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The search for alternative foods through unconventional raw materials, which largely allow import substitution, reduce competitiveness with human food and preserve the environment, has been one of the objectives set in Cuba for more than three decades. Royal palm, Roystonea regia (Kunth) O. F. Cook, is the most abundant plant from Arecaceae family in Cuba. Due to its properties, it has multiple uses: ornamental, industrial, medicinal and nutritional. It is also a provider of different products of economic importance, among which is royal palm nut.
The chemical composition of this fruit makes it to be considered as a good energy source, due to its high lipid content and, to a lesser extent, to its high fiber content (Oliva et al. 2018). Within its oil, oleic, lauric, palmitic and linoleic acids are predominant, containing 8 and 18 carbon atoms, mainly responsible for its energy source status. Royal palm nuts are one of the main alternative energy foods available in Cuba, where it is popular as an unconventional source of feed for pigs (Ly et al. 2017), but without concrete results in broilers.
As a high lipid source, to be digested in the gastrointestinal tract of broilers, royal palm nut needs lipolytic enzymes. Among them, pancreatic lipase is one of the most studied. This enzyme can be used as a digestion marker of this fraction. Therefore, the objective of this study was to determine the enzymatic activity of pancreatic lipase in broilers consuming Roystonea regia fruit meal in the ration.
An amount of 40 male broilers, eight days old (HE21), housed in metal cages, was used. Animals received experimental diets, from 8 d to 42 d. These were formulated according to requirements established by NRC (1994) for this category. Control consisted of a conventional soy bean-corn diet. In the other three treatments, 5, 10 and 15 % royal palm nut meal was included, so that the isoproteic and isoenergetic diets were maintained for beginning, growth and finish periods. During all the experimentation time, broilers had free access to water and food.
Royal palm nut, in natura, was obtained from Mayabeque province. Drying process consisted of detaching royal palm nuts manually, and then expose them under the sun, spread on a drying plate, for five consecutive days.
Dry material was stored in bags, indoors, in buildings with adequate ventilation and low humidity level. Every five days, it was ground with the rest of the components of grain and cereal diet in the corresponding proportion, with the purpose of extending the shelf life of royal palm nut meal that becomes rapidly rancid, and decreases its biological quality and palatability (Oliva et al. 2018).
At 42 d of the experiment, ten broilers were weighed and slaughtered per treatment, according to the jugular vein bleeding method. For this, the traditional procedures (stunning, bleeding and evisceration) were followed, exactly two hours and thirty minutes after the animals had ingested food, with the purpose of taking pancreas samples and obtaining the pancreatic homogenate. Immediately, pancreas was removed from each broiler, and immersed in 4 mL/g of physiological saline, between 0 and 4 °C, for further transfer to the laboratory. The organs of three animals were manually homogenized. They were centrifuged at 5,000 rpm. for 10 min at 4 °C in a Thermoscientific IEC CL31R refrigerated centrifuge. The pellet was discarded and the supernatant was stored in small aliquots at -80 °C.
Protein concentration was determined by the method of Bradford (1976) on a UV-vis Rigol spectrophotometer, Ultra-3400 series. Concentrations were calculated by interpolating absorbance values of samples in the standard curve of bovine serum albumin (BSA) (0.01- 5 mg/mL). This was standardized for all homogenates under study at 3mg/mL, for which the corresponding dilutions were made in each case. All analyzes were performed in triplicate.
To achieve maximum reliability in enzymatic activity determination, the authors worked on the linear range proposed for the enzyme, for which an incubation time of five minutes was used (León et al. 2014). Lipase activity was determined by the method proposed by Tietz and Fiereck (1966). The necessary volume of NaOH (50 mM) was determined, to neutralize released fatty acids, after enzymatic degradation. The system was kept under moderate agitation at 37 °C and 3 mL of olive oil were used as substrate. Enzyme activity was expressed as mL of NaOH/min/mg of protein.
For the statistical analysis of results, a completely randomized design was used, with four treatments and three repetitions (three pancreases each). Differences between means were determined according to Duncan (1955) test in the necessary cases. Infostat (di Rienzo et al. 2012) statistical package was used.
Significant effect of the inclusion of royal palm nut meal on the diet was observed, specifically on the activity of pancreatic lipase enzyme in broilers (figure 1). Treatment with the inclusion of 15 % royal palm nut meal increased significantly enzyme activity, compared to the rest of treatments. Treatments with 5 and 10 % did not differ from each other. Control treatment showed the lowest activity (P <0.05). This performance is probably associated with fat content in the diet, since it is known that type, origin and level of fat in the diet are important factors that affect secretion of lipolytic enzymes by pancreas (Barberá 2016).
Figure 1 Enzymatic activity of pancreatic lipase in broilers consuming different levels of royal palm nut meal
According to Gonçalves et al. (2010), nutritional factors, such as secondary metabolites, can also modify pancreatic lipase activity. These authors stated that absence of polymeric flavonoids of high molecular weight, which have been attributed the action of inhibiting pancreatic lipase activity, when exercising a specific interaction with the enzyme, are usually present in fruits of intense colors such as grapes. Although royal palm nuts show a blackish brown coloration, in the qualitative phytochemical screening performed on the meal of this fruit, the presence of secondary metabolites (Ly et al. 2017) was discarded, which would explain the results.
Barberá (2016) showed that changes in diet composition, involving different sources of lipids and fatty acids, can cause changes in lipase enzyme activity. While it is important to consider that pancreatic secretions respond to a food stimulus, the activity of these enzymes, once they enter the intestinal mucosa, can also be affected in broilers (León et al. 2014), as observed in the present study.
Increased activity of pancreatic lipase could increase lipid digestion, which may be beneficial for the animal, since it is known that these biomolecules perform several important biological functions in the body. It can be mentioned that, among these functions, they constitute structural components of membranes, as well as transportation means and storage of catabolic fuel. They also serve as a protective covering on cell surface, which is related to cell-cell recognition, species specificity and tissue immunity. They also have an intense biological activity, since some vitamins (A, B, D, E and K) and hormones are among them.
It is concluded that the inclusion of royal palm nut meal in diets for broilers increases the activity of pancreatic lipase enzyme, which makes it possible to digest lipids in the gastrointestinal tract.
