1 Introduction

The prevailing climatic conditions in the tropical regions of the world (with temperature and relative humidity over 25°C and 70% respectively) accelerate moldy growth and lipid oxidation of stored feed, (Berger, 1989; Coppen, 1989; Van den, 1990; Adaga, 2014). According to Bautista et al., (1992) and Ramezandeh et al., (1999) storage at high temperature result in an increase in both oxidative and hydrolytic rancidity with a loss in feed quality. Studies by Hamilton, (1989), Van Dan et al., (1990) and Ruiz et al., (2000) indicate that fats are intrinsically unstable when subjected to a high temperature above 300C. Under such conditions, fats are hydrolyzed to release ketonic acids, which further undergo auto-oxidation with degeneration of racial products (Hamilton 1989).

Rancidity results in loss of quality and acceptability, causing reduced feed intake in fish, (Galliard, 1989; Sanders, 1989). Similarly, NRC, (1981), reported that feed stored for longer than 90 days at ambient temperature is subjected to the breakdown of oil, and vitamins along with peroxidation of lipid component. According to FAO (2001), environmental factors during storage predispose the feeds to undergo microbial spoilage. Toxins producing fungi are also found in stored feeds, most of them producing aflatoxins, patulins, and trichotecens which are strongly carcinogenic, mutagenic and dangerous. Aflatoxins are chemicals produced by fungi as Aspergilus flavus and A. parasiticus (mould), (Russo and Yanong, 2006).

The occurrence of these microbial strains in fish feed has been reported to depend on the storage conditions of the feed (Zmyslowska, 2000). Manufactured feeds are stored under different storage conditions by commercial fish feed sellers and farmers without respect to the effect of these conditions on the nutrient profiles of the feeds. The general rule for the preservation of feed is to store them in a dry, well-ventilated area that can offer some protection from rapid changes in temperature, (Jantrarotai and Lovell, 1990). In the tropics where climatic conditions are harsh, storage of fish feeds to ensure less nutritional deterioration and shelf life longevity is often a problem, hence, different methods have been developed by different group based on what is best suitable in the various localities (Adaga, 2014). Researches on the nutritional profile of feed under different storage conditions are also scarce since the nutrient profile of a feed determines the growth performance of fish; this study was designed to determine the nutrient level of stored feed using the prevalent method of storage among fish farmers in Benue state Nigeria.

2 Materials and Methods

Eight kilograms of each of Coppens^®, Multifeed^® and vitalfeed^® was purchased from a reputable feed store along the modern market road, Makurdi Nigeria. They were divided into two halves (4kg) and stored in open and airtight conditions for six months. The open condition was done by widely opening the feed packaging from the top and left open to the atmosphere for the period of 6months. The airtight condition was created by open the feed packaging from the top however, it was squished together and tied with a rope to provide a simple airtight condition of storage. The choice of these methods of storage for this study was based on a pre-field survey done on the packaging style of most fish farmers around the state. Samples of the feed were collected one a month for different nutrient analysis. Proximate analysis of feeds was determined using the methods of AOAC, (2000). Parameters measured (in percentage) include moisture, ash, crude fiber, ether extract and crude protein. Nitrogen Free Extract (NFE) was determined by the difference between 100% and the other parameters. Organoleptic description of the feeds before, during and after the storage period was done by a team of previously trained assessors using hedonic scales designed for the study (reference).

Peroxide value (POV) and Free Fatty Acid (FFA) analysis were also done according to AOAC, (2000), oils with POV well below 10 mg/kg are considered fresh. While oils with POV between 20-40mg/kg were termed rancid. Mould growth was identified according to the method described by APHA (1998).

3 Results and Discussion

The crude protein content of Coppens^® under airtight condition remains fairly constant during a storage period of six months, while that of Multifeed^® was reduced a little as the time of storage prolonged (Table 1; Table 2; Figure 1). The same trend was observed for Vitafeed^® (Table 3; Figure 1). This could be as a result of protein aging as postulated by Shyong, (1998); however deviation of the trend of decrease observed in Coppens^® may be due to differences in feed ingredient suggesting that the susceptibility of feed to the "protein aging" phenomenon may differ with different feedstuffs. Hossen et al., (2011) reported that changes in the chemical composition and nutritive value of feed may occur during storage. Though the protein content of all commercial feeds stored in open condition reduced as the storage time increased, the marginal decrease recorded in airtight conditions (Figure 1) is an indication that nutrient deterioration may be reduced with proper storage practices. This observation agrees with Jones, (1987)and Lim et al., (2008) who reported that infestation of feed by spoilage microorganisms results in loss of dietary nutritional value owing to loss of amino acids (especially lysine and arginine), dietary lipids, and vitamins.

Lipid content of Coppens^® was higher compared to those observed in Multifeed^® (Figure 2), however, vitalfeed had the least lipid content both in open and airtight conditions of storage. Ayuba and Iorkohol, (2013) also reported differences in lipid content of Coppens^®, Dizengoff, Durante and Adolf calyx fish feeds. Types and kind of feed ingredient used in the feed formulation, as well as feed nutritional specification as determined by the manufacturer, may have led to this variation. The lipid content of Coppens^® and vitalfeed reduced as the storage time increased for open storage condition (Figure 2), however, Multifeed^® slightly varied but was not much pronounced as the other feeds. These trends could be attributed to lipid oxidation at various degrees based on the susceptibility of the lipid in the feed ingredient to deterioration. According to NRC, (1981), feed stored longer than 90 days (three months) at ambient temperature is subjected to the breakdown of oil, and vitamin along with peroxidation of lipid component. Rancidity resulting from lipid oxidation is the most outstanding deteriorative changes in feed during storage. Feed ingredients containing lipids which are highly poly-unsaturated such as fish meals are susceptible to oxidations (Pezzuto and Park, 2002; Sidhuraju and Backer, 2003). Chan, (1987) reported that poly-unsaturated fats can quickly autoxidize at ambient or sub-ambient temperatures. AIN, (1980) however reported that diets containing fish oil are more susceptible to autoxidation than diets containing other polyunsaturated fats. It is an established fact that different variables are involved in oil shelf-life; processing, storage conditions, light exposure, type of packing material, availability of oxygen and addition of antioxidants do affect the quality and characteristics of fats and lipids containing products (Lawson, 1995; Polvilloet al., 2004) this study as justified the fact that storage affects lipid content of feeds. 

The magnitude of oxidative changes was monitored by the periodical measurement of peroxide value (POV), and free fatty acid. The peroxide value levels increased in all diet groups over the storage period and was higher in Multifeed^® while least value was observed in coppen for both storage conditions (Figure 3; Figure 4; Figure 5). This could be attributed to storage time and long exposure to air. Esterbauer et al., (1986; 1991), reported that POV of fish oil diets constantly increased with time exposure to air and under normal feeding conditions. Peroxide formation is likely to occur as susceptible poly-unsaturated fatty acids are available in the oil. Increases in POV are catalyzed by free radicals attacks at sites. This might explain the observed higher rate of dietary peroxide formation attained towards the end of the storage. Fritche and Johnson, (1988), reported an extremely rapid autoxidation of diets with added fish oil as measured by peroxide value rapid oxidation (POV). Rasheed et al., (1963), reported a high oxidation in diets containing fish oil. However, the value of free fatty acid (FFA) increased slowly among the feed under airtight and open conditions throughout the storage period (Figure 6; Figure 7; Figure 8).

The observed notable difference between values of phosphorus, calcium, magnesium, and iron reported by manufacturers and those determined in the laboratory (Table 4) could be attributed to the length of storage before the experiment started, the storage conditions and environmental factors. This is justified by the observed differences noted with different storage conditions for six months. However, the values are within the recommended ranges for the culture of various fishes (Andrews et al., 1973 reported 1.50% of calcium also for I. punctatus, NRC 1983 recommended 0.50 of phosphorus for catfishes while Gatlin et al., 1982 recommended 0.04% of magnesium also for catfishes). The different commercial fish feed stored in open conditions for the period of six months were observed to have physical deterioration after the third month (Table 5), these changes includes; color change, abit soft text of the feed, mouldy growth, unpleasant smell indicating the presence of rancidity and little insect population (Table 6). This is in line with De Silva et al., (1995) that reported improper flavor and appearance changes during storage also feed lump and less palatability was reported.

The aim of feed storage is to reduce the rapidity at which feed deteriorates hence storage never enhances feed quality however proper storage is in an effort to maintain the shelf life of feed. Mold growth in stored feeds has been implicated to reduce nutritional value owing to the loss of dietary lipids, amino acids (especially lysine and arginine) and vitamins by enzymatic digestion (Jones, 1987). Mold infestation was first noticed in vitalfeed under open condition compared to other commercial feed (Table 5; Table 7). Thick mold cover on the surface of the feed was first observed in February and increased till April. This could be attributed to higher relative humidity during the period (66.38 – 71.53%) recorded in that month of the year (Table 8). Hence, Feed under open condition had higher chances of mold infestation compared to feed stored in an airtight condition. This is in accordance with Osuji, (1985) who reported that beyond 70% relative humidity mold infestation is set in. This observation is also in accordance with Cockrell et al., (1971); NRC, (1981); New, (1987); Effiong and Eyo, (1997); and Eyo, (2001). The feed under airtight condition had no such problem except for vitalfeed. Vitafeed^® became damp and musty with less moldy growth. However, this observation came later than the time of mold infestation recorded for other opened fish feeds. This could be explained by the influence of factors such as the combination of the feed ingredients, the chemical composition, and interaction of the feed, also interaction of the above-mentioned factors may as well be the cause. Moreso the above observation may be explained by the high relative humidity created within the closed chambers of the storage system (Vitalfeed) thereby increasing dampness and moisture content of the feed (Figure 9) which in turn created a conducive environment that leads to mold proliferation. However, the inside of the other feed under airtight condition was noticed to be dry throughout the storage period unlike the Vitafeed^®, hence, highlighting the role packaging plays in fish feed preservation. This is in line with the observations of NRC, (1981).

Seven insects' species were identified to infest the feed, they were; Gryllus assimilis, Blatta orientalis, Solenopsis germinate, Delia plutura, Phenicia sericata, Damistid bettle and Larder beetle (Table 6). The occurrence and development of an insect infestation are dependent on many factors such as the source of insect available, temperature, moisture, air condition of the feed and presence of other organisms, (Chow, 1980). In spite of the insect infestation of the feed, no weight loss was observed among the feed stored under airtight and open conditions throughout the storage period. This could be as a result of less number of insect infestations to the feed. An actively reproducing population of insect can quickly consume a significant amount of feed and deteriorate the physical quality (Pederson, 2000).The observed nutrition changes and pathogenic content of stored feed demonstrated in this need to be taken into consideration when feeds are been stored by farmers for long times, the aftermath effect of feeding fish with such feed need to be investigated to provide information on the performance of the potential effect on fish.

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