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The Open Agriculture Journal, 2014, 8, 48-51

Open Access

Potential Use of Proximate and Fatty Acid Composition to Distinguish Between Cultured and Wild Largemouth Bass (Micropterus salmoides)
Bobban Subhadra1, Rebecca Lochmann2,* and Andrew Goodwin3
1

Quorum Innovations, Sarasota, Florida, USA

Department of Aquaculture and Fisheries, University of Arkansas at Pine Bluff, USA

Fish Health Program, U.S. Fish and Wildlife Service, Pacific Region, Portland, Oregon, USA
Abstract: Cultured largemouth bass (LMB) cannot be sold as food in some lucrative markets due to regulatory restrictions that protect wild LMB. Distinguishing between cultured and wild fish could open food markets for cultured fish.
Wild LMB eat freshwater fish and cultured LMB eat diets high in marine fish meal and oil, which should produce differences in flesh composition. We analyzed the proximate and fatty acid composition of wild and cultured LMB muscle to
determine the potential for distinguishing fish origin analytically. Protein and moisture were higher in wild fish, while
lipid was higher in cultured fish. The n-3 and n-6 fatty acids, long-chain polyunsaturated fatty acids (LC-PUFA), and the
ratio of n-3 to n-6 fatty acids all differed between cultured and wild fish. The n-3 to n-6 ratio and n-3 LC-PUFA were
higher in cultured fish, while elevated arachidonic acid (20:4n-6) in wild fish was a key distinguishing feature.

Keywords: Arachidonic acid, Fatty acids, Largemouth bass, Lipid, Proximate, Wild and cultured.
INTRODUCTION
The largemouth bass (LMB)(Micropterus salmoides) is
native to the midwestern and southeastern United States and
northeastern Mexico, and has been introduced throughout the
USA and other countries. This species is economically important as one of the most popular sportfishes in the USA. It
is also valuable as a foodfish sold live in fish markets patronized by Asian customers in major cities in the US and Canada. With the dual importance of maintaining wild populations of LMB and satisfying foodfish markets, state wildlife
managers are concerned that wild LMB will be caught and
sold as cultured foodfish. This would deplete wild stocks, so
many states prohibit the sale of all LMB to protect the sportfishing industry [1]. The development of a test that would
distinguish cultured from wild LMB unambiguously could
provide the basis for regulations that would allow the sale of
cultured LMB while protecting wild populations.
Diets of wild and cultured LMB are distinctive, which
might produce differences in body composition. Fatty acid
composition has been used successfully to distinguish cultured and wild members of other fish species [2-4]. The fatty
acid composition of fish tissues correlates directly with dietary fatty acids [5, 6]. Currently, culture of LMB depends
mostly on salmonid diets that contain high levels of marine
fish meal and oil [7]. These diets contain high concentrations

*Address correspondence to this author at the Department of Aquaculture

and Fisheries, University of Arkansas at Pine Bluff, USA;
Tel: 1-870-575-8124; Fax: 1-870-575-4639;
E-mail: [email protected]

1874-3315/14

of n-3 long-chain polyunsaturated fatty acids (n-3 LCPUFA), including eicosapentaenoic acid (EPA; 20:5n-3), and
docosahexaenoic acid (DHA; 22:6n-3). Previous growth
trials with LMB fed a commercial trout diet showed that
LMB retain these LC-PUFA preferentially in the tissues [8].
Freshwater wild fish generally contain higher levels of C-18
fatty acids (such as linolenic acid, 18:3n-3), but also substantial concentrations of EPA and DHA [9-12]. In addition,
freshwater fish are characterized by high proportions of n-6
fatty acids such as linoleic acid (18:2n-6) and arachidonic
acid (20:4n-6) [13]. Freshwater algae, crustaceans, and
aquatic fish larvae are rich in 18:2n-6, 18:3n-3, and 20:5n-3,
although there are considerable differences among species,
seasons, and geographical locations [14-16]. Largemouth
bass have a limited ability to modify dietary fatty acids, and
the fatty acids in the diet are invariably reflected in the fatty
acid profile of the tissues. Therefore, we analyzed the
proximate and fatty acid composition of the muscle of cultured and wild LMB to determine the feasibility of this approach.
MATERIALS AND METHODOLOGY
Seven cultured LMB were obtained live from a producer
who raises fish in outdoor ponds with a commercial diet
similar to those used for trout. The labeled composition of
the diet (Skretting USA, formerly Silvercup, Murray, Utah,
USA) was 45% protein and 20% lipid. Most of the protein
and lipid was from marine fish meal and oil, respectively,
and the majority of largemouth bass producers in the region
(midwestern and southeastern United States) used this diet.

2014 Bentham Open

Potential Use of Proximate and Fatty Acid Composition

The Open Agriculture Journal, 2014, Volume 8

49

Table 1. Hepatosomatic Index (HSI) and Proximate Composition (g/100g) of the Muscle of Cultured and Wild LMBa.
Analyzed composition

Cultured

Wild

HSIb,c

0.78 0.06

0.81 0.07

76.7 0.5**

79.6 0.1*

18.1 0.1**

18.9 0.1*

3.1 0.2*

0.9 0.0**

1.1 0.02

1.1 0.03

Moisture
Protein

Lipidc
Ash

The values are means SEM of 6-7 fish per treatment. A students t-test was performed to compare the difference in the means. Means within rows with different numbers of asterisks (*) are significantly different (P<0.05).
b
Hepatosomatic index (HSI) = Liver weight (g)/ body weight (g) x 100.
c
Wet weight basis.

The diet contained key LC-PUFA including 20:4n-6 (2.0%),

20:5n-3 (13.8%), and 22:6n-3 (5.3%). Seven wild LMB were
caught by hook and line in the Arkansas River near Pine
Bluff, Arkansas. Both wild and cultured fish were collected
during June and July, 2005. The cultured fish were in their
second growing season (about 18 months old). All wild and
cultured fish were marketable size (393-793 g). Mean (SEM)
weights of whole fish were 502.6 (93.4) and 564.4 (122.3) g
for farmed and wild fish, respectively, and were not statistically different (P>0.05).
Fish were handled according to the Guidelines for the
Use of Fishes in Research [17]. Fish were euthanized with an
overdose of MS-222 and kept completely covered with ice
until dissection (up to 2 hours). Individual weights of whole
fish were determined and the fillets were removed and stored
frozen until proximate and fatty acid analysis. The livers
were removed and weighed, and the hepatosomatic index
(HSI) was calculated as liver weight (g) / body weight (g) x
100. The muscle from each fish was homogenized separately
prior to analysis. Proximate analysis of muscle was conducted on at least six fish per treatment (cultured or wild),
and fatty acid analysis was conducted on muscle from four
fish per treatment. Muscle samples were analyzed for nitrogen to determine protein content using the Kjeldahl method,
and dry matter and ash were analyzed using standard methods [18]. Total lipids from muscle were extracted and quantified with chloroform and methanol [19], then saponified and
methylated using potassium hydroxide and boron trifluoride
prior to fatty acid analysis [20, 21]. Fatty acid methyl esters
were quantified using a flame ionization gas chromatograph
(Varian, Model CP3800, Walnut Creek, California, USA)
equipped with a capillary column (50 m x 0.25 mm) (Varian
CP Select for Fame #CP7420) as described previously [20,
21]. Concentrations of the fatty acids were expressed as
grams/ 100 g to perform the calculations. Data were subjected to an unpaired students t-test to determine differences
(P<0.05) between treatment means.
RESULTS AND DISCUSSION
The percentage of total lipid in muscle of cultured LMB
(3.1%) was significantly higher than that of wild LMB
(0.9%) (Table 1). Cultured fish usually receive a commercial
diet on a regular basis, but wild fish forage in areas where

prey availability varies. In general, wild fish are more susceptible to periods of inadequate energy intake. Therefore,
cultured fish frequently contain a higher concentration of
lipid than wild fish [22-24]. Muscle lipid in wild and cultured red porgy (Pagrus pagrus) was 0.65% and 3.03%, respectively [25], which is comparable to our results. Similarly, muscle lipid in wild and cultured sharpsnout sea bream
(Diplodus puntazzo) was 1.73% and 3.87%, respectively [2].
Fillets of cultured salmon (Oncorhynchus kisutch), rainbow
trout (Oncorhynchus mykiss), and channel catfish (Ictalurus
punctatus) are all higher in lipid than their wild counterparts
[26]. Nevertheless, based on our data and the classification
cited by Ackman [27] both cultured and wild LMB can be
considered lean. Moisture and protein in muscle of wild
LMB were higher than those of cultured LMB, but muscle
ash and hepatosomatic index did not differ between treatments (Table 1). The sizes of cultured and wild LMB in this
study were similar, so differences in proximate composition
were most likely due to diet and possibly other environmental factors, but not fish size [28, 29].
Cultured and wild LMB could be distinguished on the
basis of muscle saturates, monounsaturates, total n-3 and n-6
fatty acids, total n-3 LC-PUFA, and the ratio of n-3 to n-6
fatty acids (Table 2). Of the individual fatty acids, arachidonic acid (20:4n-6) was exceptionally high in the wild fish
(12.6 g/ 100g) compared to the cultured fish (2.9 g/ 100g).
Similar results were obtained in red porgy [25] and gilthead
sea bream (Sparus aurata) [3]. Concentrations of 20:4n-6 are
low in cultured fish because practical dietary lipid sources
such as marine fish oils contain small amounts of this fatty
acid [5]. In contrast, the 20:5n-3 was lower in wild bass (1.8
g/ 100g) than in the cultured ones (7.1 g/ 100g). This difference is also attributable to the use of marine fish oils in practical diets, which are rich in 20:5n-3. The natural diets of
wild freshwater fish such as largemouth bass contain more
22:6n-3 than 20:5n-3, since 22:6n-3 is the predominant n-3
LC-PUFA in insects, other invertebrates, and the fish that
consume them [28, 30]. The concentration of DHA did not
differ between wild and cultured fish, which is consistent
with the maintenance of DHA at a physiologically optimal
level regardless of dietary concentrations [31, 32]. Total n-3
fatty acids and n-3 LC-PUFA were higher in cultured than in
wild LMB, which reflects the marine fish meal and oil content of commercial diets typically fed to cultured LMB.

50 The Open Agriculture Journal, 2014, Volume 8

Subhadra et al.

Table 2. Fatty Acid Composition of Muscle (g/100g of total fatty acids by weight) of Cultured and Wild Largemouth Bassa,b.
Fatty Acids

Cultured

Wild

16:0

14.4 (0.2)

15.6 (0.8)

4.0 0.3**

7.8 0.2*

27.3 0.5**

29.7 0.8*

6.8 0.4*

3.9 0.7**

26.9 0.9*

20.1 2.9**

MUFA e

36.7 1.6*

27.5 3.4**

18:2n-6

8.3 0.1*

5.3 0.3**

18:3n-3

1.1 0.1**

3.1 0.3*

20:4n-6

2.9 0.3**

12.7 1.3*

20:5n-3

7.1 0.3*

1.8 0.2**

22:6n-3

15.9 0.6

16.5 1.5

n-6

12.0 0.4**

21.6 2.6*

n-3g

29.1 0.7*

25.1 2.1**

n-3 LC-PUFAh

27.9 0.7*

22.0 2.1**

n-3/n-6 ratio

2.4 0.1*

1.2 0.1**

18:0
Saturates

16:1
18:1

Values are means SEM of 4 fish per treatment. A students t-test was used to compare treatment means. Means within rows followed by different numbers of asterisks (*) were
significantly different (P<0.05).
Fatty acids detected at
0.1 g/ 100g of total fatty acids by weight are not included.
c
Saturates included 14:0, 16:0, 18:0 and 20:0.
d
Total n-9 and n-7 isomers.
e
Monounsaturates included 14:1, 16:1, 18:1 and 20:1.
f
Total n- 6 fatty acids included 18:2n-6, 20:3n-6 and 20:4n-6.
g
Total n-3 fatty acids included 18:3n-3, 20:5n-3 and 22:6n-3.
h
Total n-3 LC-PUFA included 20:5n-3 and 22:6n-3. The only n-6 LC-PUFA detected was 20:4n-6.
b

CONCLUSION

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There were numerous differences in both proximate and

fatty acid composition of cultured and wild LMB. Differences in muscle lipid composition and fatty acid composition
(especially arachidonic acid) could be used as key determinants of fish origin. Application of this assay to test fish
samples from markets would ensure that wild bass populations are protected. These results are based on a small sample
of LMB consisting of cultured fish from a single farm and
wild fish from a known location in the same region. Additional studies with larger numbers of fish are needed to determine how consistent and robust these differences are with
respect to geographical location, season, and diet.

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CONFLICT OF INTEREST
The authors confirm that this article content has no conflict of interest.

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ACKNOWLEDGEMENTS
We thank Aaron Watson and Ben Batten for obtaining
the wild fish, and Nick Kinsey for processing samples and
assisting with laboratory analysis.

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Revised: November 22, 2014

Accepted: November 24, 2014

Subhadra et al.; Licensee Bentham Open.

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