Meat Science 193 (2022) 108944

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Meat Science
journal homepage: www.elsevier.com/locate/meatsci

Feeding silkworm (Bombyx mori L.) oil to growing rabbits improves the
fatty acid composition of meat, liver and perirenal fat
A. Dalle Zotte a, *, Y. Singh a, Zs. Gerencsér b, Zs. Matics b, Zs. Szendrő b, S. Cappellozza c,
M. Cullere a
a
Department of Animal Medicine, Production and Health, University of Padova, Agripolis, Viale dell’Università 16, 35020 Legnaro, Padova, Italy
b
Hungarian University of Agriculture and Life Sciences, Kaposvár Campus, H-7400 Kaposvár, Guba Sándor u. 40, Hungary
c
Council for Agricultural Research and Economics, Research Centre for Agriculture and Environment (CREA – AA), Sericulture Laboratory of Padova, Via Eulero 6a,
35143 Padova, Italy

A R T I C L E I N F O A B S T R A C T

Keywords: Silkworm oil was used in a rabbit diet to evaluate its benefit on the fatty acid (FA) profile of different carcass
Insect oil portions. Two experimental diets were prepared: a control diet (commercial diet with 13 g/kg sunflower oil) and
Monogastric feeding a silkworm oil diet (SWO) (commercial diet with 13 g/kg silkworm oil). Rabbits received the experimental diets
Oryctolagus cuniculus
the last three weeks before slaughter, which occurred at 10 weeks of age. At slaughter, hind leg meat, liver, and
Edible portion
Fatty acid profile
perirenal fat were sampled for FA profile analysis. The SWO diet significantly increased the n-3 FA of all three
TBARs sampling sites and halved the n-6/n-3 ratio of hind leg meat and perirenal fat. Furthermore, the liver of SWO
rabbits was also richer in C22:6 n-3 compared to that of control rabbits. In conclusion, this study demonstrated
that the total dietary replacement of sunflower oil with silkworm oil in fattening rabbits positively changed the
FA profile of the considered carcass tissues.

1. Introduction According to the FAO/WHO (2008), there is no rationale for a spe­

One of the main objectives of meat researchers is to improve the
dietetic and health traits of the meat, in particular by reducing the
saturated fatty acid (SFA) fraction in favor of the polyunsaturated fatty
acids (PUFAs) in both meat and fat deposits. The relationship between
the dietary fatty acid (FA) profile and rabbit meat FA profile has been
widely and thoroughly evaluated over the last 40 years (Cavani, Pet­
racci, Trocino, & Xiccato, 2009; Dalle Zotte, 2002; Dalle Zotte, 2014). It
is now known that in monogastric terrestrial livestock, the FA profile of
their meat partly reflects that of their diet, and in this regard, many
scientific articles have dealt with the incorporation of PUFAs of the n-3
series in fattening rabbit diets, mainly aiming to improve the quality
traits of their meat (Cullere & Dalle Zotte, 2018; Dalle Zotte & Szendrő,
2011). Rabbit meat is already a good source of unsaturated fatty acids
(UFA) and linoleic acid (18:2 n-6; LA), which account for 60% and 22%
of total FA, respectively; furthermore, manipulating the rabbit diet is
very effective in increasing the n-3 PUFAs, thus obtaining functional
meat (Dalle Zotte & Szendrő, 2011).
cific recommendation for n-6 to n-3 ratio, or LA to alpha-linolenic acid
(18:3 n-3; ALA) ratio intake in humans, given that intakes of n-6 and n-3
FA lie within the recommendation of the upper value of acceptable
macronutrient distribution range (the upper portion of an intake range
for an energy source associated with reduced risk of chronic disease) for
total n-6 FA (2.5–9% energy) and total n-3 FA (2% energy) (Sioen et al.,
2017).
To this end, oil/fat obtained from some insect species could be an
interesting energy source to replace sunflower oil and soybean oil in
rabbit diets (Dalle Zotte et al., 2018; Gasco et al., 2019; Benzertiha et al.,
2020; Cullere et al., 2022). This is the case for silkworm (Bombyx mori L.)
oil (SWO), which is rich in PUFAs, especially the n-3 series. The silk­
worm is a byproduct of sericulture and is used as food and medicine in
some Asian countries; however, in the countries with the most intense
production, its disposal constitutes a serious environmental problem
(Venkatesh Kumar & Srivastava, 2020). At present, the difficulty in
obtaining silkworm oil determines its high cost, thus precluding its use
in conventional feeds. However, given the recent EU legislation that

* Corresponding author.
E-mail addresses: antonella.dallezotte@unipd.it (A. Dalle Zotte), yazavinder.singh@phd.unipd.it (Y. Singh), gerencser.zsolt@uni-mate.hu (Zs. Gerencsér), matics.
zsolt@uni-mate.hu (Zs. Matics), szendro.zsolt@uni-mate.hu (Zs. Szendrő), silvia.cappellozza@crea.gov.it (S. Cappellozza), marco.cullere@unipd.it (M. Cullere).

https://doi.org/10.1016/j.meatsci.2022.108944
Received 9 May 2022; Received in revised form 5 July 2022; Accepted 4 August 2022
Available online 8 August 2022
0309-1740/© 2022 Elsevier Ltd. All rights reserved.
A. Dalle Zotte et al. Meat Science 193 (2022) 108944

allowed insect PAPs (including silkworm pupa) to be included in fish,
poultry and swine feed (Commission Regulation EU, 2021) together
with the strong input in the search for alternative feedstuffs, it is plau­
sible that considerable quantities of silkworm oil, which has never been
banned by the EU, will become available in the near future at compet­
itive costs. Therefore, SWO defatted meal and oil could represent rich
and sustainable sources of dietary protein and lipids, respectively, for
livestock (Dalle Zotte, 2021).
The objective of the present study was to investigate and discuss the
effect of the dietary replacement of sunflower oil with SWO on the FA
profile and content of rabbit meat, liver and perirenal fat.
2. Materials and methods
2.1. Animals and experimental design
obtained by using the chromatographic peak area according to the in­
ternal standard (C19:0) method and the total lipid content of the sample.
The lipid atherogenic (AI), thrombogenic (TI) peroxidability (PI), and
hypocholesterolemic/hypercholesterolemic (HH) indices of meat sam­
ples were calculated as reported by Dalle Zotte et al. (2018). The FA
profile of the SWO and of the experimental diets (control, SWO) for
growing rabbits is presented in Table 1.
An aliquot of hind leg meat sample was used to determine the extent
of muscle lipid oxidation. To this end, samples were analyzed in dupli­
cate, and the absorbance was read at 532 nm using a spectrophotometer
(Hitachi U-2000; Hitachi, Mannheim, Germany). The thiobarbituric
acid-reactive substance (TBAR) values were calculated from a standard
calibration curve of 1,1,3,3-tetraethoxypropane, and the values were
expressed as mg of malondialdehyde (MDA)/kg meat (Botsoglou et al.,
1994).

The animals were farmed in the experimental rabbitry of the Hun­ 2.3. Statistical analysis
garian University of Agriculture and Life Sciences, Kaposvár Campus
(Hungary), and it was not a permission-needed activity. All rabbits were The fatty acid profiles of meat, liver and perirenal fat samples were
handled according to the principles stated in the European Directive subjected to one-way ANOVA with the experimental diet (control and
2010/63/EU regarding the protection of animals used for experimental SWO) as a fixed effect following the GLM (General Linear Model) pro­
and other scientific purposes (EC, 2010) and according to the Hungarian cedures of SAS 9.1.3 statistical analysis software for Windows (SAS -
legal requirements (32/1999. /III. 31./ and 178/2009. /XII. 29./). Statistical Analysis Software for Windows, 2008). Least square means were
A detailed description of the farming, slaughtering and carcass dis­ obtained, and post hoc pairwise comparison was performed using the
secting procedures is provided in Cullere et al. (2022). Briefly, at Bonferroni correction. The significance was considered at the 95%
weaning (5 weeks of age), n = 64 rabbits (Pannon White breed) were confidence level.
pair-housed in commercial wire-mesh growing cages and fed a com­
mercial diet containing 13 g/kg sunflower oil until 7 weeks of age. From
3. Results and discussion
7 to 10 weeks of age, rabbits were assigned to two experimental dietary
treatments (32 rabbits/treatment): a control diet or a diet containing 13
Bombyx mori chrysalis is very rich in n-3 PUFA constituted exclu­
g/kg silkworm (Bombyx mori L.) oil (SWO) to replace the same amount
sively of alpha-linolenic acid (18:3 n-3; ALA), and its oil presents a
of sunflower oil. The experimental diets were isonitrogenous (control:
favorable n-6/n-3 ratio of 0.17, which is closely related to the chemical
186 g CP/kg; SWO: 190 g CP/kg) and isoenergy (control:17.5 MJ GE/kg;
composition of mulberry leaves (Olteanu et al., 2015), the typical feed
SWO: 17.2 MJ GE/kg), and their nutritional value corresponded to the
for silkworm. Thus, even at a low inclusion level (1.3% in the present
requirements of growing meat-type rabbits (de Blas & Mateos, 2010).
study), it favorably lowered the n-6/n-3 ratio of the enriched diet (12.9
The chemical composition of the experimental diets has been reported in
Cullere et al. (2022).
Table 1
At slaughter (10 weeks of age), one rabbit per cage was randomly
Fatty acid profile (% of total fatty acid methyl esters - FAME) of the silkworm
selected (16 rabbits/treatment), and from each rabbit, the hind legs,
(Bombyx mori L.) oil (SWO), and of the experimental diets for growing rabbits.
perirenal fat, and liver were dissected, individually vacuum-packaged in
polyethylene bags and transported frozen to the Department of Animal SWO Experimental diets

Medicine, Production and Health (MAPS) of the University of Padova, Control SWO
Italy for FA analysis. C6:0 0.00 0.18 0.13
C14:0 0.00 0.13 0.14
2.2. Fatty acid and lipid oxidation analyses C16:0 24.7 12.3 18.1
C17:0 0.00 0.00 0.10
C18.0 4.57 3.27 3.88
Lipid extraction was performed by modified accelerated solvent C20:0 0.00 0.39 0.42
extraction (M-ASE). For experimental diets, extraction was carried out C24:0 0.00 0.00 0.25
using petroleum ether as the solvent, while for silkworm oil, rabbit meat, C16:1 0.00 0.13 0.41
perirenal fat, and liver extraction was performed using a binary solvent C18:1 n-9 31.5 27.8 25.1
C18:1 n-11 0.00 0.10 0.12
mixture of chloroform/methanol at a 1:2 ratio, according to the method C20:1 n-9 0.00 0.23 0.13
of Lee, Trevino, and Chaiyawat (1996). The fat content of the samples C24:1 n-9 0.00 0.00 0.00
was determined gravimetrically after vacuum evaporation under a ni­ C18:2 n-6 5.39 47.3 32.3
trogen stream. Samples were trans-methylated using a methanolic so­ C18:3 n-3 31.7 3.32 11.5
C20:3 n-6 0.00 0.34 0.30
lution of H2SO4 (4%) to determine FA methyl esters (FAME). A biphasic
C20:4 n-6 0.00 0.18 0.15
separation was obtained by adding 0.5 ml of distilled water and 1.5 ml of C20:3 n-3 0.00 0.18 0.16
n-heptane to each sample. FAMEs were quantified by gas chromatog­ C20:5 n-3 0.00 0.20 0.15
raphy (Shimadzu GC17A) equipped with an Omegawax (Sigma–Aldrich C22:6 n-3 0.00 0.22 0.22
Co. LLC., Saint Louis, USA) 250 column (30 m × 0.25 μm × 0.25 μm) and Total SFA 29.2 16.4 23.0
Total MUFA 33.0 28.4 25.7
flame ionization detector. Helium was used as the carrier gas at a con­ Total PUFA 37.1 51.7 44.8
stant flow of 0.8 ml/min. The injector and detector temperatures were n-6 5.39 48.0 32.8
260 ◦ C. Peaks were identified based on commercially available FAME n-3 31.7 3.72 12.0
mixtures (37-Component FAME Mix; Supelco Inc., Bellefonte, PA, USA). n-6/n-3 0.17 12.9 2.70
The results were expressed as % of total detected FA methyl esters SFA: Saturated fatty acids; MUFA: Monounsaturated fatty acids; PUFA: Poly­
(FAME). unsaturated fatty acids;
Furthermore, the quantitative determination (w/w) of FA was also Control: diet contains 1.3% sunflower oil; SWO: diet contains 1.3% silkworm oil.

2
A. Dalle Zotte et al. Meat Science 193 (2022) 108944

vs. 2.7, for control and SWO, respectively) due to the concurrent Table 3
decrease in linoleic acid (LA) and increase in ALA (Table 1). In general, Effect of the dietary inclusion either with 0% (Control) or 1.3% of silkworm
the results of the present research showed that the inclusion of 1.3% (Bombyx mori L.) oil (SWO) on the lipid (g/100 g of meat) and fatty acids content
SWO in the rabbit diet as a complete replacement for sunflower oil (mg/100 g of meat) of rabbit hind leg meat.
affected the FA profile of rabbit meat, liver and perirenal fat, either Experimental Diets RSD1 P-value
expressed as % total FAME or as mg/100 g sample. Authors emphasize Control SWO
that their specific results refer to the fatty acids considered, which, being
N. 16 16
compositional data, do not allow direct comparison with other works. Lipids, g 5.58 5.52 0.92 0.8507
C6:0 6.70 5.70 1.85 0.1369
3.1. Hind leg meat C10:0 8.57 9.17 4.05 0.6745
C12:0 7.39 7.40 3.10 0.9933
C14:0 68.5 62.2 21.0 0.4027
The results in Tables 2 and 3 present the effect of the dietary C15:0 22.9 22.6 3.70 0.8196
replacement of sunflower oil either with 0% (control) or 1.3% SWO on C16:0 937 958 200 0.7642
the FA profile (% total FAME and mg/100 g, respectively) of rabbit hind C17:0 26.6 26.2 4.06 0.7958
leg meat. The oxidative status of the hind leg meat is also presented in C18.0 323 327 54.2 0.8462
C20:0 9.62 10.2 3.31 0.6126
Table 2.
C14:1 1.98 0.67 2.76 0.1914
The total SFA, total monounsaturated FA (MUFA) and total PUFA C15:1 3.65 3.92 1.33 0.5681
were not affected by the dietary treatment (Table 2). Nevertheless, C16:1 63.0 64.8 30.7 0.8653
single PUFAs, such as LA and ALA, were significantly (P < 0.0001) lower C17:1 8.18 6.25 4.86 0.2702
C18:1 n-9 1010 992 198 0.7957
C18:1 n-11 49.7 47.6 8.23 0.4744
Table 2 C20:1 n-9 9.50 10.3 2.34 0.3294
Effect of the dietary inclusion either with 0% (Control) or 1.3% of silkworm C18:2 n-6 1507 1360 224 0.0732
(Bombyx mori L.) oil (SWO) on the fatty acid profile (% of total FAME) and C20:2 n-6 17.4 16.6 4.19 0.5775
oxidative status (TBARs: mg MDA/kg meat) of rabbit hind leg meat. C18:3 n-6 5.95 5.38 1.80 0.3805
C20:3 n-6 12.4 12.7 3.85 0.7968
Experimental Diets RSD1 P-value C20:4 n-6 101 100 13.8 0.8668
Control SWO C18:3 n-3 81.0 190 29.1 <0.0001
C20:5 n-3 6.51 7.27 1.45 0.1482
N. 16 16 Total SFA2 1409 1428 282 0.8446
C6:0 0.15 0.13 0.03 0.0841 Total MUFA3 1146 1125 238 0.8074
C10:0 0.19 0.20 0.07 0.6834 Total PUFA4 1731 1692 265 0.6792
C12:0 0.16 0.16 0.06 0.9700 n-6 1626 1478 236 0.0863
C14:0 1.50 1.37 0.24 0.1324 n-3 87.5 197 30.2 <0.0001
C15:0 0.51 0.51 0.03 0.6723
1
C16:0 20.8 21.4 1.11 0.2010 RSD: residual standard deviation; 2SFA: Saturated fatty acids; 3MUFA: Mono­
C17:0 0.57 0.59 0.10 0.4727 unsaturated fatty acids; 4PUFA: Polyunsaturated fatty acids.
C18.0 7.29 7.33 0.51 0.8398
C20:0 0.22 0.23 0.07 0.4737
C14:1 0.04 0.01 0.05 0.1813 and higher in the SWO group than in the control group, respectively.
C15:1 0.08 0.09 0.03 0.4629 Nutritionally, meat from SWO rabbits was 2.3 times higher in n-3 FA
C16:1 1.34 1.42 0.48 0.6716 (197 vs. 87.5 mg/100 g, for SWO and the control group, respectively; P
C17:1 0.18 0.14 0.10 0.2761 < 0.001, Table 3).
C18:1 n-9 22.5 22.2 0.90 0.3267
Considering the amount of ALA found in rabbit hind leg meat (190
C18:1 n-11 1.12 1.07 0.08 0.0909
C20:1 n-9 0.21 0.23 0.04 0.0977 mg/100 g meat) through a limited dietary inclusion of SWO (13 g/kg)
C18:2 n-6 33.8 30.6 1.67 <0.0001 (Table 3), it can be hypothesized that the dietary claim “source of
C20:2 n-6 0.39 0.38 0.08 0.7524 omega-3 FA”, set by the EC - Commission Regulation EU, 2010 of 300
C18:3 n-6 0.13 0.12 0.04 0.3527
mg ALA/100 g food, could be achieved if the inclusion level of the SWO
C20:3 n-6 0.28 0.29 0.07 0.7568
C20:4 n-6 2.31 2.28 0.33 0.7631
is doubled. No other comparison with the literature is possible, as this
C18:3 n-3 1.80 4.26 0.33 <0.0001 study, to our knowledge, is the first to link dietary SWO inclusion with
C20:5 n-3 0.15 0.16 0.02 0.0709 the FA profile of rabbit meat. In a study, Gugołek, Strychalski, and
Total SFA2 31.4 31.9 1.26 0.3249 Kowalska (2019) fed growing rabbits’ diets supplemented with silk­
Total MUFA3 25.5 25.1 1.30 0.4819
worm pupa meal (containing 26.45% ether extract) at 5% and 10% in­
Total PUFA4 38.9 38.1 2.06 0.2966
n-6 36.9 33.7 1.94 <0.0001 clusion levels, replacing soybean meal. This supplementation almost
n-3 1.95 4.42 0.34 <0.0001 doubled the ALA percentage in the hind leg meat (5.02% total FAME at
n-6/n-3 18.9 7.69 1.04 <0.0001 10% silkworm pupa meal inclusion level, estimating 2.4% of lipids
FA indexes:
coming from silkworm pupa meal), reaching a level similar to the one
AI5 0.42 0.43 0.03 0.5611
TI6 0.80 0.70 0.04 <0.0001
we found with silkworm oil in our study (4.26% total FAME). Recently,
PI7 49.4 51.0 2.99 0.1333 another study tested a lower inclusion level (4%) of silkworm pupa
HH8 2.72 2.62 0.19 0.1780 meal, and the total n-3 FA in the hind leg meat was significantly
TBARs9 0.73 0.73 0.03 0.7853 enhanced (Kowalska, Gugołek, & Strychalski, 2020).
1
RSD: residual standard deviation; 2SFA: Saturated fatty acids; 3MUFA: Mono­ It must be stressed that silkworm chrysalis contains both chitin and a
unsaturated fatty acids; 4PUFA: Polyunsaturated fatty acids; 5AI: Atherogenicity bioactive substance called 1-deoxynojirimycin (1-DNJ), which may limit
index = (C12:0 + 4 x C14:0 + C16:0)/[total MUFA + total (n-6) + total (n-3)]; the digestion of essential nutrients (also of ether extract), as recently
6
TI: Thrombogenicity index = (C14:0 + C16:0 + C18:0)/[(0.5 x total MUFA) + observed in Japanese quails (Dalle Zotte et al., 2021); thus, it is plausible
0.5 x (n-6) + 3 x (n-3/n-6)]; 7PI: Peroxidability index = (% monoenoic x 0.025) that meat enrichment in n-3 FA is much more efficient with the use of
+ (% dienoic x 1) + (% trienoic x 2) + (% tetraenoic x 4) + (% pentaenoic x 6) + pure SWO than with silkworm meal. Due to the dietary inclusion of
(% hexaenoic x 8); 8HH: Hypocholesterolemic / Hypercholesterolemic index =
SWO, the n-6/n-3 ratio halved, thus improving (7.69 vs. 18.9, for SWO
(C18:1 n-9 + C18:2 n-6 + C20:4 n-6 + C18:3 n-3 + C20:5 n-3 + C22:5 n-3 +
and control, respectively; P < 0.0001) and reaching a ratio considered
C22:6 n-3)/(C14:0 + C16:0); 9TBARs: thiobarbituric acid-reactive substances;
MDA: malondialdehyde.
dietetically healthy. The improved healthiness of SWO hind leg meat

3
A. Dalle Zotte et al. Meat Science 193 (2022) 108944

was also highlighted by the result of the TI, the latter being lower and Table 5
thus better in SWO (0.70 vs. 0.80 in SWO and control, respectively; P < Effect of the dietary inclusion either with 0% (Control) or 1.3% of silkworm
0.0001). (Bombyx mori L.) oil (SWO) on the lipid (g/100 g liver) and fatty acids (mg/100 g
The oxidative status of the meat did not change between the two of liver) content of rabbit liver.
groups of rabbits fed the experimental diets, and this was expected, as Experimental diets RSD1 P-value
the meat of the two dietary groups did not differ in terms of the pro­ Control SWO
portion of the FA classes. The observed TBARs value was low (0.73 mg
N. 8 8
MDA/kg meat) and comparable to values found in the literature (Dalle Lipids, g 6.11 6.47 0.63 0.2671
Zotte et al., 2018), partly because rabbit meat contains antioxidants of C14:0 11.8 11.6 3.20 0.8800
alimentary origin, derived by the peculiarity of the feedstuffs composing C15:0 18.6 19.4 2.37 0.5200
the diet (i.e., alfalfa), which is able to control oxidation during short C16:0 782 843 85.3 0.1711
C17:0 56.5 58.2 6.16 0.6001
storage periods.
C18.0 1009 1070 118 0.3138
C16:1 15.9 17.7 3.18 0.2852
C18:1 n-9 534 570 65.6 0.2999
3.2. Liver C18:1 n-11 44.0 46.4 5.51 0.3960
C20:1 n-9 13.8 15.0 4.39 0.5887
C18:2 n-6 1726 1785 167 0.4905
The effect of the dietary replacement of sunflower oil either with 0%
C20:2 n-6 66.2 76.7 15.1 0.1851
(control) or 1.3% SWO on the FA profile of rabbit liver is depicted in C18:3 n-6 32.1 33.0 4.40 0.7203
Table 4 (% total FAME) and in Table 5 (mg/100 g). C20:3 n-6 52.9 52.2 8.51 0.8733
The liver has a very important role in FA biosynthesis, especially in C20:4 n-6 439 431 53.5 0.7841
C18:3 n-3 47.0 87.6 7.88
11-week-old rabbits, where 60% of total lipogenesis has been reported <0.0001
C20:3 n-3 7.24 10.0 2.65 0.0520
to occur, while only 35% was observed in adipose tissue (Benatmane, C20:5 n-3 8.70 8.72 1.95 0.9826
Kouba, Youyou, & Mourot, 2011). Key enzymes for FA biosynthesis are C22:6 n-3 6.96 10.0 2.48 0.0260
Δ9-desaturase for generating MUFA from SFA, elongases 2 and 5 that Total SFA2 1941 2002 286 0.6754
lengthen the carbon chain of essential FA, and Δ5- and Δ6-desaturase Total MUFA3 626 649 87.8 0.0617
Total PUFA4 2467 2495 350 0.8738
n-6 2395 2378 343 0.9263
Table 4 n-3 71.9 116 9.76 <0.0001
Effect of the dietary inclusion either with 0% (Control) or 1.3% of silkworm 1
RSD: residual standard deviation; 2SFA: Saturated fatty acids; 3MUFA: Mono­
(Bombyx mori L.) oil (SWO) on the fatty acid profile (% of total FAME) of rabbit
unsaturated fatty acids; 4PUFA: Polyunsaturated fatty acids.
liver.
Experimental diets RSD1 P-value
that increase double bonds of essential FA (Castellini et al., 2016; Kouba,
Control SWO Benatmane, Blochet, & Mourot, 2008). The FA composition of the diet is
N. 8 8 an external factor that plays a relevant role in modulating liver function:
C14:0 0.24 0.22 0.06 0.5422 dietary PUFAs inhibit hepatic Δ9-desaturase activity, which results in a
C15:0 0.37 0.37 0.03 0.7398
lower content of MUFAs in the liver, fat depots and Longissimus thoracis
C16:0 15.6 15.9 0.50 0.3181
C17:0 1.13 1.10 0.07 0.3568
et lumborum meat of rabbits fed extruded linseed (Benatmane et al.,
C18.0 20.2 20.1 0.93 0.9157 2011; Kouba et al., 2008). The enzyme Δ6-desaturase is required by
C16:1 0.32 0.33 0.05 0.5709 both n-6 (LA) and n-3 (ALA) essential FAs, therefore determining
C18:1 n-9 10.7 10.7 0.75 0.9089 competition for its utilization. In the past, it was thought that Δ6-
C18:1 n-11 0.88 0.88 0.06 0.8904
desaturase had a higher specificity for n-3 FA (Maniongul et al., 1993),
C20:1 n-9 0.28 0.28 0.08 0.8588
C18:2 n-6 34.6 33.6 0.61 0.0081 but later it was demonstrated that this is not always the case: the FA
C20:2 n-6 1.32 1.45 0.26 0.3428 profile of the diet, age of the animal, species and breed are relevant
C18:3 n-6 0.64 0.62 0.06 0.5562 sources of variability in this sense (Suburu et al., 2013).
C20:3 n-6 1.05 0.99 0.10 0.2221
As a consequence, not all animal species present a sufficient synthesis
C20:4 n-6 8.76 8.13 0.39 0.0058
C18:3 n-3 0.94 1.65 0.11 0.0001
of long chain (≥ 20C) PUFA of the n-3 series from the precursor ALA.
C20:3 n-3 0.15 0.19 0.06 0.1826 Among monogastric species, the rabbit showed a good response in this
C20:5 n-3 0.17 0.17 0.04 0.6561 sense (Bernardini, Dal Bosco, & Castellini, 1999), although recent
C22:6 n-3 0.14 0.19 0.04 0.0300 findings showed that genetic selection for growth traits depressed the
Total SFA2 37.6 37.7 0.65 0.6523
production of n-3 long-chain PUFA in favor of n-6 ones, the latter group
Total MUFA3 12.2 12.2 0.86 0.8888
Total PUFA4 47.8 47.0 0.62 0.0297 requiring less metabolic expenditure (Castellini et al., 2016).
n-6 46.4 44.8 0.66 0.0012 Of course, independent of the genetic factor, differences in FA pro­
n-3 1.40 2.19 0.12 <0.0001 portions among the different tissues exist, as highlighted in this study.
n-6/n-3 33.2 20.5 2.42 <0.0001
Compared to rabbit meat, rabbit liver presented higher PUFA contents
FA indexes:
AI5 0.28 0.29 0.01 0.2904
due to the higher content of n-6 FA, to the detriment of MUFA, as in the
TI6 1.23 1.26 0.03 0.0526 latter FA class in which the C18:1 n-9 content almost halved. These
PI7 79.0 77.3 1.71 0.0702 findings were hypothetically attributed to the abovementioned inhibi­
HH8 3.49 3.33 0.12 0.1050 tory effect of dietary PUFAs on Δ9-desaturase enzyme activity, evident
1
RSD: residual standard deviation; 2SFA: Saturated fatty acids; 3MUFA: Mono­ in the liver being the main site for FA biosynthesis.
unsaturated fatty acids; 4PUFA: Polyunsaturated fatty acids; 5AI: Atherogenicity The total n-3 FA content in the two matrices was affected by SWO
index = (C12:0 + 4 x C14:0 + C16:0)/[total MUFA + total (n-6) + total (n-3)]; dietary inclusion, showing a lower content in the liver than in the hind
6
TI: Thrombogenicity index = (C14:0 + C16:0 + C18:0)/[(0.5 x total MUFA) + leg meat (116 mg/100 g vs. 197 mg/100 g, in liver and meat, respec­
0.5 x (n-6) + 3 x (n-3/n-6)]; 7PI: Peroxidability index = (% monoenoic x 0.025) tively; Tables 5 and 3), which could be attributed to the different
+ (% dienoic x 1) + (% trienoic x 2) + (% tetraenoic x 4) + (% pentaenoic x 6) + metabolic functions of the two considered tissues.
(% hexaenoic x 8); 8HH: Hypocholesterolemic / Hypercholesterolemic index =
Considering that the eicosapentaenoic acid (EPA) and docosahexa­
(C18:1 n-9 + C18:2 n-6 + C20:4 n-6 + C18:3 n-3 + C20:5 n-3 + C22:5 n-3 +
enoic acid (DHA) proportions in the control and SWO diets were similar,
C22:6 n-3)/(C14:0 + C16:0).

4
A. Dalle Zotte et al. Meat Science 193 (2022) 108944

the significantly higher content of DHA found in the liver of the SWO Table 6
group compared to the control group (10.0 vs. 6.96 mg/100 g; P < 0.05) Effect of the dietary inclusion either with 0% (Control) or 1.3% of silkworm
indicates effective biosynthesis via ALA, with the latter being prepon­ (Bombyx mori L.) oil (SWO) on the lipid (g/100 g perirenal fat) and fatty acids (g/
derant in the SWO diet compared to the control diet (11.5 vs. 3.32% total 100 g of perirenal fat) content of rabbit perirenal fat.
FAME). Thus, an increase in the precursor in the diet resulted in an in­ Experimental Diets RSD1 P-value
crease in long-chain PUFAs of the n-3 series in the liver. Specifically, Control SWO
ALA increased by 1.86-fold (P < 0.0001) and DHA by 1.44-fold (P <
N. 16 16
0.05) in the SWO group compared to the control group, thanks to the Lipids 90.6 90.6 3.08 0.9958
highest ALA proportion in the SWO diet. As a result, FA indices C6:0 0.19 0.18 0.04 0.1967
improved, and the n-6/n-3 ratio significantly decreased in the SWO C10:0 0.34 0.34 0.12 0.9625
group (33.2 vs. 20.5, for control and SWO, respectively; P < 0.0001). C12:0 0.32 0.29 0.11 0.4459
C14:0 1.83 1.72 0.21 0.1695
However, the ratio remained higher with respect to the value recom­
C15:0 0.56 0.57 0.03 0.8382
mended by nutritionists. C16:0 21.3 22.6 1.23 0.0067
Although no other research has tested the effect of dietary SWO on C17:0 0.62 0.65 0.10 0.4873
the rabbit liver FA profile, a few studies have focused on the effect of C18.0 5.40 5.76 0.40 0.0179
different dietary incorporations of n-3 FA-rich sources on the rabbit liver C20:0 0.15 0.14 0.10 0.8134
C24:0 0.19 0.13 0.06 0.0111
FA profile. A study testing different dietary lipid sources (sunflower, C14:1 0.04 0.02 0.05 0.4862
flaxseed, fish oil) in rabbit diets observed a linear n-3 PUFA increase in C15:1 0.12 0.14 0.01 0.0062
the liver as a function of the n-3 PUFA dietary content (Bernardini et al., C16:1 1.25 1.33 0.43 0.6010
1999), which is similar to what was found in our study. When rabbit C17:1 0.33 0.17 0.05 0.8688
C18:1 n-9 25.7 25.8 0.74 0.6740
diets were formulated including 10% ground flaxseed (dietary ALA
C18:1 n-11 0.87 0.84 0.08 0.2175
content: 30% total FA), this determined an increase in the liver content C20:1 n-9 0.23 0.23 0.09 0.7799
of ALA but not of other longer-chain PUFAs (EPA and DHA). In parallel, C18:2 n-6 36.6 32.3 2.17 <0.0001
a reduction in n-6 PUFAs was also observed (Ander et al., 2010). A more C20:2 n-6 0.25 0.22 0.07 0.3717
recent study showed that rabbits fed either 10% extruded flaxseed or 3% C20:4 n-6 0.15 0.18 0.06 0.2723
C18:3 n-3 2.16 5.01 0.40
fish oil had a liver richer in n-3 PUFA and poorer in n-6 PUFA (Mattioli
<0.0001
C20:5 n-3 0.16 0.13 0.04 0.0735
et al., 2019), which was in line with the findings of this study as well as C22:6 n-3 0.05 0.08 0.07 0.2079
with the abovementioned literature. The liver is part, although small, of Total SFA2 30.9 32.3 1.45 0.0091
the edible portion of the rabbit carcass, and it may contribute to the Total MUFA3 28.4 28.5 1.09 0.6682
Total PUFA4 39.3 37.8 2.23 0.0721
daily n-3 FA intake, especially if rabbits are fed n-3 rich diets.
n-6 37.0 32.6 2.16 <0.0001
n-3 2.37 5.22 0.38 <0.0001
n-6/n-3 15.7 6.32 0.89 <0.0001
3.3. Perirenal fat FA indexes:
AI5 0.43 0.45 0.04 0.1532
TI6 0.79 0.81 0.05 0.1389
Perirenal fat, being a storage tissue, strictly reflects the dietary FA PI7 43.7 45.3 2.29 0.0661
composition, as already observed for rabbits long ago (Bernardini et al., HH8 2.82 2.62 0.24 0.0259
1999). In fact, among the three carcass tissues considered in this study, 1
RSD: residual standard deviation; 2SFA: Saturated fatty acids; 3MUFA: Mono­
the dietary treatment had the greatest quantitative effect on the peri­
unsaturated fatty acids; 4PUFA: Polyunsaturated fatty acids; 5AI: Atherogenicity
renal fat (Table 6). The inclusion of 13 g SWO/kg diet significantly index = (C12:0 + 4 x C14:0 + C16:0)/[total MUFA + total (n-6) + total (n-3)];
increased C16:0 (P < 0.001) and C18:0 (P < 0.05), reduced C24:0 (P < 6
TI: Thrombogenicity index = (C14:0 + C16:0 + C18:0)/[(0.5 x total MUFA) +
0.05) among SFA, and increased the MUFA C15:1 (P < 0.01). However, 0.5 x (n-6) + 3 x (n-3/n-6)]; 7PI: Peroxidability index = (% monoenoic x 0.025)
the most relevant changes concerned linoleic acid (C18:2 n-6), which + (% dienoic x 1) + (% trienoic x 2) + (% tetraenoic x 4) + (% pentaenoic x 6) +
decreased from 36.6 to 32.3 g/100 g fat (for control and SWO, respec­ (% hexaenoic x 8); 8HH: Hypocholesterolemic / Hypercholesterolemic index =
tively; P < 0.0001), and ALA (C18:3 n-3), which increased by 2.32 times (C18:1 n-9 + C18:2 n-6 + C20:4 n-6 + C18:3 n-3 + C20:5 n-3 + C22:5 n-3 +
compared to the control group (2.16 vs. 5.01 g/100 g, for control and C22:6 n-3)/(C14:0 + C16:0).
SWO, respectively; P < 0.0001). SWO perirenal fat was poorer in SFA (P
< 0.01) and total n-6 FA (P < 0.0001) and richer in total n-3 (5.22 vs. 4. Conclusions
2.37 g/100 g, for control and SWO, respectively; P < 0.0001) compared
to the control group. Therefore, the n-6/n-3 ratio was significantly Silkworm oil proved to be an excellent source of energy and linolenic
lowered and thus improved in the SWO group (6.32 vs. 15.7 for SWO and acid for growing rabbits and was able to further enhance the nutritional
control, respectively; P < 0.0001). The ratio n-6/n-3 was even lower quality of different rabbit tissues. The advantage of using the SWO is to
than that obtained for the hind leg meat of the SWO-fed group. The deliver healthy fatty acids avoiding the possible negative side effects
hypocholesterolemic/hypercholesterolemic (HH) index was also signif­ caused by the presence of 1-DNJ in silkworm meal. The latter reduces
icantly improved by the SWO diet (P = 0.0259). the absorption of sugars, therefore potentially reducing the nutritional
Perirenal fat represents a dissectible adipose tissue and rarely be­ value of the diet with possible negative effects on production
comes part of the edible portion of rabbit carcass. However, the results performance.
of this study suggest that it could be an additional source of healthy
omega-3 FAs, especially if its FA profile and content are enhanced by CRediT authorship contribution statement
dietary means. Considering that the dietary claim “high in omega-3 FA”
is set by the EC - Commission Regulation EU, 2010 for 600 mg ALA/100 A. Dalle Zotte: Conceptualization, Methodology, Resources, Vali­
g and per 100 kcal food, our results indicate that the amount of ALA in dation, Supervision, Project administration, Funding acquisition,
the rabbit perirenal fat largely surpasses the claim for both the control Writing – review & editing. Y. Singh: Formal analysis, Data curation,
(2160 mg/100 g fat) and the SWO (5010 mg/100 g fat) groups. Visualization. Zs. Gerencsér: Resources, Writing – review & editing. Zs.
Reasonably, consumers could benefit from the claim (600 mg ALA/day) Matics: Formal analysis, Writing – review & editing. Zs. Szendrő: Re­
of a perirenal fat daily intake of 27.8 g from control rabbits or of 12 g sources, Writing – review & editing. S. Cappellozza: Resources, Writing
from SWO rabbits only. – review & editing. M. Cullere: Formal analysis, Investigation, Data

5
A. Dalle Zotte et al. Meat Science 193 (2022) 108944

curation, Writing – original draft, Writing – review & editing, Dalle Zotte, A. (2021). Do insects as feed ingredient affect meat quality? Theory and
Practice of Meat Processing, 6(3), 200–209. https://doi.org/10.21323/2414-438X-
Visualization.
2021-6-3-200-209
Dalle Zotte, A., Cullere, M., Martins, C., Alves, S. P., Freire, J. P. B., Falcao-e-Cunha, L., &
Bessa, R. J. B. (2018). Incorporation of black soldier fly (Hermetia illucens L.) larvae
fat or extruded linseed in diets of growing rabbits and their effects on meat quality
Declaration of Competing Interest
traits including detailed fatty acid composition. Meat Science, 146, 50–58. https://
doi.org/10.1016/j.meatsci.2018.08.002
The authors declare no conflicts of interest. Dalle Zotte, A., Singh, Y., Squartini, A., Stevanato, P. G., Cappellozza, S., Kovitvadhi, A.,
Subaneg, S., Bertelli, D., & Cullere, M. (2021). Effect of a dietary inclusion of full-fat
or defatted silkworm (Bombyx mori L.) pupa meal on the nutrient digestibility and
Acknowledgments faecal microbiome of fattening quails. Animal, 15, Article 100112. https://doi.org/
10.1016/j.animal.2020.100112
EC - Commission Regulation EU. (2010). Commission Regulation (EU) No 116/2010 of 9
This research was supported by the University of Padova (Italy) February 2010 amending Regulation (EC) No 1924/2006 of the European
funds (DOR2082232/20). The authors are grateful to Sandro Tenti for Parliament and of the Council with regard to the list of nutrition claims. Official
technical support. Journal of the European Union, L37/16-18. if 10.2.2010 https://eur-lex.europa.eu/le
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