A nitrogen factor for commercial Pangasius (Pangasius hypophthalmus) fillets

Abstract

The Analytical Methods Committee has received and approved the following report from the Nitrogen Factors Sub-Committee.

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Summary

Pangasius (Pangasius hypophthalmus) is a catfish farmed in the brackish waters of the Mekong Delta in Vietnam. It is imported into the UK under the commercial names Pangasius, Basa, Panga or river cobbler. All the Pangasius fish are killed, filleted and packed in Vietnam, and there has been evidence that some of the product exported to the EU has been soaked in water retaining agents (polyphosphates and citrates), but not declared as such. Therefore, an accepted nitrogen factor would help to ensure that consumers are buying correctly labelled or described fish.

The experimental plan involved the preparation of a total of 150 samples in two sessions, one in March and the other in September 2012. Seventy five samples of whole fish were obtained directly from two main aquaculture centres in Vietnam, and 75 samples of commercial Pangasius fillets were obtained from processors in Vietnam known to use good manufacturing practices.

All the samples were analysed for nitrogen content, moisture, ash and fat according to established British Standards Institute methods (or their equivalents) and the results analysed statistically. All three laboratories participating in the study used the rapid Dumas combustion method for the determination of nitrogen content. This method gives slightly higher results (a factor of 1.014) than the Kjeldahl digestion method. The difference between the average nitrogen content for whole Pangasius and the commercial fillet was only 5%, indicating that good manufacturing practice had been used in their preparation, and water uptake had been kept to a minimum. The recommended nitrogen factor for commercial Pangasius fillets is 2.7 (Kjeldahl 2.65). The nitrogen factor determined in this project compares favourably with a German study of organic and conventionally farmed Pangasius fillets. The values also compare favourably with historic data provided by Young's Seafood.

Report

The membership of the Nitrogen Factors Sub-Committee responsible for the preparation of this report was: Dr Mark Woolfe, (Chairman), Dr Roger Wood (Secretary), Mr Andrew Caines, Ms Bhavna Parmar, Mrs Theresa Ekong (until 11 Oct 2013), Mr Charles Boardman, Mr David Keeble, Mr Duncan Campbell, Mr Jeremy Hall, Prof. Michael Thompson, Mr Robbie Beattie, Mrs Selvarani Elahi, Mr Steve Lamming, Mr Steve Moore, Ms Henrietta Tambo, Mr Andrew Furmage, and Ms Liz Moran. Participants in this study were: Dr Mark Woolfe, Mr Steve Lamming, Mr Robert Olivant, Mr Keith Flint, Mr Charles Boardman, Mr Chris Hunt, Dr Joanna Topping, and Prof Michael Thompson, who was responsible for the statistical evaluation.

Introduction

Labelling rules¹ require that for meat products, the species of meat used should be declared. For fish products there is still the option to declare the species of fish or use the generic description “fish”. Where ingredients are highlighted in the name of the food, the amount of that ingredient must be declared as a percentage of the final product (QUID – quantitative ingredient declaration). Although the amount of an ingredient is calculated on a recipe basis, enforcement authorities usually check the declaration by analysis of the finished product. The analysis determines the nitrogen content (mainly on a fat free basis) of a meat or fish ingredient, and converts this to a meat or fish content using a previously determined nitrogen factor. Added water in a fish ingredient can be calculated by difference. Manufacturers of fish products usually check the specification of their raw materials, and nitrogen factors are used to determine if the fish raw material contains excess added water. The use of nitrogen factors to calculate the fish content of coated fish products has also been accepted at an international level as detailed in Codex Alimentarius.² A considerable amount of research has already been undertaken to determine robust nitrogen factors for common meat and poultry species, and some fish species. In this project the nitrogen factor for a new important species of fish for the UK market was determined.

Pangasius (Pangasius hypophthalmus) production

Vietnam is the world's third largest aquaculture producer (2.7 million tonnes) after China (36.7 million tonnes) and India (4.6 million tonnes). It is the fourth largest fish exporter after China, Norway, and Thailand. Around half of Vietnam's aquaculture production is Pangasius, a catfish farmed in ponds in the brackish waters of the Mekong Delta. There has been a dramatic rise in production and export of fillets of this fish over the past 15 years (see Table 1), and the EU takes around 40% of this production. Pangasius hypophthalmus is imported into the UK under the commercial names Pangasius, Basa, Panga or river cobbler.³ Another species, Pangasianodon bocourti (commercial name Royal Basa), is not marketed in the UK. All the Pangasius fish are killed, filleted and packed in Vietnam, and there has been some evidence⁴ that some of the product exported to the EU has been soaked in water retaining agents (polyphosphates and citrates), but not declared as such.

Table 1 Vietnamese Pangasius production 2001–10 in million tonnes of fillets, and value in billion US dollars^a

2001	2002	2003	2004	2005	2006	2007	2008	2009	2010
a FAO Yearbook Fisheries and Aquaculture Statistics 2010 – FAO Rome 2012.
0.114	0.135	0.163	0.255	0.376	0.520	0.850	1.250	1.050	1.140
$0.17	$0.20	$0.24	$0.38	$0.56	$0.78	$1.27	$1.87	$1.57	$1.71

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Experimental

Pangasius samples

(1) Whole Pangasius. Seventy five specimens of gutted whole Pangasius (750–850 g weight) were obtained from two large UK fish processors. Originally the fish were reared in two main aquaculture areas of the Vietnamese Mekong Delta. One group of 40 farmed fish were obtained in October 2011 from Tan Thanh farms located north of Cao Lanh in the Mekong Delta. In July 2012, 35 whole fish that had been farmed either north of Long Xuyen or in Cao Lanh were obtained through Agifish. The two Vietnamese companies use different fish feeds. October to December is in the wet season, and February to May is the dry season in Vietnam. Fish were collected from both seasons in case there were differences in the growth of the fish. The fish were transported frozen and stored at −18 °C at the Young's Seafood site in Grimsby. The whole fish were thawed slowly over a 2 day period, to −2 °C in order to minimise any drip loss, which was in fact unnoticeable. The fish were filleted whilst still stiff, and the fillet trimmed. The two fillets obtained from one fish were homogenised and a 250 g sample transferred to a sample pot.

(2) Commercial Pangasius fillets. Seventy five commercial fillet samples were obtained from three separate processing plants based in Cao Lanh and Long Xuyen in Vietnam. The samples were taken over two periods from the end of September to the end of December 2011, and during March and April 2012. These fillet samples would have come from fish reared in the same two main tributaries of the Mekong as the whole fish samples. The fillets came in 10 kg cartons. The individually quick frozen fillets (IQF) were mostly in the weight range of 110–130 g for each fillet, with some samples in the 90–110 g range. Three fillets were picked at random from the carton, thawed and homogenised to give a 250 g sample. Each carton provided three separate samples. All samples were randomised and sent to three labs for analysis.

(3) Methods of analysis. Samples of whole Pangasius and commercial Pangasius fillets were analysed in duplicate using the following chemical analysis methods:

British Standards Institution, Analytical Methods for Meat and Meat Products

Part 1 1970 (1993) Determination of ash⁵

Part 2 1980 (1993) Determination of nitrogen or equivalent method.⁶ All labs used an equivalent method by determining nitrogen by Dumas combustion using a LECO 2000 CNS.

Part 3 1970 (1997) Determination of moisture⁷

Part 4 1970 (1993) Determination of total fat⁸

The detailed requirements for analysis included:

Two replicates of the four analyses are required per sample, which should add up to 100% ± 2%. The duplicates should be randomised within each batch. Analytical agreement is required between the duplicates – the repeatability limits are 0.5 g per 100 g (moisture and fat) and 0.1 g per 100 g (nitrogen and ash).

The laboratories analysed a standard reference material in duplicate with each batch of fish samples. Laboratories assessed their performance against the above limits, and decided whether repeats were necessary. All three laboratories used the rapid Dumas method for nitrogen determination. This measures the non-protein nitrogen as well as the protein nitrogen, and hence gives higher results than the Kjeldahl method.⁹ The difference is small, and Dumas is higher by a factor of 1.014, which is more important for fish than for meat. Therefore it is usual to quote the nitrogen result for both methods of analysis as Kjeldahl is still widely used in laboratories worldwide.

Results

Whole Pangasius

Table 2 shows the overall chemical results of the 75 samples. The average nitrogen content is 2.84 g per 100 g, and the average fat content is 2.91 g per 100 g. The chemical results of the 3 labs were compared in Table 3, and no significant differences were found for any of the analyses. In terms of the nitrogen and fat content, there was a significant difference between the Pangasius fish supplied from the Tan Thanh farm (north of Cao Lanh) and those supplied by Agifish, from both the An Giang and Dong Thap regions, presumably because of differences in feeding regimes. The results were combined to give an overall mean.

Table 2 Summary of chemical analyses

Analyte	No.	Mean	SD	SEM
Nitrogen %	75	2.84	0.16	0.019
Fat %	75	2.91	1.09	0.13
Moisture %	75	78.7	1.21	0.14
Ash %	75	1.09	0.11	0.013

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Table 3 Two way ANOVA analysis of means from the 3 labs and origin

	N %	Fat %	Moisture %	Ash %
* = Differences significant at 95% confidence.
Lab
A	2.85	2.93	78.3	1.07
B	2.82	2.96	78.8	1.08
C	2.86	2.76	79.0	1.12
Farm	*	*
Tan Thanh	2.76	3.15	78.51	1.065
Others	2.89	2.61	78.90	1.113

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Commercial Pangasius fillets

The chemical analyses of the 75 samples gave an average nitrogen content of 2.69 g per 100 g, and fat content of 1.23 g per 100 g (Table 4). The commercially prepared samples had a significantly lower fat content than the fillets prepared from whole fish by hand because of differences in trimming the fillets.

Table 4 Summary of chemical analyses

Variable	No.	Mean	SD	SEM
Nitrogen %	75	2.69	0.21	0.024
Fat %	75	1.23	0.46	0.052
Moisture %	75	81.4	1.28	0.15
Ash %	75	0.89	0.10	0.012

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In Table 5 a comparison was made between:

• the results of the 3 laboratories,

• fillets from fish harvested in the wet and dry season,

• the fish processed by the Agifish filleting plant, and those from the other two filleting plants (DL 147 and 355).

Table 5 One way ANOVA analysis of results from 3 labs, origin and season

	N %	Fat %	Moisture %	Ash %
* = Difference(s) significant at 95% confidence.
Lab	*
A	2.60	1.28	81.6	0.86
B	2.74	1.14	81.2	0.90
C	2.75	1.28	81.5	0.91
Season	*		*
D	2.65	1.20	81.7	0.89
W	2.76	1.29	80.9	0.88
Fillet plant	*		*
Agifish	2.57	1.15	82.4	0.91
DL147	2.76	1.28	80.8	0.89
DL355	2.70	1.25	81.4	0.84

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All showed significant differences in the nitrogen content. There also were significant differences in moisture content with season and origin (between Agifish and DL147). All the results were combined to give an average figure for each parameter that was measured.

Discussion and conclusions

It was not possible to obtain an equal distribution of samples covering all the variables of season and origin for Pangasius. However, sufficient samples were obtained to look at trends in the overall mean and uncertainty and show that where significant differences were found, these were not of practical significance. The recommended nitrogen factors for whole and commercial Pangasius fillets for both the Dumas and Kjeldahl methods are given in Table 6. These values have been rounded up or down to the nearest 0.05. The values take into account the low fat contents of the fish, with the standard uncertainty (2 × SEM) for means of duplicate analyses. Whole Pangasius is not widely sold on the UK market, therefore, its nitrogen factor is used as a comparison with the processed product, commercial Pangasius fillets. The nitrogen content of the fish ingredient produced by good manufacturing practice for commercial Pangasius fillets is 5% less than for the whole fish. This figure compares favourably with the interim factors in the UK Code of Practice,¹⁰ which indicated a 7% difference in nitrogen factor between whole fish and commercial fillets.

Table 6 Recommended nitrogen factors for Pangasius

Product	Nitrogen factor (Kjeldahl)
Whole Pangasius	2.85 (2.80) ± 0.04
Commercial Pangasius fillets	2.70 (2.65) ± 0.05

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The recommended nitrogen factor for Pangasius commercial fillets of 2.69 (Kjeldahl 2.65) compares favourably to a previous German study,⁴ which found that organically produced Pangasius fillets had a Kjeldahl nitrogen content of 2.72 g per 100 g. Non-organic commercial Pangasius fillets had a Kjeldahl nitrogen content of 2.13–2.52 g per 100 g and were deemed to have been previously soaked in either polyphosphate or citrate before freezing. The nitrogen factor values obtained in this study also compare favourably with historic data from 2007–2013 provided by Young's Seafood¹¹ for 31 samples of commercial fillets with an average nitrogen content of 2.82 g per 100 g (SD 0.208).

Acknowledgements

This project was undertaken as part of the programme of work of the RSC-Analytical Methods Committee – Nitrogen Factors Sub-Committee, and the contribution of members of the Sub-Committee is acknowledged. In particular, Young's Seafood is thanked for its organisation and preparation of the samples, and providing historic nitrogen data. The project (FA0115) was funded by Defra as part of its Food Authenticity Programme.

References

1. The Food Labelling Regulations (SI 1996 No. 1499), as amended, ISBN: 0-11-035941-0, 1996.
2. Codex Standard for Quick Frozen Fish Sticks (Fish Fingers), Fish Portions and Fish Fillets – Breaded or in Batter. CODEX STAN 166 – 1989, Rev 2011.
3. The Fish Labelling (England) Regulations 2010, SI 210 No. 420, HMSO.
4. H. Karl, I. Lehmann, H. Rehbein and R. Schubring, Composition and quality attributes of conventionally and organically farmed Pangasius fillets (Pangasius hypophthalmus) on the German market, Int. J. Food Sci. Technol., 2010, 45, 56–66.
5. BS4401-1: 1998, ISO 936:1998, Methods of Test for Meat and Meat Products. Part 1: Determination of Total Ash.
6. British Standard, BS 4401:1980 ISO 937:1978, “Methods of Test for Meat and meat products – Part 2: Determination of nitrogen content (reference method)”.
7. BS4401-3: 1997, ISO 1442:1997, Methods of Test for Meat and Meat Products. Part 3: Determination of Moisture Content (Reference Method).
8. BS 4401: Part 4: Method A 1970 (Weibull Stoldt, acid hydrolysis) UKAS accredited method based on BS4401-4: 1970, Methods of Test for Meat and Meat Products. Part 4: Determination of Total Fat Content: Method A (Weibull Stoldt).
9. M. Thompson, L. Owen, K. Wilkinson, R. Wood and A. Damant, A Comparison of the Kjeldahl and Dumas Methods for the Determination of Protein in Foods, using Data from a Proficiency Testing Scheme, Analyst, 2002, 127, 1666–1668.
10. Code of Practice on the Declaration of Fish Content of Fish Products. UKAFFP, BFFF, BHA, BRC, Sea Fish Industry Authority, LACOTS, Association of Public Analysts, http://www.seafish.org/media/Publications/Fish_Content_CoP.pdf.
11. Personal communication with Young's Seafood, May 2013.

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Footnote

† Correspondence should be addressed to Dr M. Woolfe, e-mail: mjwoolfe@gmail.com.

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