American Journal of Plant Sciences, 2011, 2, 287-296
doi:10.4236/ajps.2011.23032 Published Online September 2011 (http://www.SciRP.org/journal/ajps)
287
Gross Chemical Profile and Calculation of
Nitrogen-to-Protein Conversion Factors for
Five Tropical Seaweeds
Graciela S. Diniz1,3, Elisabete Barbarino1, João Oiano-Neto2, Sidney Pacheco2, Sergio O. Lourenço1*
1Departamento de Biologia Marinha, Universidade Federal Fluminense, Niterói, Brazil; 2Laboratório de Cromatografia Líquida,
Embrapa Agroindústria de Alimentos, Rio de Janeiro, Brazil; 3Instituto Virtual Internacional de Mudanças Globais, Universidade
Federal do Rio de Janeiro—UFRJ/IVIG, Rio de Janeiro, Brazil.
Received March 6th, 2011; revised April 22nd, 2011; accepted June 28th, 2011.
ABSTRACT
Despite decades of research on marine algae, there are still significant gaps in basic knowledge about chemical com-
position of these organisms, especially in tropical environments. In this study, the amino acid composition and contents
of total nitrogen, phosphorus, lipid, carbohydrate and protein were determined in Asparagopsis taxiformis, Centro-
ceras clavulatum, Chaetomorpha aerea, Sargassum filipendula and Spyridia hypnoides. The seaweeds showed low
lipid contents (lower than 5.5% d.w. in all species) and were rich in carbohydrates (more than 16% d.w. in all sea-
weeds). The percentage of nitrogen, phosphorus and protein varied widely among species, which red algae showed the
highest concentrations. The amino acid composition was similar among the seaweeds, which glutamic acid, aspartic
acid and leucine as the most abundant. All species are poor in histidine. An average of 24.2% of the total nitrogen is
non-proteinaceous. From data of total amino acid and total nitrogen, specific nitrogen-to-protein conversion factors
were calculated for each species. The nitrogen-to-protein conversion factors calculated ranged from 4.51 to 5.21, with
an overall average of 4.86. These findings show that the traditional conversion factor of 6.25 should be avoided for
seaweeds, since it overestimates the actual protein content.
Keywords: Nitrogen-to-Protein Conversion Factors, Protein, Amino Acid, Seaweeds, Element Composition, Tropical
Environment
1. Introduction
Utilization of algae has increased considerably over the
past years as a consequence of growth of research in
various fields [1]. Because seaweed species are rich in
beneficial nutrients, in countries such as China, Japan
and Korea, they have been commonly utilized in human
nutrition for centuries [2]. They have been found to be
good sources of vitamins, carbohydrates, minerals and
proteins [3], but show wide variations among species.
Seaweeds have been increasingly viewed as potential
sources of bioactive compounds with immense pharma-
ceutical, biomedical and nutraceutical importance [4].
Moreover, they have been used in agricultural and indus-
trial research due to their high content of carbohydrates,
proteins, vitamins, and minerals [5], but show wide vari-
ations among species. However, data on the bioavailabil-
ity of these components are limited.
Protein data of marine algae presents many applica-
tions, involving both basic and applied research. How-
ever, comparisons of protein content among species are
difficult because of methodological differences [6,7].
Extraction is one of the main problems in algal protein
analysis, which is performed with variable efficiency by
different methods [8]. Differences in algae cell wall
composition and in procedures used for protein extrac-
tion establish remarkable influence on final results [9].
The most common methods used for protein determi-
nation in algae, Lowry’s method [10] and Bradford
method [11] assays, are subject to interferences from
many factors [12], which are independent of the prob-
lems related to the protein extraction. The interferences
are a consequence of the effects of some substances on
specific amino acids, since that the chemical reactions
which produce the protein quantification depends on the
reactivity of the amino acid side groups [13].

Gross Chemical Profile and Calculation of Nitrogen-to-Protein Conversion Factors for Five Tropical Seaweeds
288
By contrast, total nitrogen analysis is relatively simple
and easy to perform, and nitrogen-to-protein conversion
factors (N-Prot factors) can be used to estimate crude
protein content. The use of N-Prot factors to determine
protein content has some important advantages if com-
pared to other methodologies. Total nitrogen analysis,
carried out by Kjeldhal’s method [14], Hach techniques
[15] or CHN analysis, eliminates the necessity of ex-
tracting the protein content of the sample to be analyzed,
the major problem in protein analysis of algae [8]. Bar-
barino and Lourenço (2005) [7] showed that algal pro-
teins associated with cell membranes are hardly extracted,
confirming the difficulty of reproducing extraction figures
and increasing differences in values found with different
methods. Thus, the use of N-Prot factors also allows bet-
ter comparisons of results among researchers, since pro-
tein is estimated without a tricky previous extraction.
The use of specific N-Prot factors is widely recom-
mended in order to get more accurate estimates of protein
content [16]. The nitrogen:protein ratio does vary ac-
cording to the source considered [17]. The use of N-Prot
factors is particularly wide in food science. Except for a
list of specific N-Prot factors available for certain cereals
(e.g. 5.26 for rise, 5.47 for wheat; [18]), legumes (e.g.
4.75 - 5.87 for cassava root; [19]), mushroom (4.70; [20]),
Cheddar cheese (6.38; [21]) and milk (5.94, [22]) among
other products, the factor 6.25 calculated by reference
[23] is still used for most plant and animal sources. The
use of the traditional factor 6.25 is based on the assump-
tion that samples contain protein with 16% nitrogen and
an insignificant amount of non-protein nitrogen (NPN)
[24]. However, the amino acid composition varies from
one protein source to another, existing different N con-
tent in each amino acid. Moreover, this assumption is
invalid for organisms that contain high concentrations of
other nitrogenous compounds, such as nucleic acids,
amines, urea, inorganic intracellular nitrogen (ammo-
nium, nitrate and nitrite), vitamins and alkaloids [25].
Plant materials, fungi and algae commonly show sig-
nificant amounts of NPN [7,20,26]. In addition, it is
common to find plant materials showing total protein
with less than 16% nitrogen in total amino acid [27]. The
same trends may be applied to the nitrogen distribution in
seaweeds, and the use of the factor 6.25 tends to overes-
timate the protein data [28,29]. Despite this, several au-
thors continue to use the factor 6.25 to estimate seaweed
protein content (e.g. [30-33]).
To compensate the influence of NPN, specific N-Prot
factors must be calculated. Specific N-Prot factors have
already been proposed for 12 marine microalgae [34],
with an overall average N-Prot factor of 4.78. Studies in
this field are needed for seaweeds, since very limited
information is available (e.g. [28,29]).
In a broader sense, data on chemical composition of
seaweeds are predominantly obtained with species from
temperate (e.g. [28,35,36]), warm temperate (e.g. [37,38])
and subtropical coastal environments (e.g. [39-41]). By
comparison, information on chemical composition of algae
from tropical environments is relatively scarce [42-44]
and more data are needed from those regions. Compared
with land plants, the chemical composition of seaweeds
has been poorly investigated and most of the available
information only deals with traditional edible seaweeds
[32,45,46].
The purpose of our study was to determine specific N-
Prot factors for five tropical marine seaweeds, based on
the ratio of amino acid composition to total nitrogen (TN)
content. In addition, we also characterized and compared
the seaweed species regarding hydrosoluble protein, car-
bohydrate, lipid, nitrogen and phosphorus contents.
2. Materials and Methods
2.1. Algae
In this study five macroalgae species were analyzed. The
identification of the species was carried out following the
checklist of reference [47] and with experts’ supervision.
Chlorophyta: Chaetomorpha aerea (Dillwyn) Kützing;
Rhodophyta: Asparagopsis taxiformis (Delile) Trevisan de
Saint-Léon; Spyridia hypnoides (Bory de Saint-Vincent)
Papenfuss; and Centroceras clavulatum (C. Agardh) Mon-
tagne; Ochrophyta: Sargassum filipendula (Agardh).
2.2. Sampling
C. aerea, C. clavulatum and S. filipendula were collected
in June 2007 and S. hypnoides was collected in September
2007 at Arraial do Cabo (22˚57'S 42˚01'W). A. taxiformis
was sampled in June 2007 at Angra dos Reis (23˚00'S
44˚19'W). Both sites are located in Rio de Janeiro State,
southeastern Brazil (Figure 1) and show oligotrophic
characteristics and minor anthropic influence. Whole
thalli of adult plants were collected and washed in the
field with local seawater in order to remove epiphytes,
sediment and organic matter. Plants were packed in plas-
tic bags and kept on ice until returned to the laboratory
(ca. 150 km, Figure 1). In the laboratory samples were
gently brushed under running seawater, rinsed with dis-
tilled water, dried with paper tissue and frozen at –18˚C.
Subsequently, the samples were freeze dried in a Terroni
Fauvel, model LB1500TT device. The dried material was
powdered manually using a mortar and pestle, and it was
kept in desiccators containing silica-gel, under vacuum at
room temperature, until the chemical analyses were car-
ried out.
2.3. Tissue Analysis
The Lowry’s method [10] was used to analyze hydro-

Gross Chemical Profile and Calculation of Nitrogen-to-Protein Conversion Factors for Five Tropical Seaweeds
289
soluble protein in the samples, with bovine serum albu-
min as a protein standard. Spectrophotometric determi-
nations were performed at 750 nm, 35 min after the start
of the chemical reaction.
Total carbohydrate was extracted with 80% H2SO4,
according to reference [48]. The carbohydrate concentra-
tion was determined spectrophotometrically at 485 nm,
30 min after the start of the chemical reaction, by the
phenol-sulfuric acid method [49], using glucose as a
standard.
Total lipid was extracted according to reference [50],
and determined gravimetrically after solvent (chloroform)
evaporation.
Total nitrogen and phosphorus were determined in al-
gal tissue after peroxymonosulphuric acid digestion, us-
ing a Hach digestor (Digesdhal®, Hach Co.) [15]. Samples
were digested with concentrated sulfuric acid at 440˚C
and treated with 30% hydrogen peroxide. Total nitrogen
and phosphorus contents in the samples were determined
spectrophotometrically after specific chemical reactions.
See reference [51] for analytical details.
Total amino acid was determined by high performance
liquid chromatography with pre-column derivatization with
AccQ.Fluor® reagent (6-aminoquinolyl-N-hydroxysuccini-
midyl carbamate), reverse phase column C18 AccQ.Tag®
Nova-Pak (150 × 3.9 mm; 4 μm), ternary mobile phase in
gradient elution composed by sodium acetate 140 mM +
TEA 17 mM pH 5.05 (solvent A), acetonitrile (solvent B)
and water (solvent C), flow 1 ml·min–1 [52]. A Waters,
model Alliance 2695 chromatograph was used, equipped
with a fluorescence detector Waters® 2475 (λex. 250 nm,
λem. 395 nm). Analytical conditions were suitable to deter-
mine all amino acids, except tryptophan, cysteine + cistine
and methionine. The percent of nitrogen in each amino
acid was used to calculate nitrogen recovered from total
amino acid analysis. Aspartic acid, threonine, serine, glu-
tamic acid, proline, glycine, alanine, valine, isoleucine, leu-
cine, tyrosine, phenylalanine, histidine, lysine, and argin-
ine contents were multiplied by 0.106, 0.118, 0.134, 0.096,
0.123, 0.188, 0.158, 0.120, 0.108, 0.108, 0.078, 0.085,
0.271, 0.193, and 0.322, respectively [16].
2.4. Calculation of N-Prot Factors
N-Prot factors were determined for each species by the
ratio of amino acid residues (AA-Res) to total nitrogen
(TN) of the sample: N-Prot factor = AA-Res/TN. Thus,
for a 100 g (dry weight) sample having 16.21 g of amino
acid residues and 3.48 g of TN, an N-Prot factor of 4.66
is calculated.
The amino acid residues of the samples was calculated
by summing up the amino acid masses retrieved after
acid hydrolysis (total amino acids), minus the water mass
(18 H2O/mol of amino acid) incorporated into each
amino acid after the disruption of the peptide bonds [53].
2.5. Statistical Analysis
The results were analyzed by one-way analysis of vari-
ance (ANOVA) with significance level α = 0.05 [54] fol-
lowed, where applicable, with a Tukey’s multiple com-
parison test.
Figure 1. Map showing the sampling sites in Rio de Janeiro State, Brazil: 1. Arraial do Cabo; 2. Angra dos Reis. Location of
the laboratory at Fluminense Federal University is indicated as “3”.

Gross Chemical Profile and Calculation of Nitrogen-to-Protein Conversion Factors for Five Tropical Seaweeds
290
3. Results
The hydrosoluble protein contents ranged from 8.7% (S.
filipendula, brown alga) to 16.1% (C. aerea, green alga)
of the dry weight with intermediate and similar concen-
trations (p > 0.05) in red algae (Table 1). Carbohydrates
were the most abundant substances measured in all spe-
cies, ranged from 16.8% (S. filipendula) to 29.4% (C. aerea)
of the d.w. The values tended to be higher and similar in
green and red algae, except for the rhodophyte A. taxi-
formis, which was significantly lower than the other three
species (p < 0.001). All species studied contained low
concentration of total lipid. The highest value was re-
corded in C. aerea (5.5%, d.w.) and the lowest concen-
tration was found in C. clavulatum (2.8%). The red algae
S. hypnoides and A. taxiformis showed significantly higher
lipid concentration than C. clavulatum (p < 0.001) (Ta-
ble 1).
The percentage of nitrogen showed wide variations
among species, ranging from 1.75% (S. filipendula) to
5.56% (A. taxiformis) of the dry weight. Red algae showed
higher total nitrogen concentrations in the thalli, with
significant differences (p < 0.001) to others groups (Ta-
ble 1). The green alga C. aerea showed an intermediate
concentration of N in comparison to red and brown algae.
The concentrations of phosphorus also varied widely
among species. C. clavulatum showed the highest value
(0.54%, p < 0.001) and S. filipendula and A. taxiformis
showed the lowest concentrations (0.27% and 0.30%,
respectively) (p > 0.05). The green C. aerea and red al-
gae S. hypnoides showed intermediate and similar values
(Table 1). The tissue N:P ratios were low for all species
(N:P < 11:1), except for A. taxiformis, which showed the
highest N:P ratio (18.9:1, p < 0.001), significantly higher
than the other red algae. The lowest N:P ratios were re-
corded in the green and brown algae analyzed (5.83:1
and 6.56:1, respectively).
The amino acid profiles of seaweeds samples are pre-
sented in Table 2. Glutamic acid was the most abundant
amino acid in all species studied. The highest concentra-
tion of glutamic acid (16.3% of total amino acids) was
found in S. filipendula, while A. taxiformis had the low-
est (10.3%) concentrations. Aspartic acid was the second
most abundant amino acid in seaweeds. These values
varied from 9.59% (A. taxiformis) to 12.7% (C. aerea).
The percentage of histidine was the lowest in all species,
and only the brown algae S. filipendula achieved values
close to 2%. The red alga A. taxiformis showed higher
concentrations of valine, phenylalanine and arginine than
the others species studied. The highest concentrations of
tyrosine were observed in the red algae S. hypnoides and
C. clavulatum with values close to 5%. Percentages of
leucine and threonine were similar among all species and
the lowest concentrations of glycine were observed in red
algae S. hypnoides.
The total protein content of the samples is showed in
Table 3 as total amino acid residues. The seaweeds showed
a wide range of total protein concentration, varying from
8.62% (S. filipendula) to 25.1% (A. taxiformis) of the d.w.
The red algae recorded the highest values of total protein.
Nitrogen mass within total amino acid ranged from
1.36% (S. filipendula) to 4.14% (A. taxiformis). The rela-
tive percentage of protein nitrogen was estimated as the
ratio of nitrogen recovered from amino acid to total ni-
trogen (Table 1). Protein nitrogen ranged from 69.5% (C.
clavulatum) to 81% (S. hypnoides) and the red algae
tended to show higher percentages of NPN, except S.
hypnoides.
From the ratio of the mass of amino acid residues to
total nitrogen we calculated specific N-Prot factors for
the seaweeds. The N-Prot factors ranged between 4.51 (A.
taxiformis) to 5.21 (S. hypnoides). The others three spe-
cies recorded intermediate values of N-Prot factors. An
overall average N-Prot factor = 4.86 was calculated from
the data for all species.
Table 1. Gross chemical composition of five species of seaweeds sampled in a tropical site of Brazil. Values are expressed as
percentage of the dry mass and represent the mean of four replicates ± standard deviation (n = 4)#.
Species Hydrosoluble
protein
Total
carbohydrate
Total
lipid
Total
nitrogen
Total
phosphorus
N:P ration
(by atoms)
*** *** *** *** *** ***
Asparagopsis taxiformis 11.7 ± 0.58b 22.9 ± 1.35b 4.80 ± 0.24b 5.56 ± 0.29a 0.30 ± 0.04c 18.9 ± 1.66a
Centroceras clavulatum 11.3 ± 0.64b 27.1 ± 1.73ª 2.78 ± 0.23d 4.63 ± 0.15b 0.54 ± 0.03a 8.55 ± 0.22b
Chaetomorpha aerea 16.1 ± 0.25a 29.4 ± 0.78ª 5.49 ± 0.09ª 2.56 ± 0.13d 0.43 ± 0.04b 5.83 ± 0.16c
Sargassum filipendula 8.72 ± 0.54c 16.8 ± 0.97c 2.92 ± 0.13d 1.75 ± 0.03e 0.27 ± 0.02c 6.56 ± 0.41c
Spyridia hypnoides 10.7 ± 0.92b 27.7 ± 1.47ª 4.20 ± 0.36c 3.98 ± 0.14c 0.39 ± 0.02b 10.3 ± 0.85b
#Mean values significantly different: ***p < 0.001, a > b > c > d > e. Identical superscript letters (a, a; b, b) or absence of letters indicate that mean values are
not significantly different.

Gross Chemical Profile and Calculation of Nitrogen-to-Protein Conversion Factors for Five Tropical Seaweeds291
Table 2. Total amino acid composition of five seaweeds. Results are expressed as grams of amino acid measured in 100 g of
algal protein and represent the actual recovery of amino acids after acid hydrolysis. Values are the mean of three replicates 
SD (n = 3).
Amino acid Asparagopsis
taxiformis
Centroceras
clavulatum
Chaetomorpha
aerea
Sargassum
filipendula
Spyridia
hypnoides
Aspartic acid 9.59 ± 0.82 11.1 ± 0.49 12.7 ± 1.65 11.3 ± 0.26 11.8 ± 0.19
Threonine 5.33 ± 0.24 5.24 ± 0.11 4.89 ± 0.09 4.76 ± 0.08 4.73 ± 0.02
Serine 5.46 ± 0.29 5.15 ± 0.06 4.43 ± 0.05 4.54 ± 0.06 5.57 ± 0.10
Glutamic acid 10.3 ± 0.29 11.8 ± 0.39 12.8 ± 0.45 16.3 ± 1.74 13.4 ± 0.21
Proline 4.17 ± 0.60 4.95 ± 0.18 5.06 ± 0.32 4.38 ± 0.11 4.87 ± 0.13
Glycine 4.77 ± 0.24 5.18 ± 0.09 6.22 ± 1.14 5.48 ± 0.30 3.94 ± 0.12
Alanine 6.95 ± 0.18 6.76 ± 0.14 5.97 ± 0.31 6.16 ± 0.19 6.73 ± 0.09
Valine 7.00 ± 0.46 6.18 ± 0.17 6.31 ± 0.32 5.85 ± 0.15 6.13 ± 0.09
Isoleucine 5.91 ± 0.21 5.46 ± 0.14 4.88 ± 0.12 5.15 ± 0.15 5.34 ± 0.08
Leucine 8.43 ± 0.55 7.39 ± 0.18 8.11 ± 0.49 7.97 ± 0.24 8.16 ± 0.13
Tyrosine 3.58 ± .044 4.96 ± 0.14 3.82 ± 0.26 3.67 ± 0.05 4.95 ± 0.20
Phenylalanine 6.60 ± 0.28 5.02 ± 0.09 5.48 ± 0.05 5.41 ± 0.25 5.74 ± 0.31
Histidine 1.15 ± 0.10 1.84 ± 0.17 1.75 ± 0.19 1.91 ± 0.08 1.00 ± 0.15
Lysine 5.91 ± 0.15 6.80 ± 0.10 7.33 ± 0.21 6.02 ± 0.12 7.26 ± 0.21
Arginine 8.78 ± 0.87 7.31 ± 0.09 6.69 ± 0.50 6.07 ± 0.16 5.65 ± 0.14
Total 93.9 ± 6.60 95.1 ± 2.52 96.4 ± 6.16 95.0 ± 3.93 95.3 ± 2.16
Table 3. Calculation of nitrogen-to-protein conversion factors for five seaweeds based on the amino acid residues to total ni-
trogen ratio. Values are expressed as percentage of the dry matter. Results represent the mean of three replicates  SD (n =
3).
Species Total amino acid Amino acid residues Amino acid-N Protein-N N-Prot factors
Asparagopsis taxiformis 29.3 ± 2.41 25.1 ± 2.07 4.14 ± 0.34 74.5 ± 6.14 4.51 ± 0.37
Centroceras clavulatum 26.8 ± 0.79 23.0 ± 0.68 3.22 ± 0.09 69.5 ± 2.04 4.98 ± 0.15
Chaetomorpha aerea 14.0 ± 0.96 12.0 ± 0.82 1.94 ± 0.13 75.8 ± 5.20 4.69 ± 0.32
Sargassum filipendula 10.1 ± 0.38 8.62 ± 0.33 1.36 ± 0.05 78.0 ± 2.96 4.93 ± 0.19
Spyridia hypnoides 24.1 ± 0.58 20.7 ± 0.50 3.22 ± 0.08 81.0 ± 1.95 5.21 ± 0.13
4. Discussion
Carbohydrates are the most abundant substances in most
seaweeds, since they occur in cell wall (ex.: agar, cellu-
lose) and as storage products (ex.: starch, laminaran).
Brown algae tend to show lower carbohydrates concen-
tration than others groups of seaweeds [33] and the pres-
ence of less reactive carbohydrates may generate under-
estimates of total carbohydrate [55]. This might contrib-
ute to increase differences in comparison to both green
and red algae. Kumari et al. (2010) [4] investigated the
carbohydrate contents in eighteen species of seaweeds
with the content ranged from 15 to 43% and the value
reported for Chaetomorpha spp. (30%) was similar to our
study. Instead, the reference [56] found 33.5% of carbo-
hydrate in Sargassum polycystum, higher than the value
measured by us in S. filipendula.
The metabolism of benthic seaweeds typically in-
volves the production of large amounts of carbohydrates
as storage products [55]. The production of lipids is
greater in planktonic algal species, in which they con-
tribute for floating mechanisms. The total lipid contents
vary with all species and this may reflect the difference
capability of accumulating lipids. The fat content of
seaweeds is generally low and accounts for 1% - 6% d.w.
[57,58]. Altogether, seaweeds species were low in fat and
high in carbohydrate. Both lipid and carbohydrate con-
tents agree with previous studies [45,46]. Studies that
used Folch’s method to measure total lipid content are

Gross Chemical Profile and Calculation of Nitrogen-to-Protein Conversion Factors for Five Tropical Seaweeds
292
especially useful for comparison, such as [33,57,59].
These studies reported crude lipid values in most sea-
weed predominantly lower than 5% of d.w. The same
trend was confirmed in our study, in which all species
showed less than 5.5% of lipids.
The seaweeds show variable N and P tissue concentra-
tions. Differences are related to taxonomic traits and spe-
cies-specific differences of seaweeds in taking up dis-
solved nutrients [60]. Red algae tend to show higher N
tissues concentrations than green and brown algae [29].
Red algae contain phycoerithrin, an N-rich pigment that
increases the nitrogen budget of these species [55]. In
addition, the three red algae tested are fast-growing spe-
cies, which account for a higher N content in comparison
to other species. Conversely, S. filipendula has a complex
thallus and low rate of growth, showing typically low
content of nitrogen [61]. A similar trend dwells a sandy
substrate and it is partially burred.
S. filipendula showed the lowest P concentrations in its
thallus, and this trend can also be interpreted as a conse-
quence of its low growth rates. Chaetomorpha aerea sh-
owed the second higher P concentration, and this may be
related to its contact with sediments. The occurrence of
high tissue concentrations of phosphorus was recorded in
three seaweeds that also occur partially burred in sedi-
ments (Chaetomorpha crassa, Gracilaria cervicornis and
Gracilariopsis tenuifrons) in a seasonal study in Ara-
ruama Lagoon, a hypersaline coastal environment [51].
According to the reference [62] classification of ma-
croalgal nutrient status based on N:P ratio of tissues, a
N:P ratio < 16 indicates N limitation; a N:P ratio 16 - 24
indicates N-sufficiency and P-sufficiency—i.e. no limita-
tion and N:P > 24 indicates P-limitation. According with
this classification, the low N:P ratios found for all spe-
cies (N:P < 11:1), except for A. taxiformis (18.9:1), sug-
gests that these species trend to be N-limited. This is in
accordance to the characteristics of Brazilian coastal wa-
ters, typically oligotrophic [51,63], with low availability
of nitrogen to algal populations. However, this interpre-
tation must be taken with care, since the amount of data
in our study is small, and does not allow for conclusive
remarks on this subject.
Proteins are composed of the one or more chains of
amino acids and the nutritional quality of a protein is
basically determined by the content, proportion and
availability of its amino acids [64]. The main findings of
amino acid composition of algal proteins described here
are in agreement with previous studies [8,9,29,31,33,40].
In general, all species are rich in the acidic amino acids,
glutamic and aspartic acid and poor in histidine. All sea-
weeds samples exhibits similar amino acid patterns, in
which aspartic and glutamic acid constituted a substantial
amount of total amino acids, ranged from 19.9% (A.
taxiformis) to 27.6% (S. filipendula). These two amino
acids contribute to the flavour-related properties charac-
teristic of the marine products and are responsible for the
special taste of the seaweeds. The concentrations of these
two amino acids were higher in brown algae than in red
algae, as previously described by reference [33].The
level of glutamic and aspartic acid together can represent
up 26% and 32% of the total amino acids of the green
species Ulva rigida and Ulva rotundata [8]. The refer-
ence [29] showed that values for aspartic and glutamic
acid together varied from 20.8% to 31.1% in 19 species
of seaweeds. The highest value of lysine was observed in
Chaetomorpha aerea a green alga, in contrast to refer-
ence [31] who found in red algae higher value for lysine.
Protein content of macroalgae from tropical and sub-
tropical coastal environments frequently show low pro-
tein concentrations [40,65]. Our data indicate low protein
concentrations in the algae studied, and this agrees with
the predominantly oligotrophic condition of the Brazilian
coastal waters. According to the literature, in general the
total protein of brown seaweeds is low (3% - 15% of d.w.)
compared to green and red seaweeds (10% - 47% of the
d.w.) [9,46]. Variations in the protein content of sea-
weeds can be due to differences in species composition
and seasonal periods [9]. The level of total and hydroso-
luble proteins recorded in Sargassum filipendula agrees
with the resuts generally found for the protein content of
Sargassum species [29,59]. S. filipendula showed the
lowest concentrations of both hydrosoluble protein and
total protein, what agrees with the low TN found in its
thalli. In addition, results for total and hydrosoluble pro-
teins in S. filipendula were similar, which indicates that
the extraction of protein was efficient with this species.
On the other hand, the red algae showed both total pro-
tein content and TN concentration higher than the other
species tested here. This suggests the presence of both
high concentrations of non-protein nitrogen and variable
degrees of efficiency in the extraction of protein.
In the present study can be observed a remarkable dif-
ference between the protein concentrations obtained with
Lowry’s method and the sum of AA-Res. The protein
concentration estimated with Lowry’s method achieved
only about 50% in red algae in comparison protein con-
centration estimated with total AA-Res. The both method
to protein quantification registered similar values for
green and brown algae. This could suggest a lower effi-
ciency on extraction in red algae, than brown and green
algae. The inefficiency of the protein extraction in sea-
weeds has been discussed by reference [7], especially in
freeze-dried samples. On the other hand, total amino acid
analysis involves an acidic hydrolysis of the samples,
which eliminates problems with protein extraction.
The best estimation of protein content was the sum of

Gross Chemical Profile and Calculation of Nitrogen-to-Protein Conversion Factors for Five Tropical Seaweeds293
AA-Res, which represents the true protein in each sample.
The reference [28] analyzed the protein content from two
species of Porphyra by different methods and found that
the most accurate estimation of protein would be ob-
tained with the knowledge of the molecular weights of
the sequence of the amino acids. These authors affirm
that the sum of amino acids appeared to be the most ac-
curate method of determination of protein. Otherwise, the
protein contents obtained by N-Prot factors were in good
agreement with those of their AA-Res.
A usual way to determine N-Prot conversion factors is
based in the sum of amino acid residues and determina-
tion of the amount of the total protein nitrogen, consid-
ering the individual contribution of each amino acid [16,
22]. Therefore, organisms that have proteins rich in highly
nitrogenous amino acids (e.g. arginine) tend to have
lower N-Prot conversion factors. In contrast, if the total
protein contains large amounts of amino acids with a low
proportion of nitrogen (e.g. tyrosine), the corresponding
factors is likely to be higher. Thus, variation of the total
amino acid concentrations may markedly influence the
calculation of N-Prot factors [16]. The reference [25]
indicated that the N-Prot factors calculated for many
Japanese vegetables by total nitrogen could give a more
accurate protein value than N-Prot value calculated by
total amino acid nitrogen. This trend results from the
presence of significant amounts on NPN in vegetables.
The use of the ratio of amino acids residues to total ni-
trogen to calculate N-Prot factors was described by ref-
erence [29]. The total amino acid content of seaweeds
represents not only amino acids derived from proteins
but also those in the free form. Thus the presence of free
amino acids contributes to an overestimation of the total
protein. However, according to reference [53], the use of
data of total amino acid, without determination of free
amino acids, is a widely accepted procedure to estimate
protein, since in acid hydrolysis some amino acids are
partially or totally destroyed (e.g. tryptophan, cystine,
methionine and serine). The loss during acid hydrolysis
might compensate for the influence of free amino acids
in the quantification of protein by the sum of the total
amino acid residues.
The overall mean N-Prot factors calculated in this re-
port was 4.86. In general a remarkable similarity was
observed with the current overall N-Prot factor proposed
by reference [29]. These authors reported an average
N-Prot factor of 4.92 for 19 seaweeds studied, with av-
erage specific factors for groups: 5.13 for green algae;
5.38 for brown algae and 4.59 for red algae. Reference
[28] proposed mean N-Prot factor of 5.0 obtained for two
species of Porphyra, the seaweed used to make Japanese
sushi. The average N-Prot factors calculated for red algae
in this study was 4.9, with highest value of 5.21, calcu-
lated for S. hypnoides.
Red algae tend to show larger amounts of NPN (30.5%
in C. clavulatum and 25.5% in A. taxiformis), with ex-
ception of S. hypnoides (19%), than brown and green
algae (22% and 24.2%, respectively). As a consequence
of a high NPN in red algae, the reference [29] found that
the N-Prot factors calculated for these algae tended to be
lower than for the other algal groups. This could not be
assessed in the present study because of the small num-
ber of seaweeds studied.
5. Conclusions
The seaweeds assessed here are poor in lipid and rich in
carbohydrate. Results for hydrosoluble protein indicated
that the extraction and reaction of protein occurred in
variable degrees, with lower efficiency with A. taxiformis
and maximum with S. filipendula. The present results
showed that seaweeds have relatively high non-protein
nitrogen concentrations and reinforce that the calculation
of total protein content by the use of the traditional factor
6.25 overestimates the protein data. From the current
data set it is clear that the factor 6.25 is unsuitable for
estimating seaweeds protein contents. The present study
establishes lower N-Prot factor than the traditional factor
for all species. We recommend that the specific N-Prot
factors calculated in this work are used in researches in-
volving the species assessed here. We currently are as-
sessing the effects of temporal variations on N-Prot fac-
tors calculated for seven seaweeds throughout two years
of sampling. These new results will be published soon.
6. Acknowledgements
Authors are indebted to Brazil’s National Council for
Scientific and Technological Development (CNPq) and
Research Support Foundation of Rio de Janeiro State
(FAPERJ) for the financial support of this study. GDS
thanks Coordination of Improvement of Higher Education
Personnel (CAPES) for her scholarship. Authors thank
Dr. Renato Crespo Pereira (UFF) for the use of labora-
tory facilities and to Dr. Joel C. De-Paula (UNIRIO) for
confirming the identification of the seaweeds.
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