevent silage pH reduction during silage fermentation. In the embodiment of the present invention, it is possible to consider the possibility that the above phenomenon occurred from the fact that the population of yeast and mold was large in the control silage accompanied by the odor of decay. In the case of metabisulfite silage, the drier silage (with lower SMB throughput) showed an increase in silage pH as the en- siling period increased. In general, an increase in silage pH during the en- siling period indicates limited lactic acid fermentation, which is associated with undesirable microbial distribution and extensive nutrient loss in traditional silage. However, in these silages, the SMB load was probably sufficient to inhibit LAB growth, but as the ensilling period increased, the active form of SMB would have decreased to a level that would allow the control of undesirable microorganisms. Conversely, in the silage with the highest percentage of FVD (i.e., 4 and 3.2 g SMB / kg biomass), the silage pH was generally constant (about 4) and slightly decreased with prolonged ensylation.

In general, a suitable preservation criterion for ensiled biomass is a target pH of less than 4, at which most undesirable bacteria are inhibited. The bisulfite ion itself provides efficient preservation as opposed to the hydrogen ion (low pH) responsible for preservation in conventional silage. The above results support the conclusion that the use of SMB as a silage fermentation inhibitor is very effective in providing scented and nutritious silage.

As can be seen in Figure 16, since the pH of the FVD was acidic, dissociation of bisulfite ions, which are more toxic to microorganisms when present in an undissociated form, was suppressed. Therefore, it is expected that more effective inhibition by SMB in the natural acidity of FVD will be achieved. Overall, metabisulfite silage provides a new kind of silage that is not preserved through hydrogen ions, as occurs in traditional traditional silage fermentation. Instead, in this form of silage, preservation is mediated by metabisulfite ions, contributing to the effective inhibition of microorganisms and reducing ammonia and acid formation during the ensiding process.

The total DM loss of the moisture-controlled silage according to the storage period is as shown in FIG. The loss of DM in control silage was significant, with approximately 14.8% of total silage DM lost within 30 days of ensylation. However, from then on, the rate of disappearance decreased, and after 90 days of ensylation, it only increased to 17.6%. These observations show that the metabolism of WSC and other components continues until the silage pH drops to the point where the microbial activity slows and the silage fermentation stabilizes, the duration of fermentation and DM loss during ensylation, controlled by the silage microbiome. It is consistent with the results of the meta-analysis that found the inter-relationship.

Unlike the control silage, a metabisulfite silage found negligible levels of DM loss within 30 days of ensylation, but then began to increase slightly. Among the metabisulfite silages, the silage of 3.2 g SMB / kg biomass had the lowest total DM loss, with only 3.98% DM lost in 90 days of ensilling. This suggests that the metabisulfite load and moisture content in this silage provides efficient conservation under anaerobic conditions for a long period of time, without the need for lactic acid fermentation. Co-ensiling SMB-treated FVD with almond shell and corn gluten feed at about 50% is an effective way to control silage decay and possible transport difficulties associated with the transport of juicy substances. It can be seen, thereby increasing the usefulness of the silage and periodically transporting it to a distant location.

2-4. Visual and sensory examinations

Within 7 days of storage, the control silage began to decay, mold appeared on the surface and a slight decaying odor was detected. There was no mold spot on the silage surface with the highest FVD ratio, but mold formation was remarkable on the silage surface after 1 month of ensylation with a theoretical SMB load of 2.4 or 1.6 g SMB / kg biomass. appear. The development of surface mold spots on these silages was accompanied by a decaying odor. The amount of SMB used for silage with lower FVD ratios (ie 2.4 and 1.6 g SMB / kg biomass) may be insufficient to inhibit the growth of perishable microorganisms and subsequent spoilage. Moreover, the high DM content of these silages is likely to contribute to lower compaction and efficient air exclusion, which are important for inhibiting the growth of undesirable microorganisms. Among the SMB silages, those with higher SMB loads showed a stronger SMB odor in en-silling. However, it was found that the smell of SMB gradually decreased as the storage period increased. This can be explained by the gradual loss of SMB during the storage period. When corn feed was treated with SMB (4 g SMB / kg fresh biomass) and ensylated for 90 days, the remaining bisulfite of the treated silage was about a quarter of the initial addition. The reason for this loss of SMB in the ensylating process is explained by the fact that metabisulfite is a strong reducing agent that is oxidized to sodium bisulfate when exposed to reductive components. The approach of air or reducing materials to bisulfite in silage mass is expected to contribute to further lowering of residual sulfite content in silage. Moreover, the generation of sulfur dioxide gas can be another route to SMB loss during ensylation.

Overall, among metabisulfite silage, silage with a 3.2 g SMB / kg fresh mass equivalent to about 50% moisture provided the most desirable organoleptic properties. Compared to silage with 4 g SMB / kg biomass (about 60% moisture level), the FVD ratio (53.2%) of silage with 3.2 g SMB / kg biomass is lower and the DM content is higher, resulting in a more pleasant odor (Lower SMB odor), and the wastewater effluent is negligible and can be considered more desirable.

2-5. Free sulfite concentration

The time course of the liberated sulfite concentration according to the storage period in silage with various SMB throughputs is shown in FIG. 14. Unlike the results of Experiment 1, where a relatively high content of free sulfite was seen in the SMB silage after 90 days of storage, the free sulfite content decreased to a negligible amount after 30 days of ensilling. Although uncertain, free sulfite appears to be easily incorporated into some of the components of almond shells or corn gluten feed. Free sulfite is an active form of SMB that provides strong deterrent to the growth and development of microorganisms. Significant reductions in free sulfite throughput are potential sources of feed nutrients for incorporating ensilized biomass into moist or semi-dry total blended silage (50% -65% DM), or potential for biogas production. It provides an opportunity for efficient use as biomass.

When mixing SMB silage with ruminant feed as a dietary nutrient, it is advisable to consider the sulfur content of the diet. The National Research Council (2000) reported that the sulfur requirement for farm animals is 1.5 g / kg diet DM, and the maximum allowable concentration is about 4 g / kg diet. No metabisulfite loss during storage, 20% dietary inclusion level of metabisulfite silage (3.2 g / kg biomass) additional 0.22 g / kg in diet Assuming conversion to sulfur, this is not practical and can be easily incorporated into diets with some dietary modifications.

2-6. Aerobic stability

The chemical and microbiological properties of the final silage (after 90 days) exposed to air for 5 days are summarized in FIG. 17. In general, silage with a lower SMB concentration (dryer silage) had significantly lower stability with respect to a substantial increase in pH and carbon dioxide production. This was accompanied by the rapid spread of yeast and fungi. In contrast, silage with 4 and 3.2 g SMB / kg biomass was more stable with lower carbon dioxide production, less microbial population and less pH increase. In general, dry silage tends to be more porous than moist silage, which stimulates rapid decay when exposed to air. In general, silage with yeast exceeding 5 log cfu / g DM is expected to decay rapidly upon opening of the silage. Following the oxygenation of silage, yeast begins to multiply rapidly, consuming a significant amount of organic acid and the silage pH rises rapidly. This can explain the rapid decay of control silages and silages with lower SMB throughput, since these silages have a large number of yeasts after 90 days of ensylation (Figure 16).

It was also confirmed that the silage with more SMB throughput (ie, 4 and 3.2 g SMB / kg biomass) was more aerobicly stable as a result of this example.

In conclusion, discarded fruits and vegetables are successfully preserved by metabisulfite after 7 days of aerobic exposure and then converted into a high-moisture biomass for a long time with negligible energy costs associated with dry matter and nutrient loss. It was ensylated. FVD's co-ensiling with almond shell and corn gluten feed, which provided an SMB throughput of 3.2 g / kg biomass, corresponding to a water content of about 50%, provided the best preservation properties. Overall, treatment of metabisulfite in discarded fruits and vegetables does not require fermentation of lactic acid bacteria and can be considered effective for short-term aerobic preservation followed by long-term anaerobic preservation.

[ Example  3] SMB  And organic acid treatment FVD  Long-term preservation

Experimental methods related to long-term preservation of FVD were performed using the same method as in Example 2, except that SMB and organic acid were simultaneously treated.

The organic acids used are sodium benzoate, potassium sorbate, and sodium nitrite, the mixing ratio of which is summarized in FIG. 18, and the amount of SMB and organic acid used is summarized in FIG.

After storage for 7 days under aerobic conditions, the content characteristics of dry weight and water-soluble carbohydrate content (WSC) of FVD with or without SMB and organic acids were investigated and shown in FIG. 20.

As a result, the negative control T1 showed significant loss of building and water-soluble carbohydrate content after 7 days of aerobic storage, but both the test group treated only with SMB (T2) and the test group treated only with organic acids (T3) The loss of water-soluble carbohydrate content was found to be insignificant, and the loss of building and water-soluble carbohydrate content was also observed in the test group (T4 to T6) treated with SMB and organic acid simultaneously.

In addition, microbiological and physicochemical properties of FVD with or without SMB and organic acid after storage for 7 days under aerobic conditions are shown in FIG. 21.

Referring to FIG. 21, in the case of the negative control group, after 7 days of aerobic storage, the total number of bacteria and yeast increased significantly. This means the growth of spoilage bacteria. On the other hand, in the test groups T4 and T5 treated with SMB and organic acid at the same time, the total number of bacteria, yeast and fungi was observed to be lower than the detection level. This is in line with the slight changes in building and water soluble carbohydrates observed in Figure 20.

In addition, the pH characteristics of FVD with or without SMB and organic acid after 7 days of storage under aerobic conditions are shown in FIG. 22. Referring to FIG. 22, in the case of the negative control group, a sharp decrease in silage pH was observed after 7 days of aerobic storage. On the other hand, pH was observed to be consistently low in the T4 and T5 treatment groups, which are the test groups treated with SMB and organic acid simultaneously. Likewise, these results are in line with the minor changes in building and water-soluble carbohydrates observed in FIG. 20.

Through the above results, it was confirmed that the combined use of SMB and organic acid is effective in regulating microbial growth and preserving nutrients.

So far, the present invention has been focused on the preferred embodiments. Those skilled in the art to which the present invention pertains will appreciate that the present invention may be implemented in a modified form without departing from the essential characteristics of the present invention. Therefore, the disclosed embodiments should be considered in terms of explanation, not limitation. The scope of the present invention is shown in the claims rather than the foregoing description, and all differences within the equivalent range should be interpreted as being included in the present invention.

Claims (12)

A composition for preserving feed raw materials comprising at least one selected from metabisulfite and organic acids as an active ingredient, wherein the feed raw material is fruit and vegetable discards (FVD). According to claim 1,
The metabisulfite is a composition characterized in that it is added at a rate of 6 to 8g per 1kg of fruit and vegetable waste. According to claim 1,
The organic acid is a composition characterized in that at least one selected from sodium benzoate (Sodium benzoate), potassium sorbate (Potassium sorbate), and sodium nitrite (Sodium nitrite). A pre-treatment step of adding and mixing one or more selected from metabisulfite and organic acids in fruit and vegetable wastes in an aerobic environment, and preserving the mixture, thereby preserving the fruit and vegetable waste feedstock. The method of claim 4,
The metabisulfite is a method characterized in that it is added at a rate of 4 to 8g per 1kg of fruit and vegetable waste. The method of claim 5,
The metabisulfite is added at a rate of 6 to 8g per 1kg of fruit and vegetable waste, and the preservation is performed for 6 to 9 days. The method of claim 4,
The method characterized in that the feed is silage (silage). The method of claim 7,
And a step of ensylating the fruit and vegetable wastes that have been subjected to the pre-treatment step in a silo container and sealing them before storing. The method of claim 8,
The en-siling step is characterized in that made at 7 to 90 days at room temperature of 20 to 25 ℃. The method of claim 8,
Almond peel and corn gluten (gluten) in a silo (silo) container with the fruit and vegetable wastes that have been subjected to the pre-treatment step, characterized in that the storage after storage. The method of claim 10,
The metabisulfite used in the pretreatment step is sodium metabisulfite (SMB), and the sodium metasulfite is contained in the silo container at a rate of 2.4 to 4 g per 1 kg of biomass. . The method of claim 10,
A pre-treatment step in which a fruit and vegetable waste is added to one or more selected from metabisulfite and organic acid in an aerobic environment at a rate of 6 g per 1 kg of fruit and vegetable waste and mixed for storage for 7 days;
Crushing the fruit and vegetable waste that has been subjected to the pretreatment step;
Mixing almond shells and corn gluten in the pulverized material, wherein the pulverized materials are mixed at a ratio of 39.8 to 66.6% by weight, and the almond shells and corn gluten are mixed at the same weight percentage; And
And ensiling the mixture into a silo container, sealing and then storing.

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