24 July 2021
AperTO - Archivio Istituzionale Open Access dell'Università di Torino
Original Citation:
Black soldier fly defatted meal as a dietary protein source for broiler chickens: Effects on carcass traits, breast meat quality and safety
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DOI:10.1017/S1751731119000685 Terms of use:
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Black soldier fly defatted meal as a dietary protein source for broiler chickens:
1
Effects on carcass traits, breast meat quality and safety
2 3
A. Schiavone1,2*, S. Dabbou1, M. Petracci3, M. Zampiga3, F. Sirri3, I. Biasato4, F. Gai2, 4
L. Gasco4 5
6 1
Department of Veterinary Science, University of Turin, Largo Paolo Braccini 2, 7
10095 Grugliasco (TO), Italy 8
2
Institute of Science of Food Production, National Research Council, Largo Paolo 9
Braccini 2, 10095 Grugliasco (TO), Italy 10
3
Department of Agricultural and Food Sciences, Alma Mater Studiorum – University 11
of Bologna, Via del Florio 2, Ozzano dell'Emilia (BO), Italy 12
4
Department of Agricultural, Forest and Food Sciences, University of Turin, Largo 13
Paolo Braccini 2, 10095 Grugliasco (TO), Italy 14
15
* Corresponding author: Achille Schiavone. E-mail: [email protected] 16
17
Running head: Black soldier fly meal in broiler chicken diets 18
19
Abstract
20
Finding insect meals as alternative sources of poultry feedstuffs is a recent research 21
topic, therefore the present study aimed to evaluate the effects of defatted black 22
soldier fly (Hermetia illucens) larvae meal (HI) in broiler chicken diets on the carcass 23
characteristics and meat quality parameters, proximate composition, fatty acid profile 24
and the heavy metal content of the breast meat. Four dietary treatments were 25
designed: a control diet (HI0) and three experimental diets (HI5, HI10 and HI15), 26
corresponding to 50, 100 and 150 g/kg HI inclusion levels, respectively. The inclusion 27
of 50, 100 and 150 g HI meal per kg feed supply 16.56, 33.01 and 49.63 % of 28
required crude protein. The broilers were slaughtered at day 35, the carcasses were 29
weighed, and the breast muscles were excised from 16 birds per each feeding group 30
(2 birds per replicate pens) and used for meat quality evaluation. Linear and 31
quadratic responses were observed, for increasing HI meal levels, in the live and 32
carcass weights (maximum for HI10). As far as the color of the breast meat is 33
concerned, redness (a*) showed a linear response, while yellowness (b*) linearly 34
decreased with increasing HI meal levels (minimum for HI15). As the HI larvae meal 35
increased in the diets, the moisture content linearly decreased and the protein 36
content increased. The total saturated fatty acid and total monounsaturated fatty acid 37
proportions rose, to the detriment of the polyunsaturated fatty acid fraction. The HI 38
larvae meal, used in the current study, represents a valuable protein source for 39
broiler chickens when included by up to 100 g/kg in their diets, as an improved 40
slaughtering performance was observed without any detrimental effects on meat 41
quality parameters or heavy metal residues in the meat. 42
43
Keywords: Hermetia illucens, broiler chickens, fatty acids, insect meal, heavy
44 metals. 45 46 Implication 47
Due to their nutritive value and low environmental impact, there is an increasing 48
interest on black soldier fly larvae meal (Hermetia illucens L.) as potential feed 49
source in poultry diets. This study showed that insect meal from Hermetia illucens 50
larvae can substitute conventional ingredients in the diet for broiler chickens, without 51
any detrimental effects on meat quality parameters or heavy metal residues in the 52 meat. 53 54 Introduction 55
Nowadays, insects are considered a novel and promising alternative dietary protein 56
source for monogastric animals (Makkar et al., 2014). Insects contain high quality 57
and quantity of protein and are characterized by high feed to protein conversion rate 58
(Makkar et al., 2014). Furthermore, they can easily be reared on different secondary 59
raw materials, thus allowing to reduce their disposal costs and promote a reutilization 60
of by-products (Makkar et al., 2014; Boccazzi et al., 2017; Meneguz et al., 2018 ; 61
Ottoboni et al., 2018). Among the different insect species, black soldier fly (Hermetia 62
illucens, HI) larvae meal is already being used in developed countries as a feed for
63
pets and exotic animals, including birds and fish. In relation to the nutritive profile, HI 64
larvae contain large amounts of lipids, which show an extreme quantitative and 65
qualitative variability, depending on the chemical composition of the rearing substrate 66
(Spranghers et al., 2017; Meneguz et al., 2018). In addition, HI contains 58-72% 67
saturated fatty acids (SFA) and 19-40% mono (MUFA) and polyunsaturated fatty 68
acids (PUFA) of the total fat content (Makkar et al., 2014). Moreover, the 69
accumulation of toxic elements (i.e. heavy metals, mycotoxins and pesticides) is also 70
one of the potential hazards associated with insect production (Belluco et al., 2013) 71
but data regarding the chemical safety of reared insects are very scarce. HI has 72
recently been tested as a feed ingredient in conventional poultry diets as a fat source 73
(Schiavone et al., 2017a and 2018) and in laying hens and broiler quails and 74
chickens as protein source (Maurer et al., 2016; Cullere et al., 2016 and 2018), 75
providing satisfactory results in terms of animal performance and gut morphology. In 76
male broiler chickens, Dabbou et al. (2018) demonstrated that increasing levels (50, 77
100 and 150 g /kg) of dietary HI meal inclusion may improve the live weight (LW) and 78
dietary feed intake during the starter period. However, at the highest inclusion level, it 79
may also negatively affect the feed conversion ratio and gut morphology. This 80
suggests that low inclusion levels (50 or 100 g/kg) may be more suitable. However, 81
no significant effects on the haematochemical and histological parameters were 82
observed in relation to HI meal utilization. On the other hand, there is a lack of 83
scientific information about the impact of the use of HI larvae meal on meat safety 84
and quality traits. HI larvae meal utilization as protein source has recently been 85
reported to not significantly affect meat quality parameters in broiler quails, with the 86
only exception of a negative modulation of the fatty acid (FA) profile (Cullere et al., 87
2018). However, no studies are currently available on these aspects in broiler 88
chickens. 89
Based on the above mentioned paper and in order to provide reliable data on the 90
potential use of insect meal in broiler chicken nutrition, the present research aims to 91
evaluate the effects of dietary defatted HI larvae meal inclusion on the carcass traits 92
and meat quality parameters, proximate composition, FA profile and heavy metals 93
residues of broiler chicken breast meat. 94
95
Material and methods
96
Birds and diets
97
A detailed description of the experimental design is reported in Dabbou et al. (2018). 98
Briefly, two hundred and fifty-six one-day-old male broiler chickens were reared from 99
day 1 to day 35 and randomly allotted to four dietary treatments (8 pens/treatment 100
and 8 birds/pen). Four experimental diets were formulated to be isonitrogenous and 101
isoenergetic during the three phase-feedings. The diets were formulated according to 102
Sauvant et al. (2004) for all feedstuffs, except for HI larvae meal for which chemical 103
composition as described by Schiavone et al. (2017b) was used. The diets were 104
prepared including, as a feed basis, increasing levels of HI larvae meal (0, 50, 100 105
and 150 g/kg; HI0, HI5, HI10 and HI15, respectively). The inclusion of 50, 100 and 106
150 g HI meal per kg feed supply 16.56, 33.01 and 49.63 % of required crude 107
protein. A partially defatted HI meal derived from larvae that were fed with vegetable 108
by-products (cereals). The defatting process was performed using high pressure and 109
without solvents. 110
The ingredients and the chemical composition of the experimental diets are reported 111
in Table 1. 112
113
Fatty acid profile of the insect meal and experimental diets
114
The FA composition of the HI larvae meal and experimental diets was assessed 115
using the method described by Schiavone et al. (2007).The fatty acid methyl esters 116
were separated, identified and quantified on the basis of the chromatographic 117
conditions reported by Renna et al. (2014). The results were expressed as g/100gof 118
total fatty acids (TFA) (Table 2). 119
120
Slaughtering procedures
121
At 35 d of age, 16 animals (2 birds per replicate pens) from each feeding group 122
(chosen based on the average final LW in each pen) were individually identified with 123
a shank ring and weighed. The chickens were slaughtered at a commercial abattoir. 124
Plucked and eviscerated carcasses were obtained, and the head, neck, feet and 125
abdominal fat were removed to obtain the chilled carcass. Then, the liver, heart, 126
spleen, bursa of Fabricius, abdominal fat, thigh and breast weights were immediately 127
recorded. The breast and thigh weights were expressed as percentages of the LW. 128
Twenty-four hours after slaughtering, breasts were separated into right and left sides, 129
individually vacuum sealed and refrigerated (4±1°C). Breast meat was divided in two 130
parts and frozen at -20°C for further meat analyses. 131
132
Meat quality parameters
133
The meat quality parameters (pH, color, drip loss, cooking loss and shear force) were 134
assessed on the Pectoralis major muscles of the right breast (from the 16 135
slaughtered animals per each feeding group) following the harmonized 136
methodologies for the assessment of poultry meat quality features detailed described 137
in Supplementary Material S1. 138
139
Chemical composition and fatty acid profile
140
The left breast meat of the 16 slaughtered animals per each feeding group was used 141
to perform chemical analysis. The moisture and ashes were determined according to 142
the Association of Official Analytical Chemists procedure (AOAC, 1990). Proteins 143
were determined using the standard Kjeldahl copper catalyst method (AOAC, 144
1990).Total lipids were measured using a modification of the chloroform: methanol 145
procedure described by Folch et al. (1957). FAs were then determined as reported in 146
Supplementary Material S1. The results were expressed as g/100g of TFA. The 147
health indexes were calculated as described in the Supplementary Material S1. 148
149
Heavy metals and arsenic
The analysis of heavy metals and arsenic were performed on HI larvae meal and 4 151
breast meat pools (one pool per feeding group, each of them composed by 8 152
slaughtered animals [one bird per replicate pens]), as presented in Supplementary 153
Material S1. The results were expressed as mg/kg 12% humidity. 154
155
Statistical analyses
156
The statistical analyses were performed using the IBM SPSS software package (IBM 157
Corp. Released 2012. IBM SPSS Statistics for Windows, Version 21.0. IBM Corp, 158
Armonk, NY). The Shapiro-Wilk test was used to test the normality of the data 159
distribution. Individual bird was used as the experimental unit to analyse the carcass 160
characteristics, meat quality parameters and FA profile (n=16 per treatment; 2 161
birds/pen). The carcass characteristics, meat quality parameters and FA profile data 162
were tested by means of one-way analysis of variance (ANOVA), with experimental 163
diet (HI0, HI5, HI10 and HI15) as fixed effect, and according to the following model: 164
Yij=μi+α+ɛij 165
where Yij = the single observation; μ = overall mean; α = the effect of experimental 166
diet (i= HI0 or HI inclusion level) and ɛ= residual error. 167
The effect of dietary HI meal inclusion was evaluated by means of polynomial 168
contrasts (linear and quadratic responses). The results were expressed as the mean 169
and standard error of the mean (SEM). Significance was considered at P<0.05. 170 171 Results 172 Carcass characteristics 173
The carcass characteristics of the broilers fed the experimental diets are presented in 174
Table 3. Linear and quadratic responses were observed for increasing HI meal levels 175
in the live and carcass weights (P<0.001), with a maximum for the HI10 group. The 176
breast yield increased quadratically for increasing HI meal levels (P<0.05), and the 177
quadratic response increased to a maximum for the inclusion of 100 g/kg of HI meal. 178
The abdominal fat yield showed a linear response to increasing HI meal levels 179
(P<0.05), with a maximum for the inclusion of 150 g/kg of HI meal. On the other 180
hand, no significant effects related to HI meal utilization were observed for the other 181
carcass traits. 182
183
Meat quality parameters of the breast muscle
184
No significant differences were observed for the meat quality parameters, with the 185
exception of the redness (a*) and yellowness (b*) color traits (Table 4). The observed 186
redness showed a linear response to increasing HI meal levels (P<0.05), and the 187
linear response increased to a maximum for the HI15 group. A linear decrease of 188
yellowness was elicited for the HI meal as the level of HI increased with a minimum 189
corresponding to the inclusion of 150 g/kg of HI meal. 190
191
Chemical composition and fatty acid profile of the breast meat
192
As far as the chemical composition is concerned, the moisture content decreased 193
linearly with increasing HI meal levels (P<0.05), and the linear response decreased 194
to a minimum for the inclusion of 150 g/kg of HI meal. A linear response was 195
observed for the protein content, with a maximum for the HI15 group. No significant 196
effect, related to HI meal utilization, was observed for the lipid and ash contents 197
(Table 4). 198
The effects of the experimental diets on the FA profile of breast meat are presented 199
in Table 5. A linear increase was observed for C12, C14, C16, C14:1, C16:1 n7; 200
C20:1 ΣMUFA and atherogenicity index, and the linear response increased to a 201
maximum for the inclusion of 150 g/kg of HI meal. C15:0 and C18:1 c9 showed a 202
linear response to increasing HI meal levels (P<0.05), with a maximum for the 203
inclusion of 100 g/kg of HI meal. A linear decrease was shown for C18:0, C24:0, 204
C18:2 n6, C20:4 n6, ΣPUFA, Σn3, Σn6 and ΣPUFA/SFA. A linear and quadratic 205
decrease was observed for C22:5 n3 and C22:6 n3 with a minimum for the HI10 206
group. 207
208
Heavy metal and arsenic determination
209
The concentrations of heavy metals and arsenic in the HI larvae meal and breast 210
meat samples of the broiler chickens are summarized in Tables 6 and 7, respectively. 211
All the arsenic, cadmium, lead and mercury levels in the HI larvae meal were below 212
the European Union (EU) limits reported for animal feeds (EC, 2002). The heavy 213
metal and arsenic contents in the breast meat sample pools of the four dietary 214
treatments were below the EU limits reported for chicken meat (EC, 2006). 215
216
Discussion
217
HI larvae meal has been recently proposed as emerging and innovative feed 218
ingredient in poultry feeds (Cullere et al., 2016 and 2018; Loponte et al., 2017; 219
Altmann et al., 2018; Pieterse et al., 2019) in order to improve the sustainability of 220
poultry meat production. The defatting process results in insect meals with larger 221
protein values and can reduce the risk of lipid oxidation, allowing for a longer shelf 222
life of the product (Zheng et al., 2013). Our study investigated the effects of defatted 223
HI larvae meal on carcass characteristics and meat quality of broiler chickens as well 224
as the optimal level of inclusion. 225
The results of the relative carcass and organ yields showed a satisfactory effect of HI 226
larvae meal inclusion on the carcass traits, a result that could be positive for 227
commercial purposes. Positive linear and quadratic responses were observed up to 228
100 g/kg level of HI larvae meal for the parameters mentioned above, while HI15 229
group showed unfavorable results. The results observed in HI15 group are in 230
agreement with our previous findings, which revealed a negative modulation of 231
growth performance and gut morphology by insect meal utilization in HI15 birds 232
(Dabbou et al., 2018). In fact, the authors observed the worst gut mucosal 233
development (in terms of short villi, deep crypts and reduced villus height-to-crypt 234
depth ratio) in HI15 group, which may explain the deterioration of the growth 235
performance and consequently the carcass characteristics. 236
On the basis of the results of this research, the supplement of HI larvae meal up to 237
100 g/kg has been suggested to be suitable as a feed ingredient for broiler chicken 238
diets. 239
The results obtained in this study are in agreement with those reported by Loponte et 240
al. (2017). These authors reported greater carcass weights in Barbary partridges
241
(Alectoris barbara) fed with HI and yellow mealworm (Tenebrio molitor, TM) diets 242
than control group as a partial replacement (25 % or 50%) of soybean meal. 243
However, our results are not consistent with the majority of studies about broiler 244
chickens and quails, where all carcass traits were unaffected by dietary house-fly 245
maggots (Musca domestica, MD), TM and HI larvae meal inclusion (Bovera et al., 246
2016; Cullere et al., 2016; Biasato et al., 2017 and 2018; Pieterse et al., 2019). 247
In the current study, the redness (a*) index of breast meat was higher in the HI15 248
group, while the HI5 and HI10 groups showed intermediate values in comparison 249
with HI0 group. These results may be due to a possible accumulation of insect meal 250
pigments in the intramuscular fat. On the other hand, the yellowness value (b*) 251
decreased in a linear fashion (P<0.001), unlike the overall responses of redness to 252
dietary HI larvae meal for an increased HI larvae meal content in the diet. This finding 253
may be attributed to the decreased content of corn gluten meal in HI diets (Table 1). 254
The effects of dietary insect meal on meat color are controversial. Cullere et al. 255
(2016) observed that meat redness (a*) in the breast meat of broiler quails was 256
affected by increasing inclusion levels of HI larvae meal in diets, showing the highest 257
(1.13) and lowest (0.46) values for 100 g/kg and 150 g/kg HI groups, respectively. On 258
the contrary, Leiber et al. (2017), Altmann et al. (2018) and Pieterse et al. (2019) did 259
not find any significant effect of dietary HI meal on broiler meat color. In regard to 260
other insect meals, the use of MD larvae meal in broiler diets has also been 261
associated with a significant decrease in breast muscle lightness (L*) (Pieterse et al., 262
2014).Differently, Bovera et al. (2016)did not find any significant effect on the color 263
of raw and cooked meat, or on the skin of broiler chickens, also showing that the 264
meat from broilers fed with TM meal could be accepted by consumers. However, the 265
meat color differences found in broiler chickens fed with HI larvae meal of the present 266
study appear to be of little practical relevance and potentially incapable of affecting 267
the consumers' willingness to buy meat. 268
Orthogonal polynomial contrast test revealed that increasing the HI larvae meal of 269
diets decreased the moisture content (linear effect, P<0.05), and increased protein, 270
according to a notable linear trend, for increasing HI larvae meal inclusion levels. 271
Even the effects of dietary insect meal on poultry meat proximate composition are 272
conflicting. Cullere et al. (2018), Pieterse et al. (2019) and Schiavone et al. (2017a) 273
reported no significant effects on meat chemical composition of broiler quails or 274
chickens fed diets with HI meal and fat, respectively 275
The FA composition of broiler meat basically depends on the dietary FA profile. 276
Several factors (e.g. genotype, sex, age, slaughter weight) are also able to affect the 277
FA composition of meat, the diet component being considered as a major effect in 278
monogastric animals (Rymer and Givens, 2005; Schiavone et al., 2007 and 2010). 279
Consequently, as expected, the dietary inclusion of HI larvae meal influenced the FA 280
profile of the broiler breast meat. The lauric and myristic acid contents increased with 281
increasing levels of dietary inclusion of HI in the diet, and this increase was also 282
noticed for C16:0. However, the total SFA remained constant, that is, at C18:0 and 283
C24:0 reduced linearly as the inclusion rate increased. These results are in 284
agreement with those reported by Renna et al. (2017) and Cullere et al. (2018) for 285
rainbow trout and broiler quail, respectively, fed defatted HI larvae meal. Avian FAs 286
are typically monounsaturated, due to an active hepatic delta-9 desaturase, and an 287
oleic (C18:1 c9) predominance (Klasing, 2000). An increasing percentage of MUFA, 288
mainly due to the higher content of C18:1 c9, was observed as a result of increasing 289
inclusion levels of HI meal in the diets. On the contrary, PUFA significantly decreased 290
with greater decreases in C18:2 n6. HI larvae meal was found to be relatively scarce 291
in PUFA, which were represented almost entirely by C18:2 n6 and C18:3 n3. Such a 292
composition lowered the contents of the n-6 and the n-3 PUFA fractions as the 293
dietary HI larvae meal inclusion level increased. Despite these remarkable changes 294
in the breast meat FAs, the Σ n-6/ Σ n-3 ratio remained unaffected. Breast meat lipids 295
are mainly composed of triacylglycerol and phospholipids, the latter being rich in very 296
long chain n-3 PUFA (Eicosapentaenoic acid (EPA; C20:5 n3) and Docosahexaenoic 297
acid (DHA; C22:6 n3)), which are well known for their high biological efficiency in the 298
organism and their beneficial effects on human health (Rymer and Givens, 2005). A 299
significant decreased content was observed in HI groups for Docosapentaenoic acid 300
(DPA; C22:5 n3) (on average -34.25%) and DHA (on average -43.42%). These 301
results are in agreement with those reported by Cullere et al. (2018), who found a 302
significant reduction of DHA content in breast meat of Japanese quail fed increasing 303
levels of HI meal. Renna et al. (2017) also observed lower contents of EPA and DHA 304
in the fillet muscles of trout fed 400 g/kg of HI than other groups, while DPA was 305
reduced with the lowest inclusion level of HI meal. The present study indicates that 306
the inclusion of a defatted HI larvae meal in broiler chicken diets leads to significant 307
modifications of the breast meat FA profile, with higher MUFAs and lower PUFAs in 308
HI groups than control group. In order to balance and overcome these potential 309
negative effects related to HI larvae meal utilization, a modulation of the larva rearing 310
substrate should be recommended to obtain an improved insect larvae FA profile and 311
provide healthier meat for the modern consumer. Little information on the influence of 312
insect meals on meat sensory profile and consumer health and acceptance has been 313
provided till now. The dietary inclusion of insect meal may affect the sensory profile 314
of meat as reported by Pieterse et al. (2014), who found that the meat derived from 315
chickens fed with a 10% MD larvae meal had a higher perception of metallic aroma 316
and aftertaste but a higher sustained juiciness and a lower mealiness (dry sensation) 317
in the mouth compared to the control group. Differently, Cullere et al. (2018) reported 318
unaffected meat sensory profile of quails fed HI larvae meal. Further research should 319
be focused on the evaluation of the consumer acceptance and health, and meat 320
processing in order to successfully include HI meal in commercial poultry diets. 321
In the current research, the concentrations of heavy metals and arsenic in both the HI 322
larvae meal and in the breast meat of the birds remained below the EU limits 323
suggested for feed materials (EC, 2002) and foodstuffs (EC, 2006). HI larvae have 324
been reported to accumulate cadmium (Van der Fels-Klerx et al., 2016; Purschke et 325
al., 2017; Biancarosa et al., 2018), lead (Van der Fels-Klerx et al., 2016; Purschke et
326
al., 2017; Biancarosa et al., 2018), mercury (Biancarosa et al., 2018) and arsenic
327
(Van der Fels-Klerx et al., 2016; Biancarosa et al., 2018) when reared on 328
contaminated substrates. The results obtained in the present study suggest that the 329
HI larvae meal was fed with a feeding media that did not contain heavy metals or 330
arsenic levels that exceeded the maximum allowable limits. The cadmium and lead 331
concentrations in the breast meat also remained below the EU limits, thus 332
representing a relevant result in terms of food safety. 333
334
Conclusion
335
Overall, the present study has provided new data and knowledge on the potential use 336
of a new sustainable feedstuff for broiler chickens. The main findings of the current 337
research suggest that defatted HI larvae meal can be used to up to 100 g/kg level of 338
inclusion in broiler chicken diets, without detrimental effects on carcass and meat 339
quality parameters or heavy metals contents. Remarkable differences in the meat 340
nutritional profile were found in relation to the FA composition, with an increase of 341
MUFA content to the detriment of PUFA. Therefore, important efforts should be made 342
to evaluate new substrates capable of improving the FA profile of larvae, thus 343
potentially counteracting the negative effects on the nutritional value and perceived 344
healthiness of the poultry meat. These substrates should also be evaluated from a 345
safety point of view, as new evidence on their safety could be adopted to reduce the 346
potential toxicity of the meal. 347
348
Acknowledgements
Research supported by University of Torino (Italy) founding: SCHA_RILO_14_01 350 (2015-2017) and SCHA_RILO_16_02 (2016-2018). 351 352 Declaration of interest 353
The authors declare that there is no conflict of interest. 354
355
Ethics statement
356
The experimental protocol was designed according to the guidelines of the current 357
European and Italian laws on the care and use of experimental animals (European 358
directive 86 609/EEC, put into law in Italy with D.L.116/92) and approved by the 359
Ethical Committee of the Department of Veterinary Science of the University of Turin 360
(Italy). 361
362
Software and data repository resources
363
The data sets analysed in the current study are available from the corresponding 364
author on reasonable request. 365
366
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Table 1
503
Ingredients, apparent metabolizable energy and chemical composition of the experimental diets. 504
Items Starter period Growing period Finisher period
HI0 HI5 HI10 HI15 HI0 HI5 HI10 HI15 HI0 HI5 HI10 HI15
Ingredients (g/kg as fed)
Maize meal 508.8 526.9 545.5 566.9 536.1 557.1 574.5 606.1 537.2 590.8 620.3 653.2 Soybean meal (48% CP) 345.3 299.0 248.0 193.0 332.0 277.4 230.0 153.0 300.0 253.0 178.0 100.0
HI larvae meal 0.0 50.0 100.0 150.0 0.0 50.0 100.0 150.0 0.0 50.0 100.0 150.0
Corn gluten meal 54.0 32.0 15.0 0.0 35.0 20.0 0.0 0.0 21.0 0.0 0.0 0.0
Soybean oil 46.3 45.1 43.0 40.0 59.3 56.4 54.9 48.0 71.2 70.0 63.2 56.0 Dicalcium phosphate 6.5 8.5 10.5 12.5 4.0 6.0 8.3 10.6 3.5 6.0 8.3 10.5 Calcium carbonate 17.3 16.4 15.6 14.8 16.7 15.8 14.8 14.0 15.1 14.0 13.2 12.4 Sodium chloride 2.3 2.3 2.3 2.3 2.3 2.3 2.3 2.3 2.3 2.3 2.3 2.3 Sodium bicarbonate 1.3 1.3 1.3 1.3 1.3 1.3 1.3 1.3 1.3 1.3 1.3 1.3 DL-methionine 2 2.2 2.3 2.3 1.8 1.9 2 2.0 1.8 1.9 1.9 1.8 L-lysine 5.1 5.2 5.4 5.8 3.8 4.1 4.2 5.0 3.3 3.4 4.1 5.0
Threonine 1.9 1.9 1.9 1.9 1.5 1.5 2.0 2.0 1.1 1.1 1.2 1.3
Trace mineral-vitamin premixa 8.0 8.0 8.0 8.0 5.0 5.0 5.0 5.0 5.0 5.0 5.0 5.0
Choline chloride 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 3-phytase 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 Total 1000 1000 1000 1000 1000 1000 1000 1000 1000 1000 1000 1000 AMEb (MJ/kg DM) 12.56 12.56 12.56 12.56 12.98 12.98 12.98 12.98 13.40 13.40 13.40 13.40 Nutrient compositionc (%) CP 23.01 22.98 23.01 23.01 21.50 21.52 21.52 21.52 19.51 19.47 19.54 19.50 EE 7.30 7.33 7.28 7.14 8.63 8.50 8.50 8.01 9.86 9.89 9.41 8.88 CF 3.25 2.99 2.70 2.41 3.21 2.91 2.64 2.25 3.08 2.82 2.43 2.04 Calcium 0.96 0.96 0.96 0.96 0.87 0.87 0.87 0.87 0.79 0.79 0.79 0.79 Phosphorus 0.48 0.48 0.48 0.48 0.43 0.43 0.43 0.43 0.40 0.40 0.40 0.40 Methionined 0.56 0.56 0.56 0.56 0.51 0.51 0.51 0.51 0.47 0.47 0.47 0.47 Lysined 1.44 1.44 1.44 1.44 1.29 1.29 1.29 1.29 1.16 1.16 1.16 1.16 Threonined 0.97 0.97 0.97 0.97 0.88 0.88 0.88 0.88 0.78 0.78 0.78 0.79
HI=Hermetia illucens; AME = apparent metabolizable energy; DM =dry matter; CP= crude protein; EE=ether extract; CF =crude 505
fiber. 506
a
Mineral-vitamin premix: vitamin A (retinyl acetate), 12500 IU; vitamin D3 (cholecalciferol), 3000 IU; vitamin E (DL-a-tocopheryl 507
acetate), 60 IU; vitamin K (menadione sodium bisulfite), 1.02 mg; riboflavin, 2.0 mg; pantothenate, 8.0 mg; niacin, 6 mg; piridossin, 508
4 mg; folic acid, 0.5 mg; biotin, 0.10 mg; tiamin, 1.0 mg; vitamin B12, 20 mg; Mn, 120 mg; Zn, 80 mg; Fe, 52 mg; Cu, 15 mg; I, 1.5 509
mg; Se, 0.4 mg. 510
b
Calculated according to Schiavone et al. (2017b) and Sauvant et al. (2004). 511
c
Chemical analyses were carried out on three replicates of each feed sample. 512
d
Digestible amino acid estimated according to Schiavone et al. (2017b) and Sauvant et al. (2004) for HI meal and other ingredients, respectively.
Table 2
513
Fatty acid profile of HI larvae meal and experimental diets of broiler chickens (g/100 g of total fatty acids). 514
HI larvae
meal
Starter period Growing period Finisher period
HI0 HI5 HI10 HI15 HI0 HI5 HI10 HI15 HI0 HI5 HI10 HI15
C12:0 54.59 nd 2.73 5.46 8.19 nd 2.73 5.46 8.19 nd 2.74 5.46 8.19 C14:0 10.15 0.10 0.60 1.10 1.60 0.10 0.60 1.10 1.61 0.09 0.60 1.10 1.61 C16:0 12.03 10.37 10.43 10.48 10.55 10.45 10.51 10.58 10.64 10.10 10.55 10.62 10.70 C18:0 1.77 2.50 2.40 2.30 2.17 2.51 2.41 2.29 2.13 2.42 2.39 2.21 2.03 C18:1 c9 7.91 21.51 22.20 22.90 23.65 21.87 22.43 23.17 23.85 22.15 22.73 23.31 23.107 C18:2 n6 5.98 52.59 50.15 47.72 45.30 53.05 50.63 48.20 45.81 51.26 50.83 48.45 46.09 C18:3 n3 0.80 3.26 3.14 2.80 2.41 3.27 3.09 2.77 2.22 3.12 3.03 2.48 2.13 SFA 80.28 15.38 18.67 21.96 25.25 15.24 18.52 21.80 25.05 15.83 18.43 21.70 24.93 MUFA 12.88 24.24 24.82 25.43 26.06 24.36 24.81 25.45 26.03 24.57 25.01 25.48 26.31 PUFA 6.85 58.50 55.82 53.15 50.44 58.68 56.00 53.32 50.52 57.78 56.03 53.23 50.43 Other FA nd 9.60 8.34 7.23 6.12 8.74 7.58 6.43 5.55 10.86 7.13 6.36 6.18
HI= Hermetia illucens;nd= not detected; SFA= saturated fatty acid; MUFA= monounsaturated fatty acid; PUFA=polyunsaturated 515
fatty acid; FA= fatty acids. 516
Table 3
517
Effect of the dietary HI larvae meal inclusion level on the carcass traits of broiler chickens (n=16 animals/group). 518
Dietary treatments SEM P-value
HI0 HI5 HI10 HI15 Linear Quadratic
Live weight (LW) (g) 2260.56 2259.44 2266.87 2070.12 15.69 <0.001 <0.001 Carcass weight (g) 1594.84 1601.01 1607.84 1469.65 11.73 <0.001 <0.001 Carcass weight (% LW) 70.55 70.86 70.92 71.00 0.20 0.453 0.785 Breast (% LW) 14.46 14.67 14.84 13.57 0.16 0.067 0.018 Thigh (% LW) 18.67 18.59 18.79 18.45 0.12 0.676 0.611 Spleen (% LW) 0.11 0.10 0.10 0.11 0.01 0.486 0.262 Liver (% LW) 2.17 2.13 2.08 2.25 0.05 0.707 0.368 Heart (% LW) 0.64 0.60 0.56 0.57 0.01 0.078 0.400 Bursa of Fabricius (% LW) 0.29 0.26 0.26 0.26 0.01 0.532 0.476 Abdominal fat (% LW) 1.21 1.17 1.43 1.44 0.05 0.042 0.813
HI= Hermetia illucens; LW= live weight. 519
Table 4
520
Effect of the dietary HI larvae meal inclusion on the meat quality and chemical composition in breast meat of broiler chickens (n=16 521
animals/group). 522
Dietary treatments SEM P-value
HI0 HI5 HI10 HI15 Linear Quadratic
pHu 6.03 5.99 6.04 5.98 0.05 0.650 0.777 Color Lightness (L*) 55.07 55.46 54.29 53.41 0.38 0.074 0.400 Redness (a*) 2.72 3.18 2.87 3.71 0.14 0.030 0.469 Yellowness (b*) 11.80 9.68 8.19 7.57 0.39 <0.001 0.174 Drip loss (%) 1.24 1.36 1.26 1.39 0.06 0.211 0.928 Cooking loss (%) 20.11 20.49 19.31 19.71 0.36 0.480 0.990
Allo Kramer shear force
(kg/g) 1.86 2.24 2.14 2.06 0.06 0.375 0.074 Proximate composition Moisture (%) 76.14 75.51 75.50 75.24 0.13 0.016 0.460 Protein (%) 22.37 22.28 22.42 23.09 0.12 0.035 0.118 Lipid (%) 1.56 1.76 1.85 1.75 0.05 0.134 0.110 Ash (%) 1.23 1.33 1.24 1.31 0.02 0.427 0.718
HI= Hermetia illucens. 523
Table 5
524
Effect of the dietary HI larvae meal inclusion on the fatty acid profile in breast meat of broiler chickens (g/100g of total fatty acids; 525
n=16 animals/group). 526
Dietary treatment SEM P-value
HI0 HI5 HI10 HI15 Linear Quadratic
ΣSFA 29.13 28.97 29.24 29. 88 0.29 0.352 0.507 C12:0 nd 0.33 0.61 1.03 0.06 <0.001 0.367 C14:0 0.23 0.43 0.54 0.74 0.03 <0.001 0.965 C15:0 nd 0.02 0.05 0.04 0.01 <0.001 0.041 C16:0 17.63 18.24 18.74 19.24 0.20 0.002 0.887 C17:0 0.15 0.12 0.13 0.14 0.01 0.705 0.315 C18:0 9.55 8.48 7.98 7.61 0.21 <0.001 0.352 C20:0 0.19 0.24 0.23 0.23 0.01 0.152 0.245 C24:0 1.38 1.09 0.96 0.85 0.06 0.001 0.416 ΣMUFA 27.61 30.00 31.57 32.17 0.47 <0.001 0.279 C14:1 nd 0.02 0.10 0.14 0.01 <0.001 0.630 C16:1 n7 1.79 2.24 2.62 3.13 0.12 <0.001 0.870 C17:1 0.05 1.11 0.07 0.09 0.01 0.287 0.355 C18:1 c9 25.26 27.08 28.28 28.21 0.36 0.001 0.152 C20:1 0.20 0.26 0.28 0.34 0.01 <0.001 0.330
C24:1 0.31 0.28 0.22 0.26 0.01 0.091 0.182 ΣPUFA 42.36 40.06 37.89 36.65 0.51 <0.001 0.524 C18:2 n6 31.44 30.62 29.61 28.32 0.41 0.004 0.763 C18:3 n6 0.20 0.24 0.22 0.25 0.01 0.355 0.974 C18:3 n3 2.31 2.40 2.47 2.20 0.06 0.641 0.150 C20:2 n6 0.56 0.57 0.57 0.47 0.03 0.319 0.345 C20:4 n6 5.86 4.76 3.90 4.05 0.24 0.002 0.152 C20:5 n3 (EPA) 0.13 0.17 0.17 0.17 0.01 0.207 0.375 C22:5 n3 (DPA) 1.09 0.81 0.62 0.72 0.05 0.002 0.039 C22:6 n3 (DHA) 0.76 0.49 0.34 0.46 0.04 0.001 0.004 Others FA 0.90 0.97 1.29 1.30 0.08 0.032 0.814 ΣPUFA/SFA 1.46 1.40 1.30 1.23 0.03 <0.001 0.901 Σn3 4.30 3.87 3.60 3.56 0.08 <0.001 0.162 Σn6 38.06 36.19 34.29 33.08 0.45 <0.001 0.646 Σn6/n3 8.93 9.39 9.63 9.40 0.13 0.140 0.169 AI 0.26 0.29 0.31 0.34 0.01 <0.001 0.957 TI 0.60 0.61 0.62 0.64 0.01 0.139 0.875
HI= Hermetia illucens; nd= not detected; SFA= saturated fatty acids; MUFA= monounsaturated fatty acids; PUFA= polyunsaturated 527
fatty acids; FA= fatty acids; AI= atherogenicity index; TI= thrombogenicity index. 528
Table 6
529
Heavy metals in HI larvae meal. 530 HI meal MRL (Directive 2002/32/EC) As (mg/Kg 12% h) < 0.05 2 Cd (mg/Kg 12% h) 0.32 2 Pb (mg/Kg12% h) 0.07 10 Hg (mg/Kg 12% h) < 0.02 0.1 Cr (mg/Kg) 0.23 Not legislated Fe (mg/Kg) 189 Not legislated Ni (mg/Kg) 0.18 Not legislated Cu (mg/Kg) 10 Not legislated Zn (mg/Kg) 157 Not legislated Co (mg/Kg) < 0.05 Not legislated Se (mg/Kg) < 0.02 Not legislated
HI= Hermetia illucens; MRL= Maximum Residue Limit; mg/kg 12% h= mg/kg 12% 531
humidity. 532
Table 7
533
Heavy metals in breast meat of broiler chickens (one pool per feeding group, 8 534
birds/pool). 535
Dietary treatments MRL
(Regulation 881/2006/EC)
HI0 HI5 HI10 HI15
As (mg/Kg 12% h) < 0.05 < 0.05 < 0.05 < 0.05 Not legislated Cd (mg/Kg 12% h) < 0.05 < 0.05 < 0.05 < 0.05 0.050 (EU n. 488/14) Pb (mg/Kg 12% h) < 0.05 < 0.05 < 0.05 < 0.05 0.10 (EU n. 1005/15) Hg (mg/Kg 12% h) < 0.02 < 0.02 < 0.02 < 0.02 Not legislated Cr (mg/Kg) < 0.05 < 0.05 < 0.05 < 0.05 Not legislated Fe (mg/Kg) 9.1 6.5 6.5 6.7 Not legislated Ni (mg/Kg) 0.08 < 0.05 < 0.05 < 0.05 Not legislated Cu (mg/Kg) 0.64 0.46 0.41 0.46 Not legislated Zn (mg/Kg) 13 13 13 13 Not legislated Co (mg/Kg) < 0.05 < 0.05 < 0.05 < 0.05 Not legislated Se (mg/Kg) 0.28 < 0.25 < 0.25 < 0.25 Not legislated HI= Hermetia illucens; MRL= Maximum Residue Limit; mg/Kg 12% h= mg/kg 12% 536
humidity. 537
Animal Journal
Black soldier fly defatted meal as a dietary protein source for broiler chickens: Effects on carcass traits, breast meat quality and safety
A. Schiavone, S. Dabbou, M. Petracci, M. Zampiga, F. Sirri, I. Biasato, F. Gai, L. Gasco
Supplementary material S1
Material and methods
Meat quality parameters
pH
The ultimate pH (pHu) measurement, at 48 h post mortem, was established using the dispersed phase method developed by Jeacocke (1977). Approximately 2.5 g of finely minced meat was homogenized for 30 seconds, by means of an Ultraturrax device, in 25 ml of sodium iodoacetate (5 mM) and 18 ml of potassium chloride (150 mM), previously equilibrated at pH 7 with the aid of a potassium hydroxide solution (0.1 N) and of hydrochloric acid (0.1 N). The pH of the homogenate sample, which was previously calibrated using buffer solutions at pH 4 and 7, was determined using a pH meter (Jenwey 3519; Electrode 924001). The values of 2 replicates were considered, and their mean was subsequently utilized.
Color
The meat color was measured at room temperature (20°C) on the inner surface of the
Pectoralis major muscle using a portable colorimeter Chroma Meter CR-400 Konica
Minolta Sensing device (Minolta Sensing Inc., Osaka, Japan). The color measurements were reported in terms of lightness (L*), redness (a*) and yellowness (b*) in the CIELAB color space model (Commission Internationale de l'Éclairage, 1976). The values were recorded for CIE standard illuminant D65 and the CIE 2° standard observer. The color values were obtained considering the average of three readings per sample.
Drip loss
A sample weighing about 80 g (approximately 7 × 4 × 3 cm), obtained from the cranial portion of each fillet, was individually weighed and suspended in a plastic box at 2 to 4°C. After 48 h, the samples were blotted to remove the excess surface fluids, reweighed and the drip loss was determined as the percentage of weight lost by the sample during the refrigerated storage period (Petracci and Baéza, 2011).
Cooking loss
After determination of the drip loss, the meat samples were weighed individually, packaged in a plastic bag under vacuum, and cooked by immersion in a water bath (80°C) until their final internal temperature reached 80°C, according to the recommendations of Petracci and Baéza, (2011). The cooked samples were cooled under running water for 30 min. The samples were then removed from the bags, blotted and weighed. The cooking losses were determined by calculating the difference in weight of the samples before and after cooking and they were expressed as percentages of the initial weight.
Shear values were determined using a TAHDi Heavy Duty texture analyzer (Stable Micro Systems Ltd., Godalming, Surrey, the UK), equipped with an Allo-Kramer shear cell, using the procedure described by Sams et al. (1990). One meat sample (approximately 2 × 4 × 1 cm) from each sample used for the cook-loss determination was cut, parallel to the muscle-fiber direction, weighed, and sheared with the blades at a right angle to the fibers using a 250-kg load cell and a cross head speed of 500 mm/min. The shear values were reported as kilograms of shear per gram of sample.
Fatty acid profile
Fatty acids (FA) were determined as previously reported by Glass and Cristopherson (1969). In short, 250 µg of lipids and 500 µl of the methylating solution (KOH/methanol 2 N) were put into a vial containing 5 ml of hexane and 1 g of anhydrous sodium sulphate. The vial was mixed for 30 s and placed in a water bath at 40 °C for 15 min. The sample was then stirred and cooled on ice. The upper phase, containing fatty acid methyl esters (FAME), was collected and used for the separation of the FA using a Shimadzu GC17A gas chromatograph (Shimadzu Corporation, Tokyo, Japan) with a WP-4 Shimadzu integration system, equipped with a Varian CPSIL88 capillary column (100 m long, 0.25 mm i.d., 0.20 mm thick film) (Varian, Walnut Creek, CA, USA) and a flame ionization detector. The operating conditions of the gas chromatograph were as follows: the oven temperature was kept at 170°C for 15 min, increased to 190°C at a rate of 1 °C/min, then increased to 220°C at a rate of 5°C/min, and kept at this temperature for 17 min. The temperature of the injector and detector were 270°C and 300°C, respectively. Helium was used as the carrier gas at a constant flow rate of 1.7 mL/min. The identification of the individual FAs was carried out using PUFA-2 fatty acid methyl ester standards (Matreya, Pleasant Gap, PA, the USA), and FA quantification was obtained using methyl nonadecanoate 98% (C19:0) (Sigma, Saint Louis, the USA) as the internal standard, which was added prior to lipid extraction. The results were expressed as a percentage of each individual FAME per total FAME detected.
The atherogenicity (AI) and thrombogenicity (TI) indexes were calculated according to Ulbricht and Southgate (1991) as follows:
AI = (C12:0 + 4 × C14:0 + C16:0) / [(Σ MUFA + Σ n-6) + Σ n-3)]
TI= (C14:0 + C16:0 + C18:0) / [(0.5 × Σ MUFA + 0.5 × Σ n-6 + 3 × Σ n-3) + (Σ n-3) / Σ n-6)] where MUFA are monounsaturated fatty acids.
Heavy metals and arsenic
The chromium, iron, cobalt, nickel, copper, zinc, selenium, arsenic, lead, cadmium and mercury concentrations in the HI larva meal and breast meat were determined by means of the accredited test MI 351 Rev. 1/2015 (UNI EN 13804:2002, UNI EN 15763:2010 and UNI EN 13805:2014). Aliquots of the samples (0.5 g) were weighed in allotted digestion vessels and a mixture of deionized water, nitric acid and hydrogen peroxide was added. The vessels were capped and the contents digested under high temperature and pressure using a single reaction chamber microwave digester system. The resulting solutions were transferred to pre-marked, acid-cleaned, plastic test tubes and diluted with deionized water. The metal content was determined by means of the ICP-MS technique (inductively coupled plasma spectrometry). Multi-element measurements were made using an Agilent 7700x ICP-MS (Agilent Technologies) with a collision cell.
CIE (CommissionInternationale de l’Éclairage) 1976. Recommendations on uniform colour spaces-colour difference equations, psychometric colour terms (supplement no. 2 to CIE publication no. 15). Commission Internationale de l’Éclairage, Paris, France.
Glass RL and Christopherson SW 1969. A method for the differential analysis of mixtures of esterified and free fatty acids. Chemistry and Physics of Lipids 3, 405-408.
Jeacocke RE 1977. Continuous measurement of the pH of beef muscle in intact beef carcasses. Journal of Food Technology 12, 375-386.
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Sams AR, Janky DM and Woodward SA 1990. Comparison of two shearing methods for objective tenderness evaluation and two sampling times for physical-characteristic analyses of early harvested broiler breast meat. Poultry Science 69, 348-353.
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