Oxidative stability of tilapia feeds containing Saccharomyces cerevisiae and Spirulina platensis

Estabilidade oxidativa de rações de tilápia contendo Saccharomyces cerevisiae e Spirulina platensis

ABSTRACT:

The aim of this study was to evaluate the effect of the inclusion of microbial biomass on the oxidative rancidity of tilapia rations stored for 12 months. Treatments included a control diet and diets supplemented with either 0.01% vitamin E, 0.25 and 0.5% of Saccharomyces cerevisiae and 0.25 and 0.5% of Spirulina platensis. Experimental diets were stored in the dark inside plastic bags at room temperature (25 °C) for 12 months. The oxidative rancidity was measured as thiobarbituric acid reactive substances (TBARS). It was concluded that the inclusions of Spirulina platensis at 0.25% (1.734 ± 0.206) and 0.5% (1.629 ± 0.181) and Saccharomyces cerevisiae at 0.5% (1.459 ± 0.305) minimized the oxidative rancidity in comparation to control diet (2.843 ± 0.109) of Nile tilapia until 12 months of storage.

Key words:
biomass; lipid oxidation; ratio; TBARS.

RESUMO:

O objetivo deste estudo foi avaliar o efeito da inclusão de biomassa microbiana sobre a rancidez oxidativa de rações de tilápia armazenadas por 12 meses. Os tratamentos incluíram uma dieta controle e dietas suplementadas com 0,01% de vitamina E 0,01%, com 0,25 e 0,5% de S. cerevisiae e com 0,25 e 0,5% de S. platensis. As dietas experimentais foram armazenadas em sacos plásticos escuros, em temperatura ambiente (25 °C), durante 12 meses. A rancidez oxidativa foi mensurada pelas substâncias reativas ao ácido tiobarbitúrico (TBARS). Concluiu-se que as inclusões de 0,25% (1,734 ± 0,206) e 0,5% (1.629 ± 0.181) de Spirulina platensis e 0,5% (1.459 ± 0.305) de Saccharomyces cerevisiae reduziram a rancidez das dietas em comparação a dieta controle de tilápias-do-Nilo após 12 meses de armazenamento.

Palavras-chave:
biomassa; oxidação lipídica; ração; TBARS

Oxidative rancidity or lipid oxidation refers to the lipid deterioration featured by the formation of chemicals that change food aroma and taste, decreasing palatability and nutritional value of diets (LAGUERRE et al., 2007). The process consists of a complex chain of reactions induced by oxygen in the presence of catalysts such as heat, free radicals, light, pigments and metallic ions (LAGUERRE et al., 2007). The first stage of the chain (initiation phase) occurs with the formation of free radicals under conditions favored by light and heat and is followed by the formation of primary products (propagation phase), especially peroxides and hydroperoxides. The latter, when exposed to prolonged oxidation conditions, give rise to side products like aldehydes, ketones, epoxides, hydroxy compounds, oligomers and polymers (termination phase). Malonaldehyde (MDA) is one of the main aldehydes originated in the process and commonly used as a marker to quantify the reaction (BARRIUSO et al., 2013).

The stability of animal feed during storage is associated with lipid oxidation and so explains the use of antioxidants to slow down the process, increase the shelf life of the product, reduce the waste of raw materials and increase the range of fats that can be used in the formulation of diets, especially those of high energy (AKLAKUR, 2018).

Synthetic vitamin E may be used to prevent rancidity in fish feeds; although, butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), tert-butyl hydroquinone (TBHQ), propyl gallate (PG) and ethoxyquinone (EQ) are the most widely used sources of additives for that purpose (AKLAKUR, 2018). However, these latter compounds may pose a risk to the health of animals and consumers since toxic and carcinogenic effects are assigned to them (PONCE-PALAFOX et al., 2004), thus giving rise to a growing demand for antioxidants of natural origin. Two microbial biomasses (MB) with antioxidant potential stand out as fish feed ingredients. Saccharomyces cerevisiae yeast is used in the preparation of various products (e.g. ethanol) and marketed as a co-product at the end of processes. The yeast biomass contains proteins, vitamins such as E and complex B, enzymes, volatile fatty acids, chelated minerals, natural antibiotics and peptides (RAN et al., 2016). Spirulina (Arthrospira) platensis biomass contains proteins, vitamins, essential amino acids, minerals, essential fatty acids and pigments, such as carotenoids and phycocyanins, which impart it high nutritional value and antioxidant properties (MORIST et al., 2001; LI et al., 2003). The microalga is able to grow on different substrates, including industrial effluents and simple and low-cost culture media, making its production totally viable (ARRUDA et al., 2006).

The purpose of this study was to evaluate the effect of microbial biomasses on the oxidative rancidity of tilapia rations stored for 12 months.

The study adopted a completely randomized design with six treatments and three replicates. Treatments included a control basal diet (CD) and five diets supplemented with either 0.01% vitamin E (VE), or S. cerevisiae at 0.25% (SC25) and 0.5% (SC50) or S. platensis at 0.25% (SP25) and 0.5% (SP50). The calculated and analyzed values of the nutrients in the diets are presented in table 1. The proximate composition of the biomasses were: 4% moisture, 45% protein, 0.2% lipid and 5% ash for S. cerevisiae; and 6% moisture, 46% protein, 3% lipid and 8% ash for S. platensis. The biomasses were dissolved in soy oil and incorporated to the basal diet inside a Y-shaped mixer. The same amount of soy oil was added to the control group. The experimental diets were stored in the dark inside plastic bags (polypropylene) at room temperature (25 °C) for 12 months.

Table 1
Experimental diet for tilapias.

Ingredients (%)	CD	VE	SC25	SC50	SP25	SP50
Feather meal	2	2	2	2	2	2
Ground corn	33.06	33.05	33.06	33.06	33.06	33.06
Poultry meal by-products	6	6	6	6	6	6
Soybean meal	10.44	10.44	10.19	9.94	10.19	9.94
Wheat meal	15.88	15.88	15.88	15.88	15.88	15.88
Meat meal	17.14	17.14	17.14	17.14	17.14	17.14
Blood meal	12	12	12	12	12	12
NaCl	0.8	0.8	0.8	0.8	0.8	0.8
Chicken oil	2	2	2	2	2	2
Premixa	0.6	0.6	0.6	0.6	0.6	0.6
Methionine	0.08	0.08	0.08	0.08	0.08	0.08
Vitamin E	0	0.01	0	0	0	0
S. cerevisiae	0	0	0.25	0.5	0	0
S. platensis	0	0	0	0	0.25	0.5
Total	100	100	100	100	100	100
--------------------------------------------------------------------Nutrients/Energy - calculated values-----------------------------------------------------
Digestible energy (kcal/kg)	3000	3000	3000	3000	3000	3000
Digestible protein (%)	27.63	27.63	27.63	27.63	27.63	27.63
Ether extract (%)	7.50	7.50	7.50	7.50	7.50	7.50
Crude fiber (%)	3.00	3.00	3.00	3.00	3.00	3.00
Mineral composition (%)	10.48	10.48	10.48	10.48	10.48	10.48
Total calcium (%)	2.74	2.74	2.74	2.74	2.74	2.74
Total phosphorus (%)	1.57	1.57	1.57	1.57	1.57	1.57
Starch (%)	27.03	27.03	27.03	27.03	27.03	27.03
Available phosphorus (%)	0.69	0.69	0.69	0.69	0.69	0.69
Arginine (%)	1.80	1.80	1.80	1.80	1.80	1.80
Lysine (%)	2.33	2.33	2.33	2.33	2.33	2.33
Threonine (%)	1.25	1.25	1.25	1.25	1.25	1.25
Tryptophan (%)	0.34	0.34	0.34	0.34	0.34	0.34
Methionine (%)	0.45	0.45	0.45	0.45	0.45	0.45
Vitamin C (mg/kg)	700	700	700	700	700	700
-------------------------------------------------------------------------Nutrients - analyzed values (%)------------------------------------------------------
Protein	32.29	32.18	32.26	32.22	32.41	32.34
Ether extract	8.11	8.15	8.14	8.11	8.18	8.09
Ash	9.79	9.84	9.68	9.82	9.69	9.84
Crude fiber	3.62	3.65	3.57	3.72	3.62	3.59
Moisture	10.75	10.63	10.65	10.85	10.93	10.86

• CD - control diet, VE - CD + 0.01% vitamin E, SC25 - CD + 0.25% S. cerevisiae, SC50 - CD + 0.5% S. cerevisiae, SP25 - CD + 0.25% S. platensis and SP50 - CD + 0.5% S. platensis. ^a Composition per kg of product - Choline 83,333 mg, vitamin A 2,083 UI, vitamin D3 500 UI, vitamin E 37,500 mg, vitamin K3 2,500 mg, vitamin B1 4,167 mg, vitamin B2 4,167 mg, vitamin B6 3,333 mg, vitamin B12 7,500 g, niacin 12,500 mg, calcium pantothenate 8,152 mg, pantotenic acid 7,500 mg, folic acid 833 mg, biotin 125 mg, vitamin C 116,667 mg, inositol 12,500 mg, iron specific source 12,500 mg, Fe 12,500 mg, Cu 2,500 mg, Mn 7,500 mg, Zn 16,667 mg, Co 42 mg, I 125 mg, Se 67 mg.

For the thiobarbituric acid reactive substances (TBARS) quantification, 1 g of feed was homogenized with 25 ml of 7.5% trichloroacetic acid using turrax for 1 min (TBARS extraction). Then, the mixture was filtered and 5 ml of the extract were transferred to a tube containing 5 ml 0.02 M thiobarbituric acid. The tubes were heated in boiling water bath (100 °C) for 40 min and cooled in running water for 10 min. The TBARS were measured at 538 nm and the values were expressed as mg malonaldehyde/kg (GRASSI et al., 2016, 2018).

All data collected were subjected to analysis of variance and the significance of differences between means was tested by Tukey’s test, using Action version 3.1^®. Broken line regression was used to estimate the biomass requirements to avoid the oxidative rancidity as a function of MB inclusion, using PPM (Practical Program for Modeling, https://sites.google.com/site/programapraticodemodelagem/). The significance level was set at 0.05.

The SC50, SP25 and SP50 had lower TBARS than the control group while SC25 and VE were not able to prevent the lipid oxidation during the storage for 12 months (Table 2). The antioxidant activity of biomass minimized the oxidative rancidity, increasing the shelflife of Nile tilapia feed. This effect may be assigned to carotenoids and phycocyanins in Spirulina platensis and to rutin, hesperidin, sakuranetin, quercetin, naringin, naringenin and vitamin E in Saccharomyces cerevisiae (SOARES et al., 2005). The inclusion of S. cerevisiae at 0.25% (SC25) had insufficient antioxidant activity to reduce the oxidative rancidity. The broken line regression indicated that 0.4392% S. cerevisiae (Figure. 1.A) and 0.2852% S. platensis (Figure. 1.B) biomasses in the rations are enough to provide the lowest TBARS in the diets.

Table 2
Oxidative rancidity of feed containing microbial biomass.

Treatment	TBARS* (mg malonaldehyde kg-1)
CD	2.483 ± 0.109a
VE	1.941 ± 0.158ab
SC25	1.900 ± 0.019ab
SC50	1.459 ± 0.305b
SP25	1.734 ± 0.206b
SP50	1.629 ± 0.181b
P	0.0155
CV (%)	19.6

• ^*Thiobarbituric acid reactive substances; CD - control diet, VE - CD + 0.01% vitamin E, SC25 - CD + 0.25% S. cerevisiae, SC50 - CD + 0.5% S. cerevisiae, SP25 - CD + 0.25% S. platensis and SP50 - CD + 0.5% S. platensis. CV - coefficient of variation; ^ab Means followed by different letters are significantly different (P<0.05).

Figure 1
Fitted broken-line plot of oxidative rancidity as a function of S. cerevisiae (A) and S. platensis (B) inclusion in the 12 months old diets.

GRASSI et al. (2016) also reported a delay in tilapia diets stored until 12 months when a bacterial biomass (Rubrivivax gelatinosus) was added up to 0.14% and assigned the effect to the presence of carotenoids in the biomass.

The need to seek alternatives to the synthetic antioxidant substances has increased the interest for the production of natural antioxidants through biotechnology. For instance, microbial biomasses of yeast Phaffia rhodozyma and microalgae Haematococcus pluvialis and Dunaliella salina have been used as sources of carotenoids in the feed of aquatic animals (PONCE-PALAFOX et al., 2004). Vitamin E produced by microorganisms also has been used in fish diets to prevent the oxidative rancidity and protect the tissue integrity of several fish species (PENG et al., 2009). Although, the antioxidant activity of vitamin E in feed is recognized, in this experiment, it was not able to prevent the lipid oxidation like the biomasses did.

More than benefits to the preservation of the ration, S. platensis and S. cerevisiae biomasses can improve fish health and meat quality. GRASSI et al. (2018) included microbial biomasses in tilapia diets and reported an improvement in the oxidative status of the animals and a delay in the fillets oxidation up to 60 days of storage. Moreover, S. platensis biomass increased redness and carotenoids in the tilapia fillets, so improving sensory and functional properties of the flesh (GRASSI et al., 2018).

Results presented herein indicated that S. cerevisiae and S. platensis biomasses can be considered as natural alternatives to replace the synthetic antioxidants currently in use. It was concluded that the inclusions of Spirulina platensis at 0.25 and 0.5% and Saccharomyces cerevisiae at 0.5% minimized the oxidative rancidity in Nile tilapia diets until 12 months of storage

ACKNOWLEDGEMENTS

Authors thank CAPES (88881.134883/2016-01) for the scholarship and Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) for the financial support (2015/25853-1) and scholarship (2015/21216-7).

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• CR-2019-0344.R2

Publication Dates

• Publication in this collection
  13 Dec 2019
• Date of issue
  2020

History

• Received
  04 May 2019
• Accepted
  01 Nov 2019
• Reviewed
  27 Nov 2019