Animal Science

Partial replacement of raw matters by natural zeolite in Clarias gariepinus feed

0000-0002-6687-8284J. E. Llanes^{1  *}  , 0000-0002-3525-3433M. Castro²  , 0000-0002-2641-1815Magaly Herrera² 

¹Empresa de Desarrollo de Tecnologías Acuícolas. Carretera Central km 20 ½, Loma de Tierra, Cotorro. La Habana, Cuba

²Instituto de Ciencia Animal. Apartado Postal 24, San José de las Lajas. Mayabeque, Cuba

ABSTRACT

The partial replacement of 3, 5 and 7 % of each raw matter by Cuban natural zeolite in formulations of small fish feeds and Clarias gariepinus fattening was evaluated. Two bioassays were carried out, according to a completely random design with three repetitions. In the growth stage, a total of 225 small fish of 1.03 ± 0.03 g of initial weight were used, distributed in three treatments (0, 3 and 5 % of zeolite). Similarly, in fattening, a total of 135 animals of 10.12 ± 0.05 g of initial weight were used in three treatments (0, 5 and 7 %). A decrease in the protein supplied/fish was recorded, when the levels of zeolite in the rations increased. However, the indicators of growth, feed efficiency and survival were not disadvantaged up to 5 % in the small fish feed and 7 % in the fattening feed. The economic analysis showed that the use of zeolite decreases the costs of the rations and contributes with amounts of earnings for feeding. It is concluded that the partial replacement of up to 5 % of the raw matters by natural zeolite in the small fish feed and 7 % in the fattening feed does not compromise the productive performance of Clarias gariepinus; in addition to having a positive economic effect.

Key words: feeding; catfish; mineral; nutrition; zeolite

Natural zeolite is a neutral crystalline microporous aluminosilicate mineral, originating from volcanic rocks. There are more than 45 types, and among them clinoptilolite is the purest, most effective and inexpensive (^{Abdel-Rahim 2017}). Zeolites are used in domestic animal rearing, in industry, in environmental protection, without dismissing their use in modern agriculture, which is due to problems related to the availability of nutrients for crops (^{Díaz et al. 2019}).

In aquaculture, its use has been aimed at improving water quality through the exchange of its mono and divalent ions with toxic wastes, such as ammonium, in water recirculation systems, aquariums and fish transportation tanks (^{Skleničková et al. 2020}). It is also applied to treat effluents and obtain acceptable levels of discharges, as well as to maintain appropriate water quality in fattening ponds (^{Abdel-Rahim 2017} and ^{Martínez et al. 2019}).

In Cuba there is little experience in the use of natural zeolite in fish feeding. ^{Llanes and Castro (2020)} reported the substitution of up to 5 % of feed for zeolite in GIFT Nile tilapia small fish (Oreochromis niloticus), without compromising the productive indicators. However, the literature reports its growth-enhancing effect and feed efficiency in numerous aquatic species (^{El-Gendy et al. 2015} and ^{Senmache and Reyes 2020}), possibly due to the absorption of toxic compounds or the efficiency increase in nutrient assimilation.

The catfish Clarias gariepinus is the main intensively crop species in Cuba. Hence, it demands the largest amount of feed. The growing need to evaluate national inputs to increase their efficiency and reduce conventional raw matters justifies the development of researches, since in intensive fish farming feeding represents the highest costs. The objective of this study was to evaluate the partial replacement of raw matters by natural zeolite in the growing and fattening feeds of Clarias gariepinus.

Materials and Methods

Experimental diets and animal management. The research was carried out in the Fish Nutrition Laboratory from Empresa de Desarrollo de Tecnologías Acuícolas (EDTA) in Havana, Cuba. The enterprise has circular cement tanks, with a capacity of 68 L, with constant water flow (replacement of 100 % daily).

Two experiments were carried out. In the first, in the growing stage, the formulation of catfish small fish feed (D-1) and the substitution of 3 % (D-2) and 5 % (D-3) of each raw matter by natural zeolite was used. In the second, in the same way, the catfish fattening feed (D-4) and the replacement of 5 % (D-5) and 7 % (D-6) were used (table 1). The experimental zeolite came from San Andrés plant, in Holguín, Cuba. The product is marketed under the name Zoad, and has a grain size of less than 0.8 mm.

Table 1 Composition and chemistry of the experimental diets, with different percentages of zeolite for Clarias gariepinus feeding 

Ingredients	Growing			Fattenig
	D-1 0 %	D-2 3 %	D-3 5 %	D-4 0 %	D-5 5 %	D-6 7 %
Fish meal	25.0	24.25	23.75	10.0	9.50	9.30
Soybean meal	36.0	34.92	34.20	45.0	42.75	41.85
Wheat	34.0	32.98	32.30	-	-	-
Corn	-	-	-	39.3	37.34	36.55
Vegetable oil	3.0	2.91	2.85	3.0	2.85	2.79
Dicalcium phosphate	1.0	0.97	0.95	1.7	1.61	1.58
*Vitamin mineral mixture	1.0	0.97	0.95	1.0	0.95	0.93
Natural zeolite	0.0	3.0	5.0	0.0	5.0	7.0
Total	100	100	100	100	100	100
Calculated analysis , %
Dry matter	89.30	88.41	88.52	89.44	88.41	90.12
Crude protein	36.00	34.92	34.21	29.44	27.98	27.42
Ether extract	6.15	5.96	5.84	5.64	5.36	5.24
Crude fiber	3.72	3.61	3.54	4.00	3.80	3.72
Ashes	8.92	11.05	12.87	7.03	11.08	12.94
Calcium	1.54	1.49	1.46	1.01	0.95	0.93
Available phosphorous	0.96	0.94	0.92	0.71	0.67	0.65
Digestible energy , MJ/kg	12.08	11.72	11.48	11.51	10.93	10.70

*Vitamin mineral mixture (kg of diet): vitamin A 500 IU, D 100 IU, E 75 000 mg, K 20 000 mg, B₁ 10 000 mg, B₃ 30 000 mg, B₆ 20 000 mg, B₁₂ 100 mg, D 60 000 mg, niacin 200 000 mg, folic acid 500 mg, biotin 0.235 mg, selenium 0.2 g, iron 80 g, manganese 100 g, zinc 80 g, copper 15 g, potassium chloride 4 g, manganese oxide 0.6 g, sodium bicarbonate 1.5 g, iodine 1.0 g, cobalt 0.25 g.

Preparation of diets. The meals were milled in a Creole hammer mill, at 250 µm, and blended in a mixer (HOBART MC-600^®, Canada). The oil, the natural zeolite and the vitamins and minerals premixture were added. The pelleting was carried out in a meat mill (JAVAR 32, Colombia). The mixture was dried in an oven (Selecta, Spain) at 60 °C for 24 h. A bromatological analysis was performed to the analysis, according to the methods described by ^{AOAC (2016)}. Digestible energy (DE) was calculated with the caloric coefficients reported by ^{Toledo et al. (2015)}.

Growth bioassays. The animals were acclimatized in the experimental facilities one week before starting the research. For the growth bioassay, a total of 225 small fish of 1.03 ± 0.03 g of initial weight were used, randomly distributed in nine tanks (25 animals per tank). The tank was the experimental unit. The diets were offered in broken pellets for the first 15 d, and later in 1 mm particles. The feeding rate was 8 % of the biomass, supplied in two daily rations for 40 d.

For fattening, a total of 135 small fish were used, with an initial weight of 10.12 ± 0.05 g, located in nine tanks (15 fish per tank). The tank constituted the experimental unit. The feeding rate was 6 % of the biomass, added in two daily rations for 60 d.

Every day the values of temperature and dissolved oxygen were taken with a portable digital oximeter (HANNA^®, Romania). At the end of the bioassay, the fish were individually weighed to calculate the following productive indicators:

• Food supplied/ fish = food supplied/ number of final animals

• Protein supplied/ fish = added protein / number of final animals

• Final average weight

• Feed conversion = food supplied / weight gain

• Protein efficiency = weight gain / protein supplied

• Survival = number of final animals / number of initial animals x 100

Statistical analysis. The statistical package InfoStat, version 2012 (^{Di Rienzo et al. 2012}) was used. Where necessary, mean values were compared using ^{Duncan (1955)} test.

An analysis of variance was performed, according to a one - way model. The theoretical assumptions of ANOVA were verified for all the variables based on ^{Shapiro and Wilk (1965)} tests for the normality of errors. ^{Levene (1960)} was applied for the homogeneity of variance. The variables fulfilled with the theoretical assumptions of the ANOVA. A ji-square proportions analysis was used for survival and the ^{Fisher-Yates (1958)} (P<0.05) test was used for comparison.

Economic analysis. It was carried out according to ^{Toledo et al. (2015)} procedure. The costs of the rations were calculated from the international prices of raw matters for November 2021, reported by ^{Indexmundi (2021)} (table 2). To the results was added 45 % of the total cost of raw matters for additional expenses (transportation, maquila and administrative) for Cuba. These figures were multiplied by the feed conversion values obtained in this study to determine feeding costs.

Table 2 Prices of raw matters used in the experimental diets, $US/t 

Raw matters	Prices, $ US /t
Fish meal	1 442.75
Soybean meal	442.42
Wheat	317.44
Corn	248.72
Vegetable oil	1 439.57
Dicalcium phosphate	533.24
Vitamin - mineral mixture	1 975.11
Natural zeolite	70.00

Results and Discussion

During the experimental period, water circulation was efficiently controlled to guarantee 100 % of daily turnover. The temperature and dissolved oxygen in the water from the tanks ranged from 26.8 to 28.1 ^oC and from 5.54 to 5.96 mg/L, respectively. These values are considered comfort values for the good productive performance of the species (^{Toledo et al. 2015}).

Rapid intake of the experimental diets by the animals was observed, suggesting that the palatability of the rations was not affected by the zeolite levels. Similarly, the pellets had good physical constitution and hydrostability.

^{Abdel-Rahim (2017)} reported that zeolite increases palatability and has a binding effect, which contributes to improving food stability and reducing wastes. In addition, the author reported that it is used in artificial foods to reduce the toxic effects of aflatoxins and microbial agents that spoil food.

In the growth stage, the inclusion of 5 % of zeolite decreased the supplied protein/fish and disadvantaged feed conversion. On the contrary, the food supplied/fish, the final weights and the protein efficiency were not affected with the same level of inclusion (table 3).

Tabla 3 Productive performance in the growing stage of Clarias gariepinus with the experimental diets 

Indicators	D-1 0 %	D-2 3 %	D-3 5 %	± SE	P
Food supplied/fish, g	13.60	14.26	13.49	0.17	0.124
Protein supplied / fish, g	4.90^a	4.98^a	4.61^b	0.07	0.046
Final weights, g	16.83 ± 0.47	17.45 ± 0.54	15.97 ± 0.49	-	0.104
Feed conversion	0.86^a	0.86^a	0.91^b	0.01	0.017
Protein efficiency	3.23	3.29	3.24	0.02	0.422

Different letters in the same row differ at P<0.05, according to ^{Duncan (1955)}

There was similar performance in fattening (table 4), a stage in which the increase in zeolite reduced the protein supplied/fish and, although the feed conversion did not has differences, it tended to deteriorate (P=0.076). Therefore, a longer culture time is required to observe the effect of the highest concentration (7 %), once the fattening is six or seven months. These results show the efficiency of using natural zeolite in catfish feed.

Table 4 Productive performance in the fattening of Clarias gariepinus with the experimental diets 

Indicators	D-4 0 %	D-5 5 %	D-6 7 %	± SE	P
Food supplied/fish, g	75.01	73.13	75.02	0.23	0.497
Protein supplied/fish, g	22.07ª	20.48 ^b	20.42 ^b	0.11	0.028
Final weights, g	69.33 ± 2.43	68.69 ± 2.47	66.92 ± 2.11	-	0.668
Feed conversion	1.33	1.35	1.40	0.01	0.076
Protein efficiency	2.56	2.58	2.59	0.01	0.810

Different letters in the same row differ at P<0.05, according to ^{Duncan (1955)}

The amount of food supplied/fish was similar, regardless of the zeolite inclusion levels, because feeding was restricted by body weight, and these were similar between treatments during culture. Regarding the supplied protein, a decrease was recorded when increasing the zeolite levels, due to a dilution effect that reduces its concentration on the diets.

The final weights of Clarias gariepinus small fish and young fish, fed with natural zeolite and without it, were similar (tables 3 and 4). In studies with red tilapia Oreochromis sp. (^{Rafiee and Saad 2005}), Nile tilapias GIFT (^{Llanes and Castro 2020}) and river shrimp Cryphiops caementarius (^{Senmache and Reyes 2020}) there were no significant effect of this mineral on the animals growth. In contrast, its favorable action is reported on Nile tilapias (^{El-Gendy et al. 2015}), snakehead fish Channa striatus (^{Jawahar et al. 2016}), rainbow trout Oncorhynchus mykiss (^{Sheikhzadeh et al. 2017}) and red tilapia (^{Zain et al. 2018}). The reason for these differences could be given by the species, types and properties of the zeolites, as well as by the inclusion levels in the ration.

Regarding feed efficiency indicators, the increase of zeolite in the ration tends to worse feed conversion, since the mineral is not absorbed and forms part of the volume of fecal matter. However, the protein efficiency values (tables 3 and 4) showed the high efficiency of the zeolite in the metabolic use of nitrogen, as it did not disfavor weight gain and survival with lower protein concentration. The efficient use of dietary protein depends on the content and balance of aminoacids (AA), and is important for optimal fish growth (^{Rodríguez-Avella et al. 2019}). Therefore, it is evident that zeolite did not alter the nutrient balance, and can contribute to formulations that improve productive performance and health, and that are also friendly to the environment.

^{Castro et al. (2019)} reported the advantages of using zeolite in monogastric animals feeding, by slowing down the speed of chyme through the digestive tract. Cation exchange channels trap nutrients, such as nitrogen, for slower release in the intestines. In addition, its high silicon content favors the constant scraping of the intestinal villi, where those that are worn are removed, and the formation of new villi is stimulated. These conditions favor the absorption process and the most efficient use of dietary nutrients.

The literature refers to other studies on the use of zeolite in fish feeding. ^{El-Gendy et al. (2015)} evaluated different feeding rates (2, 2.5 and 3 % of body weight) in Nile tilapia at a production scale, including 2 % zeolite in the feed (25 % protein), and found the best indicators of water quality, productive and economic with 2.5 %. ^{Sheikhzadeh et al. (2017)} showed that 5 g/kg of zeolite had a potential effect on growth, digestive enzyme activity and various biochemical indices in rainbow trout. ^{Zain et al. (2018)} reported that the addition of zeolite was essential to improve water quality and growth indicators of red tilapia. This shows that an optimal concentration of dietary zeolite allows better use of diet nutrients and improves the productive performance of animals.

The survival of the fish did not show differences when increasing the zeolite levels (table 5), which shows that the use of this mineral does not compromise the animals health. ^{Jawahar et al. (2016)} reported improvements in the immune response and resistance to diseases caused by Aphanomyces invadans in snakehead fish. ^{Martinez et al. (2019)} reported that zeolite retains the ammoniacal nitrogen emitted in urine and feces to improve water quality, which is decisive for the vitality of fish. The values of this study were similar (> 92.0 %) to those reported for Nile tilapia (^{El-Gendy et al. 2015}) and Nile tilapia GIFT (^{Llanes and Castro 2020}).

Tabla 5 Results of the survival of small fish of Clarias gariepinus fed with experimental diets in the growth and fattening period 

Stages	D1 60 % HP		D2 51 % HP		D3 42 % HP		>SE ±	P
	No.	%	No.	%	No.	%
Wrowing	74	98.67	71	94.67	73	97.33	2.00	0.313
Fattening	45	100	45	100	44	97.78	1.78	0.422

The economic analysis showed that the increase in natural zeolite decreases the costs of the rations due to its low price compared to the raw matters that make up commercial feeds (table 6). In the growing stage, the results showed savings with 3 % inclusion, once the ration cost decreased and feed conversion was not disadvantaged. However, with 5 % there was an economic loss because 70 kg of more food for every 100,000 small fish is needed.

Table 6 Economic analysis of C. gariepinus production with the inclusion of natural zeolite in small fish and fattening feed, US $/t 

Indicators	Growing			Fattening
	D-1 0 %	D-2 3 %	D-3 5 %	D-4 0 %	D-5 5 %	D-6 7 %
Ration cost	1 009.43	983.44	964.03	744.02	711.87	699.04
Feeding cost	868.11	845.76	877.27	989.54	961.02	978.65
Monetary savings	-	22.35	-9.16	-	28.52	10.89

Feeding cost = Ration cost x feed conversion

In fattening, higher amounts of profits were obtained, as the levels of substitution of raw matters by zeolite were higher. The results showed savings for 5 and 7 %. However, with 5 % it was 2.6 times higher due to a better feed conversion. It is important highlighted that fattening is the stage of greatest demand of food, where a significant amount of the budget is still used to import raw matters to guarantee food sovereignty. The results of this study confirm those obtained by ^{El-Gendy et al. (2015)}, ^{Abdel-Rahim (2017)} and ^{Llanes and Castro (2020)}, who reported that the use of zeolite contributes to lower feeding costs.

The use of natural zeolite introduces important elements of efficiency in the performance of catfish, although more studies are needed to determine the effects on the digestive physiology of small fish and young catfish. The inclusion of mineral in the feed formulas replaces, in the same proportion, the diets components, and it is not necessary to correct the formula in terms of nutrient levels The results show that the partial replacement of 3 % of the raw matters by natural zeolite in the small fish feed and 5 % in the fattening feed does not compromise the productive performance of Clarias gariepinus and has a positive economic effect.

Acknowledgments

Thanks to the Centro Nacional de Producción de Animales de Laboratorio (CENPALAB) for the support provided in the acquisition of raw matters for the preparation of experimental diets.

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Received: February 25, 2022; Accepted: April 30, 2022

^*Email: [e-mail]

Conflict of interests: The authors declare that there is no conflict of interest between them.

Author’s contribution: J. E. Llanes: Conceptualization, Investigation, Methodology, Writing - original draft. M. Castro: Conceptualization, Methodology. Magaly Herrera: Formal analysis