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Construction of a free-form amino acid database for vegetables and mushrooms

Hinako Ito

Department of Food and Nutritional Sciences, Graduate School of Humanities and Sciences, Nara Women's University, Nara 630-8506, Japan

E-mail : bhuvaneswari.bibleraaj@uhsm.nhs.uk

Hiroshi Ueno

Laboratory of Biochemistry and Microbiology, School of Agriculture, Ryukoku University, Seta, Otsu 520-2194, Japan

Hiroe Kikuzaki

Department of Food and Nutritional Sciences, Graduate School of Humanities and Sciences, Nara Women's University, Nara 630-8506, Japan

DOI: 10.15761/IFNM.1000186

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Abstract

Free-form amino acids were quantitatively measured in over 30 species of vegetables and mushrooms. Significant amounts of free-form amino acids were found in each species analyzed. Additionally, there appear to be no rules or patterns of free-form amino acid distribution among species. Highly functional free-form amino acids, like γ-aminobutyric acid, are found in significant levels and suggest that amino acid analysis could play an informative role in nutrition. Our analysis should provide valuable data for the establishment of nutritional databases that include free-form amino acids and more accurately represent the entire amino acid profile of foods.

Key words

free-form amino acid, -aminobutyric acid, mushroom, supplements, vegetable

Introduction

Amino acids play significant biological roles in human. The amino acids required for human metabolism arise from the digestion and utilization of amino acids from food, and from free-form amino acids within food components (For a list of amino acid abbreviations used in this study: Table 1). Free-form amino acids may be utilized as nutrients and/or act as messengers that target specific receptors.

Table 1. List of amino acids and their abbreviations

Amino acid

3 Letter abbreviation

Alanine

Ala

β-Alanine

β-Ala

Arginine

Arg

Asparagine

Asn

Aspartic acid

Asp

Citrulline

Cit

Cysteine

Cys

Cystine

Cys-Cys

γ-Amino butyric acid

GABA

Glutamic acid

Glu

Glutamine

Gln

Glycine

Gly

Histidine

His

Hydroxyproline

HYP

Isoleucine

Ile

Leucine

Leu

Lysine

Lys

Methionine

Met

Ornithine

Orn

Phenyl alanine

Phe

Proline

Pro

Serine

Ser

Threonine

Thr

Tryptophan

Trp

Tyrosine

Tyr

Valine

Val

Free-form amino acids exist in foods derived from plants and animals. β-alanine (β-Ala), citrulline (Cit), g-aminobutyric acid (GABA), and ornithine (Orn) usually exist as free-form amino acids and perform various biological functions including coenzyme components, control of osmoregulation, ammonia detoxification, and as neurotransmitters [1,2]. In human, tryptophan (Trp), a precursor of serotonin and melatonin, acts to regulate circadian rhythm [3]. Some free-form amino acids act as neurotransmitters: in the central nervous system GABA and glycine (Gly) function as inhibitory signals while glutamic acid (Glu) is an excitatory signal [4-8] and may have similar functions in non-neuronal cells [5]. The roles of free-form amino acids in dietary supplements have been extensively explored and dietary branched chain amino acids (BCAA; including valine (Val), leucine (Leu), and isoleucine (Ile)), improve exercise performance, endurance performance, and upper body power in human [9]. Moreover, GABA can reduce high blood pressure [10], and in Japan is authorized as “Tokuho”, which is a “food for specified health uses”, and other health foods containing GABA are widely available.

Glu receptors have been identified on taste buds and the gastrointestinal tract, including stomach cells [11], which suggests that free-form Glu may be involved in enhancing digestion, nutrient absorption, and/or utilization of nutrients [12]. The study was extended to examine the use of daily dietary Glu supplements in the meals of elderly people in hospital [12]. The results of this analysis showed that Glu supplementation improved the nutritional status of elderly and nutritionally deficient patients through improved food intake [12-14]. Therefore, free-form Glu is a highly effective supplement with the potential to improve quality of life (QOL) [12,15,16]. It is likely that free-form amino acids exert useful effects on physiologic functions, particularly during food digestion and nutrient absorption.

Some free-form amino acids act as palatable compounds and are key determinants in the taste of food. Free-form Glu, also known as monosodium glutamate or MSG, is an umami compound isolated from kelp, and is also rich in tomatoes and cheese [16]. Other free-form amino acids that confer taste include Gly, alanine (Ala), and proline (Pro) which provide sweetness, and Leu, Ile, and Val which provide a bitter taste [17,18]. L-serine (Ser) gives mainly sweetness and minor umami taste, and D-Ser, enantiomer of L-Ser, has sweetness. As for taste of D-amino acids, it is known that D-Ala and D-Leu are sweeter than their L-enantiomers [19]. Amino acids elicit not only single basic tastes but also complex taste when they are combined.

Information about the "amino acid" content of various agricultural products is available in databases, typically called "Standard Tables of Food Composition". However, amino acid databases usually list amino acid composition measured in acid hydrolyzed samples, so most of these databases do not accurately reflect free-form amino acid contents [20]. The Stein-Moore laboratory, at Rockefeller University, has shown that acid hydrolysis destroys glutamine (Gln), asparagine (Asn), Trp, and some other amino acids such as GABA, Orn, Cit [21,22]. Thus, little is known about the kinds and amounts of free-form amino acids present in foods.

Studies of nutrition, particularly concerning the quality of proteins, have progressed extensively since information about the amino acid composition of foods has become readily available. However, information about the amino acid composition of both hydrolyzed samples and free-form amino acids in the original food samples are necessary for analyses of palatability and the beneficial functions of amino acids. The Japan Society of Nutrition and Food Science website lists some information about free-form amino acids in food, but the data presented was collated from literature published about 30 years ago [23-26]. Therefore, the amino acid contents listed in some of these databases are likely to be misleading or incomplete.

Amino acid analysis has advanced over the last few decades, and the time has come to renew these data using modern techniques that provide increased sensitivity and reliability. Currently, only a few databases specializing in free-form amino acids are available. To understand the amount of free-form amino acids in our daily foods, free-form amino acids must be "extracted" and then measured. Previously, we reported initial attempts to establish a free-form amino acid database for approximately 140 species of fruits and vegetables [27]. We showed that distribution of free-form amino acids is characteristic for each sample and significantly different from the amino acid distribution recorded in acid hydrolyzed databases. Here, we have extended our analysis of free-form amino acid contents to include additional vegetables and some mushroom species.

Materials and methods

Extraction of free amino acids from foods

We examined 38 species of vegetables and mushrooms (Table 2). Some samples were purchased from local supermarkets in Shiga Prefecture and others were supplied by local farmers. The food samples came from various geographic regions. All food samples were washed in water and the edible and inedible parts, including peels, seeds, and skins, were separated. Samples were stored at -25°C until use.

Table 2. List of foods analyzed for free-form amino acids

 

Scientific Name

Food

Vegetables

Abelmoschus esculentus

Okra “Yashiro-shio okura”

Allium cepa

Onion

Allium cepa

Red onion

Capsicum annuum

Paprika

Chrysanthemum morifolium

Edible chrysanthemum

Chrysanthemum morifolium

Edible purple chrysanthemum “Enmeiraku”

Cucurbita maxima

Winter squash

Cucurbita moschata

Japanese squash

Daucus carota

young carrot (Leaf)

Daucus carota

Moso bamboo

Phyllostachys heterocycla

Pea

Pisum sativum

Peas (Tutankhamen)

Pisum sativum

Podded pea

Raphanus sativus

Radish sprout

Rheum rhabarbarum

Garden rhubarb

Solanum mlycopersicum

Cherry tomato

Solanum melongena

Egg plant “Beinasu”

Solanum melongena

Egg plant “Kamonasu”

Solanum melongena

Egg plant “Kyokonasu”

Solanum melongena

Egg plant “Marunasu”

Solanum melongena

Egg plant “Zebranasu”

Spinacia olerace

Spinach “Okame”

Vicia faba

Broad bean

Vigna unguiculata

Cowpea

Vigna unguiculata

 

Cowpea beans

Pumpkin “Korinnki”

 Mushrooms

Agaricus bisporus

Button mushroom (brown)

Agaricus bisporus

Button mushroom (white)

Auricularia auricula-judae

Jew’s ear

Boletus edulis

Porcini mushrooms (Dried)

Flammulina velutipes

Winter mushroom

Grifola frondosa

Mushroom “Maitake”

Hypsizygus marmoreus

 Beech mushroom

Hypsizygus marmoreus

 Beech mushroom(Tanba)

Letinula edodes

 Mushroom “Shiitake”

Lyophyllum shimeji

 Mushroom “Honshimeji”

Pleurotus eryngii

King oyster mushroom

Sample preparation

Samples (5-10 g) were ground to a fine powder with a mortar and pestle. The powdered samples were added to 100 mM HEPES-Na buffer, pH 7.0, and homogenized on ice with a Tissue-Tearor (Biospec). The homogenate was centrifuged at 10,000×g for 10 min and the supernatant was collected. Suitable supernatant aliquots were treated with 60% perchloric acid to remove proteins. The protein content of samples was estimated by Bradford protein assays (BioRad). Ultrafiltration was performed using CENTRICON®10 (Millipore) when further removal of protein contamination was necessary. After ultrafiltration, the protein content was measured by Bradford protein assay.

Amino acid standards

GABA, hydroxyproline (HYP), L-Gln, L-Asn, β-Ala, L-cysteine (Cys), and L-Trp were purchased from Wako Chemicals. Purchased amino acids were added to type H amino acid mixture standard solution (Wako Chemicals), to produce a working standard solution of each amino acid (100 µmol/L in 0.1 N HCl).

Amino acid analysis

Free-form amino acids extracted from food samples were analyzed on a Hitachi Ultra High Speed Liquid Chromatography system equipped with UV-Vis detector. Samples were reacted with 4-fluoro-7-nitrobenzofurazan (NBD-F) and derivatized NBD-amino acids were separated on a Zorbax SB-C18 column (3.0 × 50 mm, 2.7 µm; Agilent Technologies) and elution was monitored at 470 nm. Derivatization of amino acids was performed according to the Hitachi Ultra High Speed Liquid Chromatography manual. NBD-amino acids were eluted using a stepwise gradient of increasing B solution: equilibration with 15%, 15-25% for 2.7 min, 25-35% for 3.5 min, 35-70% for 1.0 min, 70-85% for 0.1 min, and kept at 85% for 1.5 min, in sequence. Elution solutions A and B were purchased from Hitachi High-Technologies, where solution A mainly consists of ammonium formate and solution B of acetonitrile. The flow rate was maintained at 0.550 mL/min throughout the analysis.

Linearity of the dose response was evaluated using an amino acid standard solution within the range of 10 to 60 pmol. Minimum and maximum correlation coefficients (r2) for Asn, as an example, were 0.974 and 0.997, respectively. Ten consecutive standard solution injections were analyzed to evaluate reproducibility (Table 3). The relative standard deviation of peak area for Tyr and His was in the range of 0.3% and 5.1%, respectively. The mean limit of detection was 0.8 pmol.

Table 3. Means of peak area and retention time (min) 10 replicates analysis

 

Peak Area

Elution Time (min)

Mean ± SE

%RSD

Mean ± SE

%RSD

His

107839 ± 1724

5.05

0.784 ± 0.001

0.290

Arg

100894 ± 1427

4.47

0.909 ± 0.004

1.250

HYP

113930 ± 589

1.64

1.715 ± 0.004

0.740

Asn

34137 ± 497

4.61

2.003 ± 0.001

0.216

Gln

50739 ± 470

2.93

2.158 ± 0.002

0.224

Cit

79837 ± 342

1.36

2.250 ± 0.001

0.166

Ser

53349 ± 478

2.83

2.331 ± 0.002

0.301

Asp

48268 ± 530

3.47

2.760 ± 0.001

0.156

Glu

51824 ± 418

2.55

2.927 ± 0.001

0.101

Thr

107560 ± 300

0.88

3.073 ± 0.001

0.094

Pro+β-Ala

106552 ± 302

0.90

3.156 ± 0.001

0.132

Gly

90638 ± 637

2.22

3.254 ± 0.002

0.166

GABA

62614 ± 510

2.57

3.551 ± 0.002

0.141

Ala

53683 ± 593

3.49

3.817 ± 0.001

0.124

Val

78636 ± 237

0.95

5.465 ± 0.002

0.139

Met

61485 ± 95

0.49

5.612 ± 0.001

0.080

Leu

68875 ± 316

1.45

6.363 ± 0.002

0.099

Ile

84374 ± 233

0.87

6.519 ± 0.003

0.140

Cys-Cys

56979 ± 197

1.09

6.689 ± 0.002

0.118

Trp

69741 ± 295

1.34

6.820 ± 0.002

0.078

Orn

74663 ± 277

1.17

6.864 ± 0.002

0.089

Phe

80193 ± 134

0.53

6.947 ± 0.002

0.069

Lys

79550 ± 210

0.83

7.026 ± 0.001

0.063

Cys

82425 ± 178

0.68

7.226 ± 0.001

0.029

Tyr

61109 ± 62

0.32

7.450 ± 0.001

0.031

Amino acids are listed as abbreviated according to Table 1.

Results

Chromatograms for the amino acid mixture standard and eggplant "kamonasu" (Solanum melongena) were obtained and analyzed (Figures 1 and 2). In our system, the typical time to baseline NBD-amino acid separation, with the exception of Pro and β-Ala, was about 8 min.

Figure 1. HPLC chromatogram of the amino acids standard mixture derivatized with NBD-F and UV-VIS detection

Figure 2. HPLC chromatogram of the “kamonasu” derivatized with NBD-F and UV-VIS detection

The free-form amino acid contents of the vegetables and mushrooms analyzed varied considerably (Tables 4 and 5). The total free-form amino acids found in the extracts ranged from 59.91 (Garden rhubarb) to 4,093 and 424.6 (Jew's ear) to 12,773 µmol per 100 g of vegetables and mushrooms, respectively. Of the vegetables and mushrooms analyzed, the podded pea (4,093 µmol per 100 g) and dried porcini mushrooms (12,773 µmol per 100 g) had the highest total free-form amino acids of vegetables and mushrooms, respectively. The most abundant free-form amino acids found in the vegetables examined were Asn, Gln, Glu, and GABA, all of which are important metabolites in human. In mushrooms, arginine (Arg), Glu, Gln, and Ala were the most abundant free-form amino acids. Orn was found in mushrooms, but rarely in vegetables and HYP, Cys, and methionine (Met) were found at low levels in both vegetables and mushrooms.

Table 4. Contents of free-form amino acids assayed in vegetables (µmol/100 g sample on a wet weight basis)

 

Okra"Yashiro-shio okura"

Onion

Red onion

Paprika

Edible chrysanthemum 
"Kinkaramatsu"

Edible chrysanthemum
"Enmeiraku"

His

39.5 ± 0.977

16.06 ± 4.234

8.394 ± 0.674

10.68 ± 1.84

13.44 ± 3.137

9.415 ± 1.225

Arg

86.12 ± 2.473

83.6 ± 5.363

53.49 ± 4.528

44.01 ± 2.39

32.75 ± 7.504

17.23 ± 3.145

HYP

nd

nd

nd

0.32 ± 0.261

nd

nd

Asn

1480 ± 40.59

79.14 ± 8.421

87.73 ± 4.638

473.1 ± 45.43

408.1 ± 116

128.1 ± 7.235

Gln

908 ± 2.846

157.3 ± 10.06

286.2 ± 19.35

255.8 ± 23.08

469.3 ± 111.9

596.7 ± 51.47

Cit

nd

nd

nd

nd

nd

nd

Ser

129.1 ± 3.901

15.97 ± 1.216

24.37 ± 1.356

99.13 ± 12.57

53.32 ± 14.24

61.87 ± 4.289

Asp

46.51 ± 0

15.3 ± 1.811

16.42 ± 0.921

83.82 ± 15.04

76.64 ± 14.9

115.3 ± 11.64

Glu

105.3 ± 5.084

29.89 ± 1.902

29.09 ± 1.11

36.91 ± 4.401

98.7 ± 19.51

143.7 ± 9.575

Thr

42.67 ± 1.133

135.9 ± 9.568

95.39 ± 5.964

36.7 ± 7.325

58.42 ± 15.05

55.44 ± 3.696

Pro+β-Ala

18.15 ± 0.806

4.231 ± 0.167

6.233 ± 0.567

19.2 ± 3.532

103.1 ± 17.66

24.09 ± 1.438

Gly

22.59 ± 0.228

4.777 ± 0.204

3.875 ± 0.332

17.28 ± 5.297

6.604 ± 3.816

7.871 ± 0.686

GABA

79.61 ± 11.49

4.982 ± 0.523

2.74 ± 0.237

36.21 ± 4.637

0.831 ± 0.831

nd

Ala

53.62 ± 3.453

14.66 ± 1.369

22.93 ± 1.376

21.45 ± 2.529

17.69 ± 5.758

33.81 ± 3.291

Val

35.36 ± 2.323

19.74 ± 1.286

19.77 ± 0.994

40.05 ± 7.5

26.74 ± 6.647

23.73 ± 2.163

Met

nd

nd

nd

4.971 ± 0.58

nd

nd

Leu

21.44 ± 0

28.89 ± 1.684

31.6 ± 1.652

15.5 ± 1.354

5.079 ± 2.957

nd

Ile

13.34 ± 0.802

15.14 ± 0.701

13.66 ± 1.315

9.44 ± 0.669

14.58 ± 5.23

6.451 ± 0.718

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

16.61 ± 0.042

9.72 ± 1.496

6.809 ± 0.757

nd

3.274 ± 1.375

nd

Orn

nd

nd

nd

nd

nd

nd

Phe

44.37 ± 0.51

21.26 ± 1.237

20.46 ± 1.604

9.36 ± 0.673

21.3 ± 5.56

4.035 ± 0.349

Lys

4.88 ± 0.641

25.19 ± 3.425

27.54 ± 2.864

4.736 ± 2.777

0.456 ± 0.456

4.164 ± 0.705

Cys

0.442 ± 0.312

nd

nd

25.03 ± 5.24

0.647 ± 0.373

nd

Tyr

23.56 ± 0.98

31.61 ± 1.768

20.01 ± 0.273

11.63 ± 0.219

4.63 ± 1.658

nd

 

Winter squash

Japanese squash

Corinky squash

Young carrot (Root)

Young carrot (Leaf)

Moso bamboo

His

74.06 ± 1.997

57.78 ± 1.118

77.19 ± 4.327

0.665 ± 0.47

7.221 ± 0.163

36.11 ± 2.543

Arg

37.9 ± 10.38

15.37 ± 1.592

26.6 ± 3.459

1.141 ± 0.807

0.485 ± 0.343

52.1 ± 5.874

HYP

1.134 ± 0

nd

nd

nd

nd

nd

Asn

82.25 ± 2.595

1025 ± 37.8

162.1 ± 16.04

2.948 ± 0.458

17.41 ± 1.325

995.7 ± 151.2

Gln

90.75 ± 6.974

367.3 ± 14.21

281.9 ± 35.3

16.13 ± 5.61

55.48 ± 6.236

368.6 ± 68.51

Cit

nd

nd

nd

nd

nd

nd

Ser

63.67 ± 3.586

231.2 ± 15.89

90.86 ± 9.239

11.83 ± 0.405

24.09 ± 1.003

232.7 ± 42.35

Asp

192.5 ± 15.57

118.9 ± 5.924

179.7 ± 17.7

18.45 ± 4.924

51.98 ± 3.635

101.7 ± 10.36

Glu

97.7 ± 4.456

95.33 ± 14.87

21.5 ± 2.688

14.09 ± 0.914

59.27 ± 5.786

63.96 ± 6.861

Thr

23.66 ± 2.029

46.92 ± 2.309

44.8 ± 5.556

7.677 ± 0.336

24.43 ± 2.651

103.1 ± 13.78

Pro+β-Ala

14.37 ± 1.468

59.87 ± 1.693

15.03 ± 1.776

1.222 ± 0.096

8.544 ± 1.058

132.6 ± 26.76

Gly

8.521 ± 0.167

34.47 ± 1.265

9.774 ± 1.11

0.286 ± 0.202

2.966 ± 0.239

32.78 ± 4.574

GABA

112.2 ± 10.17

82.02 ± 3.727

70.72 ± 6.325

40.13 ± 6.319

40.4 ± 1.376

75.83 ± 12.81

Ala

43.4 ± 1.337

181.8 ± 9.326

50.15 ± 4.89

9.717 ± 2.299

49.69 ± 4.37

128.9 ± 31.91

Val

14.99 ± 1.734

55.74 ± 2.064

56.93 ± 5.973

5.45 ± 0.333

21.05 ± 2.243

140.8 ± 26.69

Met

7.215 ± 1.251

26.21 ± 1.322

20.58 ± 2.241

nd

nd

44.02 ± 5.688

Leu

40.01 ± 1.073

42.37 ± 5.847

54.94 ± 6

1.852 ± 0.413

5.699 ± 0.683

110.6 ± 11.53

Ile

15.85 ± 2.566

28.14 ± 0.981

35.2 ± 3.677

2.252 ± 0.407

10.44 ± 1.489

88.01 ± 12.15

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

4.827 ± 0.255

20.78 ± 1.077

5.034 ± 0.615

nd

nd

39.6 ± 2.042

Orn

nd

nd

nd

nd

nd

nd

Phe

29.65 ± 1.843

30.89 ± 1.656

41.53 ± 4.733

2.879 ± 0.04

5.631 ± 0.047

55.34 ± 5.782

Lys

2.929 ± 0.313

1.216 ± 0

14.8 ± 2.154

1.134 ± 0.802

4.007 ± 0.486

23.13 ± 2.955

Cys

0.476 ± 0

nd

nd

nd

nd

1.128 ± 0.142

Tyr

34.79 ± 4.246

54.89 ± 3.071

44.66 ± 5.257

2.195 ± 0.367

4.417 ± 0.317

361.9 ± 32.95

 

Pea

Peas “Tutankhamen”

Podded pea

Radish sprout

Garden rhubarb

 

His

49.61 ± 9.239

28.71 ± 1.118

54.13 ± 4.181

70.19 ± 12.28

0.598 ± 0.226

Arg

1304 ± 77.94

703.6 ± 38.32

70.3 ± 12.99

33.05 ± 7.199

1.391 ± 0.452

HYP

nd

nd

nd

1.762 ± 1.205

nd

Asn

291 ± 60.56

171.5 ± 12.55

1257 ± 61.92

60.09 ± 13.23

7.21 ± 1.421

Gln

688.1 ± 50.39

546 ± 60.41

890.1 ± 7.863

438.7 ± 77.95

16.42 ± 1.526

Cit

nd

nd

nd

nd

nd

Ser

153.8 ± 11.01

118.1 ± 9.087

444.1 ± 24.78

60.98 ± 12.41

3.174 ± 0.397

Asp

161.4 ± 10.26

53.84 ± 7.253

114.5 ± 0.625

5.169 ± 1.378

4.756 ± 0.497

Glu

257 ± 18.73

233.9 ± 4.07

84.87 ± 6.466

27.8 ± 3.028

10.9 ± 1.067

Thr

407 ± 30.14

239.9 ± 11.59

355.7 ± 2.423

58.36 ± 11.07

1.149 ± 0.366

Pro+β-Ala

14.21 ± 0.639

28.73 ± 5.658

42.94 ± 8.26

17.98 ± 3.882

0.704 ± 0.231

Gly

35.94 ± 2.387

26.26 ± 0.778

34.17 ± 2.539

40.21 ± 7.198

nd

GABA

105.8 ± 7.17

40.19 ± 1.837

84.76 ± 3.756

9.175 ± 1.978

7.235 ± 0.31

Ala

253.4 ± 27.27

351.5 ± 5.481

387.7 ± 10.44

39.58 ± 6.501

2.866 ± 0.252

Val

159.4 ± 9.839

155.4 ± 3.191

140.8 ± 4.633

50.55 ± 10.05

nd

Met

12.49 ± 0.862

4.772 ± 0.17

8.843 ± 0.528

0.461 ± 0.461

nd

Leu

37.07 ± 2.496

32.55 ± 2.118

25.58 ± 1.322

15.18 ± 2.881

0.16 ± 0.16

Ile

45.71 ± 2.607

50.4 ± 0.83

57.76 ± 6.047

29.32 ± 5.613

nd

Cys-Cys

nd

nd

nd

nd

nd

Trp

3.819 ± 0.777

4.433 ± 0.697

nd

11.58 ± 1.086

nd

Orn

2.935 ± 0.371

2.347 ± 0.009

nd

3.12 ± 0.643

nd

Phe

26.63 ± 2.33

9.983 ± 0.101

15.8 ± 0.034

15.76 ± 2.987

nd

Lys

28.73 ± 3.061

17.24 ± 0.54

13.65 ± 2.164

9.518 ± 1.765

nd

Cys

1.588 ± 0.085

1.596 ± 0.055

1.26 ± 0.069

2.105 ± 0.142

nd

Tyr

49.61 ± 9.239

12.5 ± 0.283

9.671 ± 0.02

16.45 ± 3.433

3.353 ± 0.684

 

Cherry tomato

Eggplant "Beinasu"

Eggplant "Kamonasu"

Eggplant "Kyokonasu"

Eggplant "Marunasu"

Eggplant "Zebranasu"

His

13.39 ± 5.62

31.46 ± 8.565

39.41 ± 3.139

88.56 ± 4.134

15.98 ± 2.025

30.67 ± 1.806

Arg

19.27 ± 5.129

26.41 ± 1.75

80.53 ± 8.906

76.24 ± 2.669

16.53 ± 4.35

90.2 ± 8.215

HYP

nd

nd

2.386 ± 0.125

nd

nd

nd

Asn

166.6 ± 71.7

355.4 ± 28.42

472.1 ± 63.92

992.8 ± 29.96

98.11 ± 19.69

152 ± 10.4

Gln

472.9 ± 214.5

206 ± 22.98

269.1 ± 36.24

130.6 ± 7.851

89.43 ± 13.8

204.1 ± 10.87

Cit

nd

nd

nd

nd

nd

nd

Ser

48 ± 16.45

10.26 ± 0.677

25.77 ± 2.991

45.42 ± 2.354

10.78 ± 1.583

18.25 ± 0.891

Asp

81.52 ± 41.2

29.51 ± 2.832

21.19 ± 0.597

60.67 ± 3.534

21.88 ± 2.575

25.66 ± 4.704

Glu

687.4 ± 290.4

20.54 ± 1.197

20.41 ± 0.961

32.95 ± 2.064

17.63 ± 2.032

16.21 ± 1.467

Thr

24.85 ± 7.489

23.64 ± 1.108

33.09 ± 3.141

62.48 ± 0.968

18.51 ± 2.113

20.87 ± 1.14

Pro+β-Ala

22.29 ± 1.618

12.98 ± 0.407

17.22 ± 1.132

30.29 ± 1.987

9.24 ± 1.061

18.5 ± 2.429

Gly

7.274 ± 2.767

4.403 ± 0.265

9.574 ± 0.55

7.413 ± 0.306

4.307 ± 0.463

7.901 ± 0.476

GABA

93.59 ± 55.17

86.84 ± 7.584

221.5 ± 18.1

74.7 ± 3.888

184.2 ± 15.42

83.94 ± 6.821

Ala

37.04 ± 16.65

3.619 ± 0.614

6.567 ± 0.482

9.47 ± 0.887

5.975 ± 0.901

8.224 ± 1.059

Val

8.005 ± 3.296

21.67 ± 0.835

45.94 ± 2.538

61.48 ± 0.896

20.98 ± 2.597

35.03 ± 1.74

Met

3.382 ± 1.15

nd

nd

nd

nd

nd

Leu

19.01 ± 6.342

4.738 ± 0.545

6.691 ± 0.134

10.81 ± 0.493

3.824 ± 0.9

16.46 ± 1.966

Ile

7.223 ± 2.122

12.46 ± 0.639

20.22 ± 1.233

27.8 ± 0.593

11.12 ± 1.615

23.61 ± 1.435

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

1.438 ± 4.39

4.056 ± 0.645

7.87 ± 1.805

14.45 ± 1.306

2.713 ± 0.182

4.411 ± 0.285

Orn

nd

1.772 ± 0.13

7.206 ± 2.17

5.766 ± 0.577

1.46 ± 0.072

9.496 ± 1.016

Phe

27.81 ± 6.499

17.34 ± 0.356

37.74 ± 1.647

97.05 ± 2.003

12.98 ± 1.54

18.1 ± 1.283

Lys

8.565 ± 4.092

12.77 ± 1.369

10.92 ± 0.899

30.36 ± 1.895

8.691 ± 1.686

9.977 ± 1.494

Cys

7.715 ± 0.878

nd

nd

1.626 ± 0

nd

nd

Tyr

11.72 ± 2.1

8.541 ± 0.919

11.25 ± 1.104

31.58 ± 0.765

7.867 ± 1.1

9.702 ± 0.373

 

Spinach "Okame"

Broad bean

Cowpea

Cowpea beans

 

His

nd

126.6 ± 23.9

38.08 ± 6.123

163.6 ± 4.632

Arg

0.372 ± 0.372

654.7 ± 54.74

38.2 ± 7.142

162.4 ± 3.91

HYP

2.259 ± 1.311

nd

nd

nd

Asn

26.24 ± 1.131

945 ± 66.41

613 ± 30.48

392.6 ± 25.88

Gln

35.31 ± 9.173

154.1 ± 4.755

551.4 ± 28.61

248.4 ± 8.96

Cit

nd

2.675 ± 1.693

nd

nd

Ser

24.18 ± 1.985

142.1 ± 9.879

73.27 ± 2.145

72.32 ± 1.861

Asp

48.59 ± 3.46

146.2 ± 13.99

46.51 ± 0

46.51 ± 0

Glu

85.43 ± 7.502

181.6 ± 11.21

46.48 ± 1.804

265.7 ± 7.392

Thr

9.409 ± 1.653

143.3 ± 14.71

45.86 ± 0.463

90.39 ± 1.414

Pro+β-Ala

4.336 ± 0.561

36.5 ± 2.915

18.17 ± 0.047

67.29 ± 3.878

Gly

1.165 ± 0.681

31.17 ± 1.757

6.967 ± 0.301

16.37 ± 0.785

GABA

22.75 ± 1.147

226.6 ± 16.13

34.2 ± 1.668

15.39 ± 1.526

Ala

21.94 ± 2.911

654.5 ± 64.95

31.8 ± 4.071

125.9 ± 3.136

Val

4.754 ± 1.102

79.84 ± 2.213

41.82 ± 1.037

120 ± 4.047

Met

nd

1.7 ± 0.558

8.761 ± 0.393

25.52 ± 0.461

Leu

3.405 ± 0.362

27.39 ± 0.715

21.44 ± 0

21.44 ± 0

Ile

nd

25.34 ± 0.754

23.29 ± 0.467

74.98 ± 2.901

Cys-Cys

5.861 ± 3.487

nd

nd

nd

Trp

nd

32.18 ± 2.075

3.813 ± 0.47

18.24 ± 1.499

Orn

nd

nd

nd

3.388 ± 0.124

Phe

3.291 ± 0.61

11.6 ± 0.318

20.45 ± 0.577

58.47 ± 0.536

Lys

nd

60.26 ± 0.73

7.725 ± 0.576

17.8 ± 0.988

Cys

109.3 ± 0

1.153 ± 0.126

nd

2.286 ± 0.136

Tyr

12.14 ± 1.411

60.24 ± 3.935

18.83 ± 1.493

72.12 ± 2.174

Amino acids are listed as abbreviated according to Table 1.

Average of 2~5 analyses (Means ± SE), nd: Not detected.

Table 5. Contents of free-form amino acids assayed in mushrooms (µmol/100 g sample on a wet weight basis)

 

Beech mushroom

Beech mushroom
(Tanba)

Button mushroom
(brown)

Button mushroom
(white)

Jew's ear

King oyster
mushroom

His

29.38 ± 7.477

99.7 ± 5.005

43.63 ± 2.66

133.7 ± 3.481

9.663 ± 1.963

80.38 ± 3.689

Arg

149.8 ± 8.816

311.2 ± 10.49

90.49 ± 2.751

99.8 ± 4.613

57.94 ± 13.16

302.6 ± 13.05

HYP

2.095 ± 1.236

nd

1.15 ± 0.734

1.847 ± 1.173

nd

13.52 ± 0.789

Asn

290.9 ± 31.47

113 ± 2.854

80.17 ± 3.27

178.1 ± 9.544

72.1 ± 12.47

106.5 ± 3.763

Gln

149.8 ± 10.74

283.2 ± 17.75

322.7 ± 12.09

802.7 ± 41.74

68.81 ± 14.11

382.9 ± 15.61

Cit

nd

nd

nd

nd

nd

nd

Ser

60.76 ± 4.634

184.2 ± 7.715

101.5 ± 3.719

182.6 ± 8.61

14.09 ± 2.211

151.8 ± 3.506

Asp

6.673 ± 0.615

7.738 ± 0.887

204.5 ± 8.97

210.3 ± 13.68

27.32 ± 6.626

37.37 ± 9.767

Glu

222.9 ± 21.88

62.48 ± 7.219

299.3 ± 13.01

372.5 ± 29.14

26.04 ± 3.239

190.1 ± 16.83

Thr

66.86 ± 4.54

121.2 ± 5.029

131.5 ± 5.477

206.8 ± 8.645

11.01 ± 1.4

145.7 ± 4.064

Pro+β-Ala

24.26 ± 1.547

84.83 ± 2.042

97.13 ± 3.355

127.2 ± 4.381

2.392 ± 0.317

64.84 ± 1.564

Gly

53.36 ± 3.844

141.4 ± 8.039

88.39 ± 3.428

163.2 ± 10.44

8.306 ± 1.602

129.2 ± 10.84

GABA

nd

98.47 ± 7.632

59.07 ± 2.68

140.9 ± 20.12

32.61 ± 8.053

35.12 ± 0.667

Ala

86.72 ± 8.508

123.5 ± 9.437

240.8 ± 9.665

598.3 ± 50.35

43.95 ± 11.21

507.6 ± 61.07

Val

33.9 ± 3.485

85.08 ± 5.763

94.7 ± 3.492

151.4 ± 8.963

4.24 ± 2.007

103.8 ± 6.616

Met

nd

15.67 ± 0.605

5.684 ± 0.26

8.567 ± 0.423

 

4.674 ± 0.199

Leu

39.65 ± 4.621

96.53 ± 6.645

93.28 ± 3.86

144.5 ± 10.15

8.475 ± 3.134

144 ± 21.41

Ile

18.91 ± 1.756

46.8 ± 2.982

46.45 ± 1.692

83.98 ± 5.49

3.667 ± 2.126

75.18 ± 6.733

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

nd

nd

nd

32.86 ± 1.011

nd

14.15 ± 0.772

Orn

156.3 ± 20.96

242.2 ± 25.8

110.1 ± 8.37

98.03 ± 15.28

23.93 ± 8.903

199.2 ± 29.78

Phe

38.54 ± 2.916

103.1 ± 3.677

100.2 ± 3.305

118.4 ± 4.251

nd

73.4 ± 1.9

Lys

12.8 ± 1.983

18.78 ± 3.628

25.74 ± 2.02

44.98 ± 7.77

6.433 ± 3.884

52.82 ± 9.439

Cys

nd

4.687 ± 0.72

nd

 

nd

1.773 ± 0.293

Tyr

24.48 ± 1.605

75.12 ± 4.388

65.53 ± 2.636

50.22 ± 2.619

3.627 ± 0.616

76.28 ± 2.503

 

Mushroom "Honshimeji"

Mushroom "Maitake"

Mushroom "Shiitake"

Porcini mushrooms (Dried)

Winter mushroom

 

His

178.5 ± 10.14

41.05 ± 2.678

123.7 ± 5.593

527.7 ± 54.46

118.2 ± 3.371

 

Arg

410.6 ± 18.82

47.35 ± 2.468

226.4 ± 12.59

724.1 ± 35.73

114 ± 3.03

 

HYP

nd

0.312 ± 0.312

nd

244.2 ± 43.95

19.8 ± 1.552

 

Asn

44.37 ± 3.853

16.44 ± 1.439

87.64 ± 3.504

1277 ± 142.8

101.7 ± 2.697

 

Gln

755.9 ± 55.49

119.2 ± 9.674

1237 ± 83.49

629.6 ± 72.69

414.9 ± 11.71

 

Cit

nd

nd

nd

nd

nd

 

Ser

93.53 ± 8.26

35.49 ± 3.123

97.06 ± 4.664

1499 ± 83.72

78.06 ± 2.904

 

Asp

25.04 ± 0.899

11.71 ± 0.468

187.3 ± 15.24

325.5 ± 230.1

19.19 ± 3.483

 

Glu

125.7 ± 4.428

75.59 ± 4.98

180.1 ± 13.53

462.6 ± 84.5

95.03 ± 3.556

 

Thr

113.4 ± 8.527

42.18 ± 3.339

169.8 ± 10.55

694.9 ± 26.45

113.1 ± 4.342

 

Pro+β-Ala

87.9 ± 6.65

17.79 ± 2.266

71.66 ± 1.074

396.4 ± 42.3

68.92 ± 3.138

 

Gly

155.5 ± 6.678

21.9 ± 1.938

255.7 ± 15.22

1222 ± 144

123 ± 2.055

 

GABA

46.12 ± 1.717

0.467 ± 0.467

80.73 ± 4.854

1378 ± 91.32

183.9 ± 5.589

 

Ala

185.9 ± 6.48

92.2 ± 6.107

113.7 ± 6.708

2088 ± 148.1

252.6 ± 5.196

 

Val

66.5 ± 3.585

21.51 ± 1.703

128.8 ± 7.157

262.2 ± 40.33

95.39 ± 1.62

 

Met

10.53 ± 1.111

nd

3.345 ± 1.156

346.9 ± 99.25

5.32 ± 0.205

 

Leu

63.38 ± 3.125

8.217 ± 0.811

80.65 ± 4.762

65.07 ± 0.218

115.6 ± 5.381

 

Ile

31.56 ± 1.551

11.55 ± 0.373

51.86 ± 2.891

92.72 ± 23.11

60.88 ± 1.216

 

Cys-Cys

nd

nd

nd

nd

nd

 

Trp

nd

nd

nd

103.6 ± 18.91

20.72 ± 0.598

 

Orn

274.4 ± 13.8

62.71 ± 1.961

156 ± 14.37

66.86 ± 11.24

85.33 ± 2.289

 

Phe

70.26 ± 4.225

22.54 ± 0.784

58.04 ± 2.473

123.4 ± 23.3

217.3 ± 5.183

 

Lys

22.34 ± 1.081

7.209 ± 0.438

49.39 ± 5.58

96.48 ± 7.537

27.24 ± 1.79

 

Cys

nd

nd

0.566 ± 0.19

8.504 ± 6.013

nd

 

Tyr

39.86 ± 1.959

24.42 ± 1.461

12.55 ± 2.442

136.9 ± 17.97

142.8 ± 6.058

 

 Amino acids are listed as abbreviated according to Table 1.

The free-form amino acid compositions of five different kinds of eggplant were analyzed. Slight differences were observed in the free-form amino acid contents of the five species, but Asn, Gln, and GABA were abundant in all.

BCAAs, common ingredients in drinks designed for athletes, were not detected in most foods analyzed. However, moso bamboo and some peas, including the podded pea, contained considerable amounts of Val, Leu, and Ile BCAAs.

Discussion

Here, we identified and measured significant levels of free-form amino acids in both vegetables and mushrooms. These free-form amino acids should contribute to the taste and palatability of foods. Indeed, Glu provides umami taste and has been shown to improve appetite [12,14,15], both of which contribute to QOL. Glu is particularly important in the foods of elderly and sick people, as it improves QOL by increasing nutrient intake. It is often discussed that excess intake of amino acids is toxic for human. For example, it has been warned that Glu has a potential risk of neurotoxicity like headache. However, this warning was almost eliminated after the joint report of FAO, WHO, and the Expert Committee on Food Additives (JECFA) in 2000 [28]. Thus, Glu is considered relatively safe and could be consumed without any concerns.

GABA is known for health-promoting functions, including an anti-hypotensive effect. Some studies suggest that supplementation with 10~20 mg (equivalent to 97~194 µmol) GABA reduces blood pressure in human [29,30]. Commercially available GABA enriched "Tokuho" foods are typically supplemented with 20 mg of GABA in Japan. Our present study suggests that two pieces of eggplant “kamonasu”, equivalent to 90 g, per day provides enough GABA to maintain blood pressure at a normal level.

Free-form amino acids are significantly beneficial to human health. The presence of these amino acids in vegetables and mushrooms suggests that they also provide a benefit to those species. In plants, free-form amino acids are likely absorbed directly from soil via the roots [31]. Some of these free-form amino acids are metabolized or stored, and some are utilized as signaling molecules to regulate cellular functions including enzyme activity, gene expression, and redox-homeostasis [32]. Pro, Arg, Met, Glu, and GABA are involved in the regulation of plant responses to various stress conditions [33-35]. Asn, the most abundant free-form amino acid in the foods analyzed in our study, is accumulated under stress conditions or mineral deficiencies [36]. In plants, free-form Asn also plays a central role in nitrogen storage and transport [36,37]. The roles of free-form amino acids in mushrooms are not well understood and are still being determined.

Here, we have demonstrated the presence of free-form amino acids in plants. However, there are many factors that influence the distribution of amino acids, including cultivation conditions, geographic production location, picking season, breed, and soil. In this study, we were unable to examine, or control, these variables. Nevertheless, we believe our results provide ample information with which to establish a free-form amino acid database to serve and inform nutritional studies. 

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Editorial Information

Editor-in-Chief

Renee Dufault
Food Ingredient and Health Research Institute

Article Type

Research Article

Publication history

Received date: March 17, 2017
Accepted date: April 13, 2017
Published date: April 17, 2017

Copyright

© 2017 Ito H. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Citation

Ito H, Ueno H, Kikuzaki H (2017) Construction of a free-form amino acid database for vegetables and mushrooms. Integr Food Nutr Metab 4: DOI: 10.15761/IFNM.1000186

Corresponding author

Hiroshi Ueno

Laboratory of Biochemistry and Microbiology, School of Agriculture, Ryukoku University, Seta, Otsu 520-2194, Japan

Figure 1. HPLC chromatogram of the amino acids standard mixture derivatized with NBD-F and UV-VIS detection

Figure 2. HPLC chromatogram of the “kamonasu” derivatized with NBD-F and UV-VIS detection

Table 1. List of amino acids and their abbreviations

Amino acid

3 Letter abbreviation

Alanine

Ala

β-Alanine

β-Ala

Arginine

Arg

Asparagine

Asn

Aspartic acid

Asp

Citrulline

Cit

Cysteine

Cys

Cystine

Cys-Cys

γ-Amino butyric acid

GABA

Glutamic acid

Glu

Glutamine

Gln

Glycine

Gly

Histidine

His

Hydroxyproline

HYP

Isoleucine

Ile

Leucine

Leu

Lysine

Lys

Methionine

Met

Ornithine

Orn

Phenyl alanine

Phe

Proline

Pro

Serine

Ser

Threonine

Thr

Tryptophan

Trp

Tyrosine

Tyr

Valine

Val

Table 2. List of foods analyzed for free-form amino acids

 

Scientific Name

Food

Vegetables

Abelmoschus esculentus

Okra “Yashiro-shio okura”

Allium cepa

Onion

Allium cepa

Red onion

Capsicum annuum

Paprika

Chrysanthemum morifolium

Edible chrysanthemum

Chrysanthemum morifolium

Edible purple chrysanthemum “Enmeiraku”

Cucurbita maxima

Winter squash

Cucurbita moschata

Japanese squash

Daucus carota

young carrot (Leaf)

Daucus carota

Moso bamboo

Phyllostachys heterocycla

Pea

Pisum sativum

Peas (Tutankhamen)

Pisum sativum

Podded pea

Raphanus sativus

Radish sprout

Rheum rhabarbarum

Garden rhubarb

Solanum mlycopersicum

Cherry tomato

Solanum melongena

Egg plant “Beinasu”

Solanum melongena

Egg plant “Kamonasu”

Solanum melongena

Egg plant “Kyokonasu”

Solanum melongena

Egg plant “Marunasu”

Solanum melongena

Egg plant “Zebranasu”

Spinacia olerace

Spinach “Okame”

Vicia faba

Broad bean

Vigna unguiculata

Cowpea

Vigna unguiculata

 

Cowpea beans

Pumpkin “Korinnki”

 Mushrooms

Agaricus bisporus

Button mushroom (brown)

Agaricus bisporus

Button mushroom (white)

Auricularia auricula-judae

Jew’s ear

Boletus edulis

Porcini mushrooms (Dried)

Flammulina velutipes

Winter mushroom

Grifola frondosa

Mushroom “Maitake”

Hypsizygus marmoreus

 Beech mushroom

Hypsizygus marmoreus

 Beech mushroom(Tanba)

Letinula edodes

 Mushroom “Shiitake”

Lyophyllum shimeji

 Mushroom “Honshimeji”

Pleurotus eryngii

King oyster mushroom

Table 3. Means of peak area and retention time (min) 10 replicates analysis

 

Peak Area

Elution Time (min)

Mean ± SE

%RSD

Mean ± SE

%RSD

His

107839 ± 1724

5.05

0.784 ± 0.001

0.290

Arg

100894 ± 1427

4.47

0.909 ± 0.004

1.250

HYP

113930 ± 589

1.64

1.715 ± 0.004

0.740

Asn

34137 ± 497

4.61

2.003 ± 0.001

0.216

Gln

50739 ± 470

2.93

2.158 ± 0.002

0.224

Cit

79837 ± 342

1.36

2.250 ± 0.001

0.166

Ser

53349 ± 478

2.83

2.331 ± 0.002

0.301

Asp

48268 ± 530

3.47

2.760 ± 0.001

0.156

Glu

51824 ± 418

2.55

2.927 ± 0.001

0.101

Thr

107560 ± 300

0.88

3.073 ± 0.001

0.094

Pro+β-Ala

106552 ± 302

0.90

3.156 ± 0.001

0.132

Gly

90638 ± 637

2.22

3.254 ± 0.002

0.166

GABA

62614 ± 510

2.57

3.551 ± 0.002

0.141

Ala

53683 ± 593

3.49

3.817 ± 0.001

0.124

Val

78636 ± 237

0.95

5.465 ± 0.002

0.139

Met

61485 ± 95

0.49

5.612 ± 0.001

0.080

Leu

68875 ± 316

1.45

6.363 ± 0.002

0.099

Ile

84374 ± 233

0.87

6.519 ± 0.003

0.140

Cys-Cys

56979 ± 197

1.09

6.689 ± 0.002

0.118

Trp

69741 ± 295

1.34

6.820 ± 0.002

0.078

Orn

74663 ± 277

1.17

6.864 ± 0.002

0.089

Phe

80193 ± 134

0.53

6.947 ± 0.002

0.069

Lys

79550 ± 210

0.83

7.026 ± 0.001

0.063

Cys

82425 ± 178

0.68

7.226 ± 0.001

0.029

Tyr

61109 ± 62

0.32

7.450 ± 0.001

0.031

Amino acids are listed as abbreviated according to Table 1.

Table 4. Contents of free-form amino acids assayed in vegetables (µmol/100 g sample on a wet weight basis)

 

Okra"Yashiro-shio okura"

Onion

Red onion

Paprika

Edible chrysanthemum 
"Kinkaramatsu"

Edible chrysanthemum
"Enmeiraku"

His

39.5 ± 0.977

16.06 ± 4.234

8.394 ± 0.674

10.68 ± 1.84

13.44 ± 3.137

9.415 ± 1.225

Arg

86.12 ± 2.473

83.6 ± 5.363

53.49 ± 4.528

44.01 ± 2.39

32.75 ± 7.504

17.23 ± 3.145

HYP

nd

nd

nd

0.32 ± 0.261

nd

nd

Asn

1480 ± 40.59

79.14 ± 8.421

87.73 ± 4.638

473.1 ± 45.43

408.1 ± 116

128.1 ± 7.235

Gln

908 ± 2.846

157.3 ± 10.06

286.2 ± 19.35

255.8 ± 23.08

469.3 ± 111.9

596.7 ± 51.47

Cit

nd

nd

nd

nd

nd

nd

Ser

129.1 ± 3.901

15.97 ± 1.216

24.37 ± 1.356

99.13 ± 12.57

53.32 ± 14.24

61.87 ± 4.289

Asp

46.51 ± 0

15.3 ± 1.811

16.42 ± 0.921

83.82 ± 15.04

76.64 ± 14.9

115.3 ± 11.64

Glu

105.3 ± 5.084

29.89 ± 1.902

29.09 ± 1.11

36.91 ± 4.401

98.7 ± 19.51

143.7 ± 9.575

Thr

42.67 ± 1.133

135.9 ± 9.568

95.39 ± 5.964

36.7 ± 7.325

58.42 ± 15.05

55.44 ± 3.696

Pro+β-Ala

18.15 ± 0.806

4.231 ± 0.167

6.233 ± 0.567

19.2 ± 3.532

103.1 ± 17.66

24.09 ± 1.438

Gly

22.59 ± 0.228

4.777 ± 0.204

3.875 ± 0.332

17.28 ± 5.297

6.604 ± 3.816

7.871 ± 0.686

GABA

79.61 ± 11.49

4.982 ± 0.523

2.74 ± 0.237

36.21 ± 4.637

0.831 ± 0.831

nd

Ala

53.62 ± 3.453

14.66 ± 1.369

22.93 ± 1.376

21.45 ± 2.529

17.69 ± 5.758

33.81 ± 3.291

Val

35.36 ± 2.323

19.74 ± 1.286

19.77 ± 0.994

40.05 ± 7.5

26.74 ± 6.647

23.73 ± 2.163

Met

nd

nd

nd

4.971 ± 0.58

nd

nd

Leu

21.44 ± 0

28.89 ± 1.684

31.6 ± 1.652

15.5 ± 1.354

5.079 ± 2.957

nd

Ile

13.34 ± 0.802

15.14 ± 0.701

13.66 ± 1.315

9.44 ± 0.669

14.58 ± 5.23

6.451 ± 0.718

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

16.61 ± 0.042

9.72 ± 1.496

6.809 ± 0.757

nd

3.274 ± 1.375

nd

Orn

nd

nd

nd

nd

nd

nd

Phe

44.37 ± 0.51

21.26 ± 1.237

20.46 ± 1.604

9.36 ± 0.673

21.3 ± 5.56

4.035 ± 0.349

Lys

4.88 ± 0.641

25.19 ± 3.425

27.54 ± 2.864

4.736 ± 2.777

0.456 ± 0.456

4.164 ± 0.705

Cys

0.442 ± 0.312

nd

nd

25.03 ± 5.24

0.647 ± 0.373

nd

Tyr

23.56 ± 0.98

31.61 ± 1.768

20.01 ± 0.273

11.63 ± 0.219

4.63 ± 1.658

nd

 

Winter squash

Japanese squash

Corinky squash

Young carrot (Root)

Young carrot (Leaf)

Moso bamboo

His

74.06 ± 1.997

57.78 ± 1.118

77.19 ± 4.327

0.665 ± 0.47

7.221 ± 0.163

36.11 ± 2.543

Arg

37.9 ± 10.38

15.37 ± 1.592

26.6 ± 3.459

1.141 ± 0.807

0.485 ± 0.343

52.1 ± 5.874

HYP

1.134 ± 0

nd

nd

nd

nd

nd

Asn

82.25 ± 2.595

1025 ± 37.8

162.1 ± 16.04

2.948 ± 0.458

17.41 ± 1.325

995.7 ± 151.2

Gln

90.75 ± 6.974

367.3 ± 14.21

281.9 ± 35.3

16.13 ± 5.61

55.48 ± 6.236

368.6 ± 68.51

Cit

nd

nd

nd

nd

nd

nd

Ser

63.67 ± 3.586

231.2 ± 15.89

90.86 ± 9.239

11.83 ± 0.405

24.09 ± 1.003

232.7 ± 42.35

Asp

192.5 ± 15.57

118.9 ± 5.924

179.7 ± 17.7

18.45 ± 4.924

51.98 ± 3.635

101.7 ± 10.36

Glu

97.7 ± 4.456

95.33 ± 14.87

21.5 ± 2.688

14.09 ± 0.914

59.27 ± 5.786

63.96 ± 6.861

Thr

23.66 ± 2.029

46.92 ± 2.309

44.8 ± 5.556

7.677 ± 0.336

24.43 ± 2.651

103.1 ± 13.78

Pro+β-Ala

14.37 ± 1.468

59.87 ± 1.693

15.03 ± 1.776

1.222 ± 0.096

8.544 ± 1.058

132.6 ± 26.76

Gly

8.521 ± 0.167

34.47 ± 1.265

9.774 ± 1.11

0.286 ± 0.202

2.966 ± 0.239

32.78 ± 4.574

GABA

112.2 ± 10.17

82.02 ± 3.727

70.72 ± 6.325

40.13 ± 6.319

40.4 ± 1.376

75.83 ± 12.81

Ala

43.4 ± 1.337

181.8 ± 9.326

50.15 ± 4.89

9.717 ± 2.299

49.69 ± 4.37

128.9 ± 31.91

Val

14.99 ± 1.734

55.74 ± 2.064

56.93 ± 5.973

5.45 ± 0.333

21.05 ± 2.243

140.8 ± 26.69

Met

7.215 ± 1.251

26.21 ± 1.322

20.58 ± 2.241

nd

nd

44.02 ± 5.688

Leu

40.01 ± 1.073

42.37 ± 5.847

54.94 ± 6

1.852 ± 0.413

5.699 ± 0.683

110.6 ± 11.53

Ile

15.85 ± 2.566

28.14 ± 0.981

35.2 ± 3.677

2.252 ± 0.407

10.44 ± 1.489

88.01 ± 12.15

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

4.827 ± 0.255

20.78 ± 1.077

5.034 ± 0.615

nd

nd

39.6 ± 2.042

Orn

nd

nd

nd

nd

nd

nd

Phe

29.65 ± 1.843

30.89 ± 1.656

41.53 ± 4.733

2.879 ± 0.04

5.631 ± 0.047

55.34 ± 5.782

Lys

2.929 ± 0.313

1.216 ± 0

14.8 ± 2.154

1.134 ± 0.802

4.007 ± 0.486

23.13 ± 2.955

Cys

0.476 ± 0

nd

nd

nd

nd

1.128 ± 0.142

Tyr

34.79 ± 4.246

54.89 ± 3.071

44.66 ± 5.257

2.195 ± 0.367

4.417 ± 0.317

361.9 ± 32.95

 

Pea

Peas “Tutankhamen”

Podded pea

Radish sprout

Garden rhubarb

 

His

49.61 ± 9.239

28.71 ± 1.118

54.13 ± 4.181

70.19 ± 12.28

0.598 ± 0.226

Arg

1304 ± 77.94

703.6 ± 38.32

70.3 ± 12.99

33.05 ± 7.199

1.391 ± 0.452

HYP

nd

nd

nd

1.762 ± 1.205

nd

Asn

291 ± 60.56

171.5 ± 12.55

1257 ± 61.92

60.09 ± 13.23

7.21 ± 1.421

Gln

688.1 ± 50.39

546 ± 60.41

890.1 ± 7.863

438.7 ± 77.95

16.42 ± 1.526

Cit

nd

nd

nd

nd

nd

Ser

153.8 ± 11.01

118.1 ± 9.087

444.1 ± 24.78

60.98 ± 12.41

3.174 ± 0.397

Asp

161.4 ± 10.26

53.84 ± 7.253

114.5 ± 0.625

5.169 ± 1.378

4.756 ± 0.497

Glu

257 ± 18.73

233.9 ± 4.07

84.87 ± 6.466

27.8 ± 3.028

10.9 ± 1.067

Thr

407 ± 30.14

239.9 ± 11.59

355.7 ± 2.423

58.36 ± 11.07

1.149 ± 0.366

Pro+β-Ala

14.21 ± 0.639

28.73 ± 5.658

42.94 ± 8.26

17.98 ± 3.882

0.704 ± 0.231

Gly

35.94 ± 2.387

26.26 ± 0.778

34.17 ± 2.539

40.21 ± 7.198

nd

GABA

105.8 ± 7.17

40.19 ± 1.837

84.76 ± 3.756

9.175 ± 1.978

7.235 ± 0.31

Ala

253.4 ± 27.27

351.5 ± 5.481

387.7 ± 10.44

39.58 ± 6.501

2.866 ± 0.252

Val

159.4 ± 9.839

155.4 ± 3.191

140.8 ± 4.633

50.55 ± 10.05

nd

Met

12.49 ± 0.862

4.772 ± 0.17

8.843 ± 0.528

0.461 ± 0.461

nd

Leu

37.07 ± 2.496

32.55 ± 2.118

25.58 ± 1.322

15.18 ± 2.881

0.16 ± 0.16

Ile

45.71 ± 2.607

50.4 ± 0.83

57.76 ± 6.047

29.32 ± 5.613

nd

Cys-Cys

nd

nd

nd

nd

nd

Trp

3.819 ± 0.777

4.433 ± 0.697

nd

11.58 ± 1.086

nd

Orn

2.935 ± 0.371

2.347 ± 0.009

nd

3.12 ± 0.643

nd

Phe

26.63 ± 2.33

9.983 ± 0.101

15.8 ± 0.034

15.76 ± 2.987

nd

Lys

28.73 ± 3.061

17.24 ± 0.54

13.65 ± 2.164

9.518 ± 1.765

nd

Cys

1.588 ± 0.085

1.596 ± 0.055

1.26 ± 0.069

2.105 ± 0.142

nd

Tyr

49.61 ± 9.239

12.5 ± 0.283

9.671 ± 0.02

16.45 ± 3.433

3.353 ± 0.684

 

Cherry tomato

Eggplant "Beinasu"

Eggplant "Kamonasu"

Eggplant "Kyokonasu"

Eggplant "Marunasu"

Eggplant "Zebranasu"

His

13.39 ± 5.62

31.46 ± 8.565

39.41 ± 3.139

88.56 ± 4.134

15.98 ± 2.025

30.67 ± 1.806

Arg

19.27 ± 5.129

26.41 ± 1.75

80.53 ± 8.906

76.24 ± 2.669

16.53 ± 4.35

90.2 ± 8.215

HYP

nd

nd

2.386 ± 0.125

nd

nd

nd

Asn

166.6 ± 71.7

355.4 ± 28.42

472.1 ± 63.92

992.8 ± 29.96

98.11 ± 19.69

152 ± 10.4

Gln

472.9 ± 214.5

206 ± 22.98

269.1 ± 36.24

130.6 ± 7.851

89.43 ± 13.8

204.1 ± 10.87

Cit

nd

nd

nd

nd

nd

nd

Ser

48 ± 16.45

10.26 ± 0.677

25.77 ± 2.991

45.42 ± 2.354

10.78 ± 1.583

18.25 ± 0.891

Asp

81.52 ± 41.2

29.51 ± 2.832

21.19 ± 0.597

60.67 ± 3.534

21.88 ± 2.575

25.66 ± 4.704

Glu

687.4 ± 290.4

20.54 ± 1.197

20.41 ± 0.961

32.95 ± 2.064

17.63 ± 2.032

16.21 ± 1.467

Thr

24.85 ± 7.489

23.64 ± 1.108

33.09 ± 3.141

62.48 ± 0.968

18.51 ± 2.113

20.87 ± 1.14

Pro+β-Ala

22.29 ± 1.618

12.98 ± 0.407

17.22 ± 1.132

30.29 ± 1.987

9.24 ± 1.061

18.5 ± 2.429

Gly

7.274 ± 2.767

4.403 ± 0.265

9.574 ± 0.55

7.413 ± 0.306

4.307 ± 0.463

7.901 ± 0.476

GABA

93.59 ± 55.17

86.84 ± 7.584

221.5 ± 18.1

74.7 ± 3.888

184.2 ± 15.42

83.94 ± 6.821

Ala

37.04 ± 16.65

3.619 ± 0.614

6.567 ± 0.482

9.47 ± 0.887

5.975 ± 0.901

8.224 ± 1.059

Val

8.005 ± 3.296

21.67 ± 0.835

45.94 ± 2.538

61.48 ± 0.896

20.98 ± 2.597

35.03 ± 1.74

Met

3.382 ± 1.15

nd

nd

nd

nd

nd

Leu

19.01 ± 6.342

4.738 ± 0.545

6.691 ± 0.134

10.81 ± 0.493

3.824 ± 0.9

16.46 ± 1.966

Ile

7.223 ± 2.122

12.46 ± 0.639

20.22 ± 1.233

27.8 ± 0.593

11.12 ± 1.615

23.61 ± 1.435

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

1.438 ± 4.39

4.056 ± 0.645

7.87 ± 1.805

14.45 ± 1.306

2.713 ± 0.182

4.411 ± 0.285

Orn

nd

1.772 ± 0.13

7.206 ± 2.17

5.766 ± 0.577

1.46 ± 0.072

9.496 ± 1.016

Phe

27.81 ± 6.499

17.34 ± 0.356

37.74 ± 1.647

97.05 ± 2.003

12.98 ± 1.54

18.1 ± 1.283

Lys

8.565 ± 4.092

12.77 ± 1.369

10.92 ± 0.899

30.36 ± 1.895

8.691 ± 1.686

9.977 ± 1.494

Cys

7.715 ± 0.878

nd

nd

1.626 ± 0

nd

nd

Tyr

11.72 ± 2.1

8.541 ± 0.919

11.25 ± 1.104

31.58 ± 0.765

7.867 ± 1.1

9.702 ± 0.373

 

Spinach "Okame"

Broad bean

Cowpea

Cowpea beans

 

His

nd

126.6 ± 23.9

38.08 ± 6.123

163.6 ± 4.632

Arg

0.372 ± 0.372

654.7 ± 54.74

38.2 ± 7.142

162.4 ± 3.91

HYP

2.259 ± 1.311

nd

nd

nd

Asn

26.24 ± 1.131

945 ± 66.41

613 ± 30.48

392.6 ± 25.88

Gln

35.31 ± 9.173

154.1 ± 4.755

551.4 ± 28.61

248.4 ± 8.96

Cit

nd

2.675 ± 1.693

nd

nd

Ser

24.18 ± 1.985

142.1 ± 9.879

73.27 ± 2.145

72.32 ± 1.861

Asp

48.59 ± 3.46

146.2 ± 13.99

46.51 ± 0

46.51 ± 0

Glu

85.43 ± 7.502

181.6 ± 11.21

46.48 ± 1.804

265.7 ± 7.392

Thr

9.409 ± 1.653

143.3 ± 14.71

45.86 ± 0.463

90.39 ± 1.414

Pro+β-Ala

4.336 ± 0.561

36.5 ± 2.915

18.17 ± 0.047

67.29 ± 3.878

Gly

1.165 ± 0.681

31.17 ± 1.757

6.967 ± 0.301

16.37 ± 0.785

GABA

22.75 ± 1.147

226.6 ± 16.13

34.2 ± 1.668

15.39 ± 1.526

Ala

21.94 ± 2.911

654.5 ± 64.95

31.8 ± 4.071

125.9 ± 3.136

Val

4.754 ± 1.102

79.84 ± 2.213

41.82 ± 1.037

120 ± 4.047

Met

nd

1.7 ± 0.558

8.761 ± 0.393

25.52 ± 0.461

Leu

3.405 ± 0.362

27.39 ± 0.715

21.44 ± 0

21.44 ± 0

Ile

nd

25.34 ± 0.754

23.29 ± 0.467

74.98 ± 2.901

Cys-Cys

5.861 ± 3.487

nd

nd

nd

Trp

nd

32.18 ± 2.075

3.813 ± 0.47

18.24 ± 1.499

Orn

nd

nd

nd

3.388 ± 0.124

Phe

3.291 ± 0.61

11.6 ± 0.318

20.45 ± 0.577

58.47 ± 0.536

Lys

nd

60.26 ± 0.73

7.725 ± 0.576

17.8 ± 0.988

Cys

109.3 ± 0

1.153 ± 0.126

nd

2.286 ± 0.136

Tyr

12.14 ± 1.411

60.24 ± 3.935

18.83 ± 1.493

72.12 ± 2.174

Amino acids are listed as abbreviated according to Table 1.

Average of 2~5 analyses (Means ± SE), nd: Not detected.

Table 5. Contents of free-form amino acids assayed in mushrooms (µmol/100 g sample on a wet weight basis)

 

Beech mushroom

Beech mushroom
(Tanba)

Button mushroom
(brown)

Button mushroom
(white)

Jew's ear

King oyster
mushroom

His

29.38 ± 7.477

99.7 ± 5.005

43.63 ± 2.66

133.7 ± 3.481

9.663 ± 1.963

80.38 ± 3.689

Arg

149.8 ± 8.816

311.2 ± 10.49

90.49 ± 2.751

99.8 ± 4.613

57.94 ± 13.16

302.6 ± 13.05

HYP

2.095 ± 1.236

nd

1.15 ± 0.734

1.847 ± 1.173

nd

13.52 ± 0.789

Asn

290.9 ± 31.47

113 ± 2.854

80.17 ± 3.27

178.1 ± 9.544

72.1 ± 12.47

106.5 ± 3.763

Gln

149.8 ± 10.74

283.2 ± 17.75

322.7 ± 12.09

802.7 ± 41.74

68.81 ± 14.11

382.9 ± 15.61

Cit

nd

nd

nd

nd

nd

nd

Ser

60.76 ± 4.634

184.2 ± 7.715

101.5 ± 3.719

182.6 ± 8.61

14.09 ± 2.211

151.8 ± 3.506

Asp

6.673 ± 0.615

7.738 ± 0.887

204.5 ± 8.97

210.3 ± 13.68

27.32 ± 6.626

37.37 ± 9.767

Glu

222.9 ± 21.88

62.48 ± 7.219

299.3 ± 13.01

372.5 ± 29.14

26.04 ± 3.239

190.1 ± 16.83

Thr

66.86 ± 4.54

121.2 ± 5.029

131.5 ± 5.477

206.8 ± 8.645

11.01 ± 1.4

145.7 ± 4.064

Pro+β-Ala

24.26 ± 1.547

84.83 ± 2.042

97.13 ± 3.355

127.2 ± 4.381

2.392 ± 0.317

64.84 ± 1.564

Gly

53.36 ± 3.844

141.4 ± 8.039

88.39 ± 3.428

163.2 ± 10.44

8.306 ± 1.602

129.2 ± 10.84

GABA

nd

98.47 ± 7.632

59.07 ± 2.68

140.9 ± 20.12

32.61 ± 8.053

35.12 ± 0.667

Ala

86.72 ± 8.508

123.5 ± 9.437

240.8 ± 9.665

598.3 ± 50.35

43.95 ± 11.21

507.6 ± 61.07

Val

33.9 ± 3.485

85.08 ± 5.763

94.7 ± 3.492

151.4 ± 8.963

4.24 ± 2.007

103.8 ± 6.616

Met

nd

15.67 ± 0.605

5.684 ± 0.26

8.567 ± 0.423

 

4.674 ± 0.199

Leu

39.65 ± 4.621

96.53 ± 6.645

93.28 ± 3.86

144.5 ± 10.15

8.475 ± 3.134

144 ± 21.41

Ile

18.91 ± 1.756

46.8 ± 2.982

46.45 ± 1.692

83.98 ± 5.49

3.667 ± 2.126

75.18 ± 6.733

Cys-Cys

nd

nd

nd

nd

nd

nd

Trp

nd

nd

nd

32.86 ± 1.011

nd

14.15 ± 0.772

Orn

156.3 ± 20.96

242.2 ± 25.8

110.1 ± 8.37

98.03 ± 15.28

23.93 ± 8.903

199.2 ± 29.78

Phe

38.54 ± 2.916

103.1 ± 3.677

100.2 ± 3.305

118.4 ± 4.251

nd

73.4 ± 1.9

Lys

12.8 ± 1.983

18.78 ± 3.628

25.74 ± 2.02

44.98 ± 7.77

6.433 ± 3.884

52.82 ± 9.439

Cys

nd

4.687 ± 0.72

nd

 

nd

1.773 ± 0.293

Tyr

24.48 ± 1.605

75.12 ± 4.388

65.53 ± 2.636

50.22 ± 2.619

3.627 ± 0.616

76.28 ± 2.503

 

Mushroom "Honshimeji"

Mushroom "Maitake"

Mushroom "Shiitake"

Porcini mushrooms (Dried)

Winter mushroom

 

His

178.5 ± 10.14

41.05 ± 2.678

123.7 ± 5.593

527.7 ± 54.46

118.2 ± 3.371

 

Arg

410.6 ± 18.82

47.35 ± 2.468

226.4 ± 12.59

724.1 ± 35.73

114 ± 3.03

 

HYP

nd

0.312 ± 0.312

nd

244.2 ± 43.95

19.8 ± 1.552

 

Asn

44.37 ± 3.853

16.44 ± 1.439

87.64 ± 3.504

1277 ± 142.8

101.7 ± 2.697

 

Gln

755.9 ± 55.49

119.2 ± 9.674

1237 ± 83.49

629.6 ± 72.69

414.9 ± 11.71

 

Cit

nd

nd

nd

nd

nd

 

Ser

93.53 ± 8.26

35.49 ± 3.123

97.06 ± 4.664

1499 ± 83.72

78.06 ± 2.904

 

Asp

25.04 ± 0.899

11.71 ± 0.468

187.3 ± 15.24

325.5 ± 230.1

19.19 ± 3.483

 

Glu

125.7 ± 4.428

75.59 ± 4.98

180.1 ± 13.53

462.6 ± 84.5

95.03 ± 3.556

 

Thr

113.4 ± 8.527

42.18 ± 3.339

169.8 ± 10.55

694.9 ± 26.45

113.1 ± 4.342

 

Pro+β-Ala

87.9 ± 6.65

17.79 ± 2.266

71.66 ± 1.074

396.4 ± 42.3

68.92 ± 3.138

 

Gly

155.5 ± 6.678

21.9 ± 1.938

255.7 ± 15.22

1222 ± 144

123 ± 2.055

 

GABA

46.12 ± 1.717

0.467 ± 0.467

80.73 ± 4.854

1378 ± 91.32

183.9 ± 5.589

 

Ala

185.9 ± 6.48

92.2 ± 6.107

113.7 ± 6.708

2088 ± 148.1

252.6 ± 5.196

 

Val

66.5 ± 3.585

21.51 ± 1.703

128.8 ± 7.157

262.2 ± 40.33

95.39 ± 1.62

 

Met

10.53 ± 1.111

nd

3.345 ± 1.156

346.9 ± 99.25

5.32 ± 0.205

 

Leu

63.38 ± 3.125

8.217 ± 0.811

80.65 ± 4.762

65.07 ± 0.218

115.6 ± 5.381

 

Ile

31.56 ± 1.551

11.55 ± 0.373

51.86 ± 2.891

92.72 ± 23.11

60.88 ± 1.216

 

Cys-Cys

nd

nd

nd

nd

nd

 

Trp

nd

nd

nd

103.6 ± 18.91

20.72 ± 0.598

 

Orn

274.4 ± 13.8

62.71 ± 1.961

156 ± 14.37

66.86 ± 11.24

85.33 ± 2.289

 

Phe

70.26 ± 4.225

22.54 ± 0.784

58.04 ± 2.473

123.4 ± 23.3

217.3 ± 5.183

 

Lys

22.34 ± 1.081

7.209 ± 0.438

49.39 ± 5.58

96.48 ± 7.537

27.24 ± 1.79

 

Cys

nd

nd

0.566 ± 0.19

8.504 ± 6.013

nd

 

Tyr

39.86 ± 1.959

24.42 ± 1.461

12.55 ± 2.442

136.9 ± 17.97

142.8 ± 6.058

 

 Amino acids are listed as abbreviated according to Table 1.