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Food science

Protein enrichment of pineapple waste for animal feeds
Bioconversion of vegetable, animal, and industrial wastes by means of Fungi mycelia in an artificial rumen

Protein enrichment of pineapple waste for animal feeds

N. B. Guerra, T. M. M. Stamford, R. B. de Medeiros, C. P. de Freitas, S. R. Maia, and M. L. Cavalcante
Laboratory of Food Sciences, Department of Nutrition, Health Science Centre, Federal University of Pernambuco, Recife, Brazil


The utilization of fungi for protein production has been studied for years. The efficiency of fungi has been shown in substrates like lignin, cellulose and hemicellulose -polymers found in fruit wastes [1-6]. Stamford et al. have used passion-fruit waste for microbial protein production [7], as Senez et al. have used potato and banana wastes [8].

Considering that substantial amounts of waste products are available free or sold at very low prices by local agroindustries, the upgrading of wastes from fermented fruits for use in balanced feeds for poultry is potentially advantageous.

The purpose of this study was to investigate the conversion efficiency of pineapple-waste carbohydrates into single-cell protein (SCP).

Materials and methods

Pineapple wastes consisting of flesh alone as well as flesh and skin were obtained from Indústrias Maguary S.A., Bonito, Pernambuco, Brazil. The chemical composition of the wastes is given in table 1. The fungi used, Aspergillus niger 2228, Myrothecium verrucaria 2100, and Trichoderma viride 2596, were obtained from the culture collection of the Mycology Department, Federal University of Pernambuco. The strains were maintained at 5°C in a SABOURAUD-agar medium and reinoculated every month. The basic culture medium was Czapek's without saccharose, and, in the experiments with only partial salt addition, the KCl and FeSO4. were removed. Two processes were used: submerged liquid fermentation and moist-solid-state fermentation.

TABLE 1. Chemical composition of pineapple waste



Flesh Flesh + skin
pH 3.2 4.1
Humidity (g/100 g) 85.61 90.50
Ash (g/100g) 0.38 0.63
Protein (g/100 g) 0.42 0.60
Ether extract (g/100 g) 0.17 0.30
Fibre (g/100 g) 1.78 1.83
Carbohydrates (g/ 100 g)a 11.64 6.14
Dry matter (g/100g) 14.39 9.50
Magnesium (mg/100 g) 43.77 72.96
Calcium (mg/100 g) 40.10 -
Phosphorus (mg/100 g) 30.80 -

a. Estimated by difference.

In the preliminary assays for the submerged liquid fermentation process, twenty-seven 250 ml flasks were used, three for each treatment. Each flask contained 100 ml of the medium: 30 g of pineapple waste (flesh, flesh + skin) and 70 ml of distilled water added or not with salts. The flasks were autoclaved at 121.5 °C for 15 minutes, inoculated with a 2 per cent spore suspension, and incubated in a shaker at 30 at 100 rpm for 48, 72, and 96 hours. The filtered mycelia were dried in a 60 °C oven with air circulation until a constant weight.

Assays were performed for biomass production at a semipilot level using a 10-litre capacity Chemap fermentator and at a pilot level using a 200 kg capacity Thermo-Frigor fermentator. The mycelia were collected and dried as described previously.

With the moist-solid-state fermentation method, the pineapple waste (flesh and skin) was heated at 100 under fluent steam as a culture medium for growth of the A niger and M. verrucaria, with or without the addition of KCl and FeSO4. The waste was spread on nylon sieves in wooden frames. The uniform layers were 1 cm thick to allow sufficient aeration. The fungi were inoculated with the spore suspension, and the trays were put on open racks at room temperature, some for 48 and some for 72 hours. A part of the material was then dried in an oven with air circulation at 50 °C or 55 °C for 48 hours.

TABLE 2. Protein biomass production from fungi grown in pineapple waste (flesh) using submerged liquid fermentations

Treatment Micro-organisms Fermentation periodb
72 hours 96 hours
Dry matter Total protein Added protein Dry matter Total protein Added protein
(%) (%) (%) (%) (%1) (%)
Waste + H2O A. night 1.06 8.00 174.9 1.09 17.82 512.37
M. verrucaria 1.41 8.75 200.7 0.85 12.32 323.36
T. viride 1.15 10.18 249.8 1.51 9.45 224.47
Waste +complete
A. nigor 1.50 18.11 522.3 1.57 14.61 402.06
M. verrucario 1.45 12.18 317.46 1.48 16.76 475.94
T. viride 1.15 13.17 352.57 1.56 16.15 454.9
Waste with no KCl A. Niger 1.43 20.25 595.87 1.51 18.13 523.02
and FeSO4 added M. verrucaria 1.76 15.47 420.27 1.07 19.68 575.28
to medium T. viride 1.09 11.89 308.59 1.72 14.91 412.37

a. Pineapple waste has 2.91 g/100 g protein (dry matter).
b. No data were collected at 48 hours of incubation using the submerged liquid fermentation method because the slow growth of the fungi made a determination of protein added by the process impossible.

TABLE 3. Essential amino-acid levels of corn, sorghum and pineapple waste (flesh) treated with submerged liquid fermentation and nutritional requirements of poultry

Amino acids g/100 g sample
A. nigera M. verrucariab Cornc Sorghumc Layingd Chickened
Isoleucine 0.39 0.40 0.37 0.60 0.86 0.5
Leucine 0.60 0.56 n.d. n.d. 1.60 1.2
Lysine 0.40 0.60 0.22 0.27 1.25 0.5
Methionine 0.09 0.05 0.17 0.10 0.86 0.53
Phenylalanine 0.38 0.35 0.44 0.46 1.50 0.25
Threonine 0.43 0.34 0.34 0.27 0.80 0.40
Tryptophan 0.d. 0.d. 0.09 0.09 0.23 0.11
Valine 0.37 0.47 0.42 0.53 1.00  

a. Treated with incomplete salts at 96 hr of incubation.
b. Treated with incomplete salts at 72 hr of incubation.
c. L. G. A. Alves, "Estudos sobre o emprego do sorgo granifero em dietas pare frangos de corte," unpublished M.Sc. thesis (Santa Maria, R S., 1975).
d. Consejo Nacional de investigaciones, Comisión de Nutrición Animal, Subcomisión pare Avea, Necesidodes nutritivas de les ares de corral (Buenos Airea, Editorial Hemisferio Sur, 1975).

Total nitrogen was measured in the mycelia by the micro-Kjeldahl method [9]. The crude protein content of the mycelia was calculated by the factor 6.25 x n. The amino acids were measured in a Beckman auto-analyser reaction with nihydrine.

Results and discussion

The data from the preliminary assays with pineapple waste (flesh) using the submerged liquid fermentation process are shown in table 2. The highest content of crude protein was detected in the A. niger assay without KCl and FeSO4 at 72 hours of incubation and in the M. verrucaria assay without KCl and FeSO4 at 96 hours of incubation. These values were higher than those obtained by Stamford et al. in passion-fruit waste [7]. The highest amino-acid values were obtained with the M. verrucaria at 72 hours of incubation when the lysine content was 0.6 per cent, that is, a higher value than that shown by A. niger grown on corn and sorghum.

TABLE 4. Protein biomass production from fungi grown in pineapple waste (flesh + skin) using submerged liquid fermentationa

Treatment Micro-organisms Fermentation periodb
72 hours 96 hours
Dry matter (%) Total protein (%) Added protein (%) Dry matter (%) Total protein (%) Added protein
Waste + A. niger 2.18 9.62 53.18 1.75 6.84 8.92
H2O M. verruceris 2.03 5.67 -9.71 1.93 5.39 -0.14
T. viride 1.84 7.70 22.61 2.24 7.86 25.15
Waste + A. niger 1.67 12.69 102.07 1.79 12.55 99.84
complete M. verrucori. 1.80 6.64 5.73 2.44 10.10 60.82
salts T. virido 1.73 8.89 41.56 2.19 11.91 89.64
Waste with A. niger 1.68 12.09 92.51 1.80 10.85 72.77
salt M. vorrucorio 2.09 6.66 6.05 2.19 7.03 11.94
reduction T. virido 2.22 10.96 74.52 2.31 8.05 28.18

a. Pineapple waste has 2.91 g/100 g protein (dry matter).
b. No data were collected at 48 hours of incubation using the submerged liquid fermentation method because the slow growth of the fungi made a determination of protein added by the process impossible.

TABLE 5. Comparison of the data from the different assays in pineapple waste (flesh) fermented by A. niger using the submerged liquid fermentation process for 72 hours

Assay level Waste amount (g) Protein (%) Fibre (%)
Laboratory (Shaker) 30 20.25 --
Semi-pilot (Chemap fermentator) 3,000 9.22 29.77
Pilot (fermentator)a 21,600 8.53 27.13
Pilot (fermentator)b 21,600 5.67 26.84

a. Without salt addition.
b. With incomplete salt addition ino KCl and FeSO4).

The lysine content of the wastes fermented by the A. niger and the M. verrucaria and the lysine needs of poultry were compared. The A. niger meets 80 per cent and 24 per cent of the lysine requirements of laying hens and cockerels, respectively, and the M. verrucaria 48 per cent and 120 per cent (table 3).

The growth of the three fungi varied with the presence of KCl and FeSO4. The A. niger and the M. verrucaria grew better without these salts, demonstrating that some metallic ions can impair the growth of these fungi. The T. viride, however, required the presence of these salts to grow in the substrate used [3].

The values obtained from the enrichment of pineapple waste (flesh plus skin) with these three fungi (table 4) were always lower than those found in pineapple waste (flesh).

Based on the data from preliminary assays, the A. niger was chosen for enrichment of pineapple waste (flesh) using the submerged liquid fermentation method on the laboratory or pilot scale.

A comparison of the results from several assays (table 5) shows that the protein content decreased when the amount of the waste (flesh) increased, depending on the type of equipment used. These data agree well with those of Sakaguchi et al. [2] for 10-litre capacity fermentators in which the fermented material was affected by air flux interruptions. The crude protein content (5.67 per cent) showed, once again, the positive effect of incomplete salts on the A. niger. Since this fermentator does not offer conditions for maintaining the incubation temperature around 30 °C, A. niger's performance was impaired when compared to that of previous assays (table 5).

The highest protein content (5.60 per cent) was detected in the A. niger treated without salts at 48 hours of incubation. These data were similar to those found for the M. verrucaria incubated for 72 hours. According to Sakaguchi et al., higher quality enzyme production can be obtained using the moist-solid-state fermentation method [2]. In our experiments, however, the submerged liquid fermentation method gave the best results.


From these data, it may be concluded that:


This paper was translated into English by L. N. Pedrosa, Department of Nutrition, Health Science Center, Federal University of Pernambuco.


1. C. S. Lacaz, P. S. Minami, and A. Purchio, 0 grande mundo dos fungos (Editora da Universidade de São Paulo, São Paulo,Poligno, 1970).

2. K. Sakaguchi, T. Venara, and S. Hinoshita, Biochemical and Industrial Trial Aspects of Fermentation (Kodansha, Tokyo, 1971).

3. T. J. B. Menezes, L. A. Duchini, and I. B. Figueiredo, "Producao em laboratorio de proteina fungica em bagaco de cane," Revista brasileiro de tecnologia, 7 (4): 439-446 (1976).

4. M. Mohyuddin, T. R. Sharma, A. K. Karel, and E. G. Niemann, "Use of Aspergillus flavus to Evaluate the Relative Nutritive Value in Cultivares of Rye, Wheat and Triticale," Journal of the Science of Food and Agriculturo, 27(10): 943-950 (1976).

5. M. V. Rajagopal, "Microbial Protein from Corn Waste,'' Journalof Food Technology, 12: 663-667 (1977).

6. W. R. Stanton and A. I. Walbridge, "Fermentation of Cassava and Vegetable Substances," Chemical Abstracts, 77: 359 (1972).

7. T. L. M. Stamford, Z. F. Fernandes, M. L. Cavalcante, C. P. Freitas, N. B. Guerra, and R. L. Vieira, "Racão animal a partir de resíduos de frutos fermentados," part 1: "Maracujá," Boletim da Sociedade Brasileira de Ciência e Tecnologio do Alimentos, Campinas, 17 (1): 107-109 (1983).

8. J. C. Senez, M. Raiabovlt, F. Deschamps, "Protein Enrichment of Starchy Substrates for Animal Needs by Solid-state Fermentation," World Annual Review, 35: 36-39 (1980).

9. O. Baily, "Determination of Nitrogen," in Techniques in Protein Chemistry, 2nd ad. (Elsevier, Amsterdam, 1967).

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