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Banana fruits with high content of resistant starch: Effect of genotypes and phosphorus fertilization
Journal Pre-proofs Banana fruits with high content of resistant starch: effect of genotypes and phosphorus fertilization Magali Leonel, Ana Carolina Batista Bolfarini, Marlon Jocimar Rodrigues da Silva, Jackson Mirellys Azevêdo Souza, Sarita Leonel PII: DOI: Reference:
S0141-8130(19)37156-9 https://doi.org/10.1016/j.ijbiomac.2019.10.217 BIOMAC 13718
To appear in:
International Journal of Biological Macromolecules
Received Date: Accepted Date:
4 September 2019 24 October 2019
Please cite this article as: M. Leonel, A. Carolina Batista Bolfarini, M. Jocimar Rodrigues da Silva, J. Mirellys Azevêdo Souza, S. Leonel, Banana fruits with high content of resistant starch: effect of genotypes and phosphorus fertilization, International Journal of Biological Macromolecules (2019), doi: https://doi.org/ 10.1016/j.ijbiomac.2019.10.217
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© 2019 Published by Elsevier B.V.
1 1
Banana fruits with high content of resistant starch: effect of genotypes and phosphorus
2
fertilization
3 4
Magali Leonela,*,[email protected], Ana Carolina Batista Bolfarinib, Marlon Jocimar Rodrigues
5
da Silvab, Jackson Mirellys Azevêdo Souzab, Sarita Leonelb
6 7
aCenter
8
Botucatu, São Paulo 18610-307, Brazil
9
bSchool
for Tropical Roots and Starches (CERAT), São Paulo State University (UNESP),
of Agriculture, São Paulo State University (FCA/UNESP),
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Botucatu, São Paulo 18610-307, Brazil
11
*Corresponding
12
Highlights
13
Phosphorus plays an important role in starch accumulation in bananas;
14
Response to phosphate fertilizer is genotype dependent
15
Phosphate fertilization increases resistant starch content in green banana fruits
16
author.
2 17 18
Abstract
19
Banana fertilization practices aim to increase agricultural yield and the effects of these
20
practices on fruit components have not been well explored. This study aimed to evaluate the
21
effects of phosphate fertilizer levels on production parameters and accumulation of dry
22
matter, phosphorus, starch and resistant starch in green fruits. Four banana genotypes were
23
cultivated under the same cultural practices, with varying levels of phosphate fertilizers
24
during three production cycles. Bunches were harvested and evaluated for total fruit mass,
25
average finger mass and yield. Fresh green fruit pulps were analyzed for dry matter,
26
phosphorus, starch and resistant starch content. The results showed that the effects of
27
phosphate fertilizer were genotype dependent and that the increase of P2O5 rates applied in
28
banana fertilization promoted an increase in fruit production with higher levels of phosphorus,
29
starch and resistant starch.
30
Keywords: Musa spp; mineral nutrition; resistant starch
31 32
3 33
1. Introduction
34
Banana, as a climacteric fruit, has a very short period of post-harvest
35
commercialization, leading to considerable economic losses, which has been increasing the
36
use of green bananas as a source of functional ingredients [1, 2].
37
In most of the Brazilian regions there are favorable soil conditions for banana
38
production, but not always the chemical and physical properties of the soils are the most
39
adequate, reflecting in low quality and fruit productivity. Thus, fertility and soil physical
40
structure are fundamental factors [3].
41
Phosphorus (P) is an essential nutrient for plants and it is important in breathing,
42
energy production and transformation processes by the plant, as well as in cell division. This
43
nutrient is quickly mobilized in plants and, when there is a deficiency, is translocated from
44
older tissues to active meristematic regions [4].
45
The adequate supply of P is therefore essential from the early stages of plant growth
46
and the application of adequate P doses enables root development, besides increasing water
47
and nutrients uptake, which can contribute to fruiting [5,6].
48
Banana crop demands a high amount of fertilizers, not only because of the high
49
accumulation and the export of nutrients, but also because it is commonly grown on soils with
50
lower fertility. The phosphorus rates recommended in banana cultivation vary from 30 to 120
51
g of P2O5 per planting pit, depending on the available P content and soil texture. However,
52
banana absorbs most of the required P between three and nine months after planting, and
53
reduces to 80% in the reproductive phase [7].
54
In addition to the phosphorus functions already mentioned, this nutrient play important
55
roles in starch biosynthesis and in functional properties of the starches. Photosynthesis and
56
sucrose synthesis-related genes have been shown to change their expression following
57
phosphorus deficiency. Phosphate esters are covalently bound to starch granules, mainly in
4 58
amylopectin molecule, providing specific properties for starches and thus making them more
59
suitable for different industrial applications [5, 8].
60
Starch is the main component of unripe bananas, which undergoes important changes
61
during ripening. The average starch content drops from 70 to 80% in the pre-climacteric
62
period (prior to starch breakdown) to less than 1% at the end of the climacteric period [2].
63
Starch occurs naturally in the form of granules in plant cells and is mainly composed
64
of two polymers: amylopectin and amylose. Amylose is a linear polymer composed of
65
glucopyranose units linked through α-D- (1 → 4) glycosidic linkages, while amylopectin is a
66
highly branched polymer of high molecular weight [9].
67
Based on in vitro digestibility starch may be categorized into rapidly digestible starch
68
(RDS), slowly digestible starch (SDS), and resistant starch (RS). Resistant starch (RS) has
69
been defined as the starch fraction which escapes digestion in the small intestine of healthy
70
individuals [10].
71
Resistant starch occurs because of its inaccessibility to amylase. Depending on the
72
nature of inaccessibility, there are five classes: RS1 is found in grains and the granule may be
73
encased inside cells or in a strong protein matrix; RS2 consists of raw starch granules that
74
resist amylase digestion; RS3 is retrograded starch; RS4 is chemically modified starch, and
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RS5 is amylose complexed with lipid [11, 12, 13].
76
The use of green banana pulp as a source of resistant starch involves the study of
77
several interfering factors, such as the particular characteristics of each banana genotype, as
78
well as the interference of the growing conditions on the productivity and the intrinsic
79
characteristics of the starch.
80
Considering the growing market for functional foods and the importance of banana in
81
the multiple aspects involved in food security, this study evaluated the effect of levels of
82
phosphate fertilizers applied to the cultivation of four banana genotypes on parameters of
5 83
productivity, dry matter accumulation, phosphorus, total starch and resistant starch in fruit in
84
order to provide useful information for the expansion of production and marketing of green
85
banana pulp.
86 87
2. Material and Methods
88
2.1. Banana genotypes
89
The banana genotype Prata Anã is from the genomic group AAB. It is a very vigorous
90
plant and does not require shoring. It presents size of 2.0 to 3.5 m, good tillering, vegetative
91
cycle of 280 days and average yield of 15 t/ha. It presents small bunches (14 kg), with 100
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fruits/bunch and fruits of 13 cm with 110g of weight. It is tolerant to cold, susceptible to
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Yellow and Black Sigatoka, and to bacterial wilt (Moko), moderately susceptible to fusarium
94
wilt (Panama disease), and moderately resistant to nematodes and banana root borer [14].
95
The cultivar Fhia 18, a hybrid of PrataAnã, is tetraploid (AAAB) and was developed
96
by the Honduran Agricultural Research Foundation [15]. (FHIA, 2018). The plant presents
97
medium size, a vegetative cycle of 353 days, good tillering and average yield of 20 t/ha. Its
98
fruits are straight, thick with medium size. This cultivar is resistant to black Sigatoka and
99
Panama disease, moderately resistant to yellow Sigatoka and susceptible to Moko and banana
100
root borer [14].
101
The genotypes 'Nanicão-IAC-2001' and 'Grand Naine' are of the genomic group (AAA)
102
and show similar characteristics regarding the physical fruits characteristics. These genotypes
103
are susceptible to yellow and black Sigatoka and resistant to Panama disease [14].
104 105
2.2. Banana growth
106
The cultivation of the four banana genotypes was conducted in Red Latosol at
107
Experimental Farm from School of Agriculture (FCA) that belongs to UNESP, located in the
6 108
city of São Manuel, state of São Paulo (22°44'28"S 48°34'37"W, at an altitude of 740m). The
109
climate is humid subtropical (Cfa), that is, temperate hot (mesothermic), with concentrated
110
rains from November to April (summer) and average annual rainfall of 1376.70 mm; the
111
city´s average temperature of the warmest month exceeds 22°C [16]. Figure 1 show the
112
temperature and rainfall data in the experimental area during the study.
113
Soil samples were collected at a depth 0-20cm to determine their chemical properties:
114
pH in CaCl2, 5.5; M.O., 12 g dm-3; P resin, 16 mg dm-3; H+Al, 15 mmolc dm-3; K, 1 mmolc
115
dm-3; Ca, 13 mmolc dm-3; Mg, 5 mmolc dm-3; SB, 19 mmolc dm-3; CTC, 34 mmolc dm-3;
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V%, 57; S, 1 mg dm-3.
117
In November 2012, it was held the transplanting of micro propagated bananas plantlets
118
adopting a spacing of 4 m between rows and 2.5 m between plants, totaling 1000 plant ha-1. It
119
was opened planting holes in the dimensions of 60 cm in diameter by 60 cm in depth and
120
were applied 10 L of cattle manure and half of the recommended doses of phosphorus for
121
planting. The other half was applied 80 days after planting [17]. Also 0.5 kg of dolomitic
122
limestone was added to raise the base saturation index to 60% and the magnesium content to 9
123
mmolc dm-3 [18].
124
The experiment was conducted during three production cycles. The first production
125
cycle happened from November 2012 to April 2014; while the second from August 2014 to
126
February 2015; and the third from June 2015 to January 2016.
127
Fertilization of production with phosphorus (40 kg P2O5 - 100%) was standardized
128
according to soil chemical analyzes and expected yield of less than 20 t ha-1 [17]. Therefore,
129
P fertilization doses were: no application (control); 20 kg P2O5 ha-1 year-1; 30 kg P2O5 ha-1
130
year-1; 40 kg P2O5 ha-1 year-1; 50 kg P2O5 ha-1 year-1; and 60 kg P2O5 ha-1 year-1. Furthermore,
131
triple superphosphate (46% P2O5) was used as the source of phosphorus. The applications
132
realized in the form of semicircle (100 cm radius) in front of the youngest shoot. These
7 133
production fertilizers were carried out in January 2014, August 2014 and June 2015 for the
134
first, second and third cycles of production, respectively.
135
Supplemental fertilizations were 600 g of ammonium sulphate and 550 g of potassium
136
chloride per plant, divided in the course of the cycle, 20% at 35 days after planting (DAP),
137
50% at 80 DAP and 30% at 130 DAP [18]. The fertilizers were applied in circles of 100 cm in
138
diameter around the plant. At the time of thinning it was carried out the fertilization with 10 g
139
of boric acid in the open hole in the rhizome of the mother plant.
140
In the course of the experiment were performed: irrigation, weed control, thinning and
141
removal of dried leaves, fertilization, pest control and disease, elimination of banana heart,
142
removal of pistils and cutting pseudo stem after harvesting.
143 144
2.3. Production analysis
145
Fruits at maturity stage 1 (fully green peel) of five plants of each genotype in each
146
treatment were harvested. In order to standardize the maturation stage of the banana cultivars,
147
the ratio (ºBrix/titratable acidity) of the fruits was also determined.
148
The following variables were evaluated: mass of bunch (kg), total mass of fruits (kg),
149
average mass of fingers (g) and yield (t ha-1). The yield was calculated considering the weight
150
of the bunch in a stand of 1000 plants ha-1.
151 152
2.4. Fruits characteristics
153
The green fruits of each banana genotype, grown under the different conditions of
154
phosphate fertilization, were evaluated for phosphorus [19], dry matter [20], total starch and
155
resistant starch in the three cycles.
156
For the analysis of starch fractions in banana pulp samples of immature fresh fruits
157
(0.5 g) were disintegrated with liquid nitrogen and then treated with successive washes of
8 158
80% alcohol, 50% alcohol and water to remove sugar soluble and other soluble substrates
159
[21]. Determinations of resistant starch and total starch contents followed the methodology
160
described by Gõni et al. [22].
161 162
2.5. Experimental design and statistical analysis
163
The experimental design was completely randomized systematized in subdivided
164
plots, with the plots represented by the five rates of phosphate fertilization and the subplots by
165
the four banana cultivars, with 10 plants per experimental plot and 6 replicates in blocks,
166
totaling 60 plants of each genotype.
167
In order to study only the effect of phosphate fertilization and genotypes, the crop
168
cycles factor was eliminated. For this, we calculated the average of the results for each
169
production cycle, then being carried out the analysis of these averages.
170
The data were submitted to the test of homogeneity and normality, analysis of variance
171
(test F) and, when significant, the means of treatment of the P2O5 rates for each genotype were
172
expressed in regressions for the effects of rates of fertilizer for all variables. The criterion for
173
choosing the model was the significance of the F test up to 5% of probability and the highest
174
value of the determination coefficient (R2). Aiming to have a better visualization of the
175
physicochemical profile of the fruits of the four banana cultivars, regardless of the applied
176
phosphorus doses, the principal component analysis (PCA) was performed.
177 178
3. Results and discussion
179
There was a significant interaction between the banana cultivars and the phosphorus
180
doses for all variables analyzed (Figures 1 and 2). Thus, the effects of the two factors were
181
jointly analyzed.
182
9 183
3.1. Production analysis
184
The effect of phosphate fertilizer levels on the bunch mass, total fruit mass and fruit
185
mass was very similar in all banana cultivars studied. For all of them, the quadratic regression
186
model was better fitted, and it was verified that the highest values of the mentioned
187
production characteristics were observed when 35 kg of P2O5 ha-1 were applied. Fertilizer
188
doses lower and higher than this provided lower results (Figure 2).
189
The quadratic effect in response to increasing phosphorus fertilizer levels on this
190
parameter also were observed in other studies [23,24]. However, Silva and Rodrigues [7]
191
evaluating five rates of triple superphosphate (0, 50, 100, 200 and 300 g of P2O5) applied
192
during cultivation of ‘PrataAnã’ (AAB) in a clayey Red Latosol with low phosphorus
193
availability reported a linear increase in bunch mass in function of the P2O5 rates.
194
Despite the variations in the regression models, the results observed by these authors
195
corroborate with those obtained in this study, showing that the inadequate supply of nutrients
196
due to the use of low fertility soils has been one of the main causes of obtaining low yields in
197
banana plants. The perennial vegetative and reproductive growth of this plant requires the
198
adequate supply of nutrients throughout the crop cycle. The phosphorus absorbed by the plant
199
participates in several metabolic processes, including energy transfer, synthesis of nucleic
200
acids and starch, respiration, membrane synthesis and stability, activation and deactivation of
201
enzymes, redox reactions and carbohydrate metabolism [25,26], actions directly related to the
202
increase of productivity.
203
Significant differences in all productive characteristics were observed among the
204
cultivars studied. Regardless of the fertilizer rate, the highest values of bunch mass, total fruit
205
mass and fruit mass were obtained in the cultivar Grand Nine, followed by the cultivars
206
Nanicão and FHIA 18. On the other hand, the cultivar Prata Anã showed the lowest values for
207
production parameters. The differences observed between the banana genotypes for the
10 208
phosphate fertilizer could be due to their mechanisms of adaptation to the growing conditions,
209
as well as due to their genetic characteristics. It is known that Cavendish bananas (Grande
210
Naine and Nanicão), are generally more productive than Prata (Prata Anã and FHIA 18).
211
Plants growing at different P concentrations develop adaptive mechanisms including
212
changes in morphology and architecture of the root system as well as in physiological
213
characteristics of roots. A more developed and extensive root system allows greater
214
absorption of less mobile ions, such as phosphate [27, 28]. Thus, lower levels of phosphate
215
fertilization are required to increase the parameters of productivity.
216
Besides this, the increase in production parameters observed in this study may be
217
related to the important effects of phosphorus on photosynthesis. Photosynthesis and sucrose
218
synthesis-related genes have been shown to change their expression following P deficiency,
219
indicating their involvement in the observed increase in growth [5].
220
Another point is that in the adequate levels in the soil the phosphorus stimulates root
221
growth and development, allowing greater efficiency in water utilization and ionic absorption
222
of other elements. However, elevation of salinity and toxicity provided by high concentrations
223
of phosphate fertilizer reduce root growth and, consequently, affect nutrient uptake, which
224
may justify the decrease in production parameters with the use of fertilizer rates higher than
225
40 kg of P2O5 ha-1 year-1.
226
So, results obtained by adequate levels of phosphorus fertilizer in banana culture is so
227
important because as discussed by Zhu et al. [29] in their revision about phosphorus in
228
agricultural soils, there are a number of adverse environmental impacts associated with the
229
excessive use of inorganic phosphate fertilizers. These include increasing risk of P loss from
230
soils with elevated phosphorus concentrations, which may lead to eutrophication in water
231
bodies and the depletion of finite resources of high grade phosphate rocks.
11 232
In view of the above, it is evident the need to study the real need for phosphate
233
fertilization by the different genotypes. It was observed a better productive performance of all
234
genotypes at doses below the recommended level (40 kg P2O5 kg-1 year-1), especially in the
235
cultivar Grand Naine, in which the highest bunch weight was obtained with 27.2 kg P2O5 kg-1
236
year-1, i.e. only 68% of the recommended dose.
237 238
3.2. Fruits characteristics
239
The analysis of the levels of dry matter, phosphorus, total starch and resistant starch in
240
the fruits of the four banana genotypes showed differences among the cultivars within each
241
level of phosphate fertilization and the effect of the P2O5 rates used on these components in
242
the unripe fruits (Figure 3).
243
When we analyzed the effects of phosphorus doses on the physicochemical
244
characteristics of banana fruits, it was possible to observe that the quadratic regression models
245
were the best fit for all cultivars studied. The data analysis showed that the increase of
246
phosphate fertilizer rates applied in banana cultivation promoted an increasing in dry matter,
247
phosphorus, total and resistant starch contents. However, in the higher concentrations there
248
was a decreasing in the contents of these components. As all these parameters are directly
249
related to the metabolism of the plants and the phosphorus is intrinsically linked to it, it is
250
important to emphasize that in the extreme conditions of low or high availability of
251
phosphorus several control mechanisms are activated in the plant, reflecting in the
252
accumulation of dry matter, in the starch content and its structure characteristics, as well as, in
253
the accumulation of this mineral in the fruit.
254
When the supply of phosphorus is limited, plants grow more roots, increase the rate of
255
uptake by roots from the soil, retranslocate Pi from older leaves, and deplete the vacuolar
256
stores of Pi. In addition, mycorrhizal fungi may more extensively colonize the roots.
12 257
Conversely, when plants have an adequate supply of Pi and are absorbing it at rates that
258
exceed demand, a number of processes act to prevent the accumulation of toxic Pi
259
concentrations [30].
260
In general, the highest contents of dry matter in the fruits were obtained when the
261
bananas were fertilized with 40 kg of P2O5 ha-1. Using this dose, we observed that the fruits of
262
the cultivars Nanicão and Prata Anã averaged 42.16% of dry matter, differing significantly
263
from fruits of the cultivars Grand Naine and FHIA 18, which, with the same dose of the
264
fertilizer, presented about 38% dry matter (Fig. 2A).
265
The effect of phosphorus fertilizer rates on fingers dry matter showed that the
266
adequate availability of this nutrient in the soil influenced the production and allocation of
267
assimilates in the fruits. On the other hand, the lower phosphorus availability in soil with the
268
use of the half levels of fertilizer recommendation (20 kg of P2O5 ha-1 year-1)may have
269
decreased the dry matter content by reducing the radiation intercepted during the crop cycle
270
(IR), as cited by Jenkins and Ali [31] in their study with potatoes.
271
The levels of phosphorus in banana fruits ranged from 31.62 to 42.45 mg 100 g-1, with
272
differences between cultivars and fertilizer rates (Fig. 2B). The highest accumulations of
273
phosphorus in the fruits occurred with fertilization doses of 40 Kg ha-1 for the Grand Nine,
274
around 43 Kg ha-1 for the Nanicão and FHIA 18, and 47 Kg ha-1 for Prata Anã. Within the
275
evaluated dose of 40 kg ha-1, there was no significant difference in phosphorus content in the
276
Nanicão and Grand Naine cultivars, with a mean of 41.7 mg 100 g-1. However, these cultivars
277
differed significantly from the cultivars Prata Anã and FHIA 18, which showed on average
278
35.2 mg 100 g-1 of phosphorus in fruits.
279
For all cultivars, the increase of the fertilizer rates led to an increasing of the
280
phosphorus content, however, in the higher levels it was not observed an increasing in the
13 281
phosphorus levels in fruits pulps. Applying higher rates of phosphate fertilizer may result in
282
considerable immobilization, leaving part of the phosphate inaccessible to the plants [32].
283
The differences in phosphorus content in fruits, as well as the response to fertilization
284
among cultivars may be related to the root system. In a more developed and deeper root
285
system there is an increase in the contact area between the roots of the banana and the soil,
286
which favors the absorption of the P by the plant.
287
This increase in the phosphorus content in the fruits pulps is very important since in
288
the composition of a diet the phosphorus comes mainly from the animal protein sources.
289
Phosphorus is an essential nutrient in human health and has a variety of physiological roles.
290
The increase of its content in the banana fruits leads to nutritional benefits, since phosphorus
291
has the function of buffering acidic or alkaline systems, assisting in the maintenance of pH, in
292
the temporary storage of energy derived from macronutrient metabolism, in the form of ATP,
293
being responsible for the activation through the phosphorylation of several enzymatic
294
cascades [33].
295
The increase in the doses of phosphate fertilization led to a limited increase in the
296
content of starch in the banana fruits. For the cultivars Grand Naine and Prata Anã, the
297
highest content of starch in the fruits was obtained when the fertilization was carried out with
298
37 kg of P2O5 ha-1. On the other hand, fruits of the cultivars Nanicão and FHIA 18 showed the
299
maximum values for starch content with higher doses of the fertilizer, that is, with 44 kg of
300
P2O5 ha-1, with reduction in the starch content in fruits in higher doses of fertilizer.
301
These results evidenced the important participation of phosphorus in the biosynthesis
302
of starch in plants. AGPase is a key regulatory enzyme that catalyzes the rate limiting step of
303
starch biosynthesis and it is extremely sensitive to allosteric regulation, with glycerate-3-
304
phosphate (3-PGA) acting as an activator and Pi as an inhibitor [34].
14 305
Zhao et al. [26] in their study with the effects of nitrogen and phosphorus on
306
transcriptional regulation of genes encoding key enzymes of starch metabolism in duckweed
307
described that the activities of AGPase and soluble starch synthesis enzyme (SSS EC
308
2.4.1.21) were positively correlated with changes in starch content.
309
The global starch market is poised to register in the period of 2015 to 2023 a
310
Compound Annual Growth Rate (CAGR) of 5.85%. The market is competitive and driven by
311
an increase in the trend of “health and wellness” and growing consumer demand for all
312
natural ingredients [35]. Thus, the possibility of increasing the resistant starch content by
313
agricultural management in banana cultivation may allow Brazil a commercial differential.
314
The levels of resistant starch in the fresh pulps of unripe bananas ranged from 17.0 to
315
29.8 g 100 g-1. Studies related that banana starches have B-type crystalline pattern and
316
starches with this type of crystallinity tend to be more resistant to pancreatic amylases. This
317
increased strength is due to branches and associations of amylopectin and amylose [14, 36,
318
37].
319
Phosphate fertilization rates had effect on this fraction of the starch for all genotypes.
320
The maximum values for resistant starch contents were obtained with fertilizations at the
321
doses of 40 kg of P2O5 ha-1 for Prata Anã, around 42 kg of P2O5 ha-1 for the cultivars Nanicão
322
and FHIA 18 and 54 kg of P2O5 ha-1 to Grand Naine. The changes in phosphorus levels in
323
unripe fruits pulp can have promoted changed in phosphorus linked in starch structure with
324
can have led to a disruption of the hydrogen bonds of the amylopectin double helix reducing
325
the crystalline region of the starches and interfering in the resistant starch content in the
326
banana fruits.
327
Interference of phosphorus in starch digestibility has been evaluated and some studies
328
related that as amylolytic enzymes are incapable of bypassing the phosphorylated glucosyl
15 329
residue, phosphoryl-oligosaccharides are released from the digestion of starch with amylase
330
[38, 39].
331
The effect of phosphate fertilizer on the characteristics of banana starch was evaluated
332
by Mesquita et al. [40] and these authors observed effect of phosphate fertilization on the
333
cristallinity of the banana starch (genotype Maçã). The increase in phosphorus fertilizer led to
334
a decreasing of cristallinity of the starch and resistant starch content.
335 336
3.3. Principal components analysis
337
The effect of the different doses of phosphorus was clearly evidenced by Figures 2 and
338
3, which also showed the effects of the banana cultivars in each dose of the evaluated
339
fertilizer. However, in order to better understand the physicochemical profile of the fruits of
340
the four banana cultivars, independent of the doses of phosphate fertilizer, the principal
341
components analysis was applied, and the results showed that only the first two components
342
(PC1 and PC2) explained 93.89% of total variability (Figure 4).
343
Analyzing PC1, which explained 65.26% of the data, we observed that the fruits of the
344
cultivar Prata Anã were clearly separated from the other cultivars, mainly from Grand Naine,
345
which was positioned on the opposite side in PC1 (Figure 4A). According to PC1 loadings,
346
except for the phosphorus content, all other variables strongly contributed to this separation
347
(Figure 4B). PC1 scores and loadings indicate that the cultivar Prata Anã showed better
348
quality fruits, with a higher content of dry matter and higher concentrations of starch, total
349
and resistant. However, this cultivar was less productive, presenting lower values of bunch
350
mass and fruit, individual and total. An opposite result was observed in the cultivar Grand
351
Naine, which presented high productive indexes, however with fruits with low starch
352
contents. In addition, the analysis showed that the cultivar Nanicão was similar to Grand
16 353
Naine, but this cultivar, together with FHIA 18, showed intermediate values because they
354
were closer to the central axis of PC1.
355
The PC2, which explained 28.63% of the variability of the experiment, was evidenced
356
mainly by the phosphorus content in the fruits, as can be visualized in PC2 loadings, showing
357
the importance of this element in the distinction of the banana cultivars.
358
The analysis of PC2 scores and loadings especially suggests a separation of the
359
cultivars Nanicão and FHIA 18, since the highest levels of phosphorus were obtained in
360
bananas of the cultivar Nanicão and smaller in the fruits of FHIA 18. Thus, it can also be
361
observed that this element was observed in intermediate concentrations in the fruits of the
362
cultivars Prata Anã and Grand Naine.
363 364
4. Conclusion
365
Results obtained in this study showed that, through the management of phosphate
366
fertilization, it is possible to increase the agricultural production of green banana fruits with
367
higher levels of phosphorus and resistant starch, allowing a substantial increase in the
368
commercial value of banana pulp by the growing market in the food industry. Therefore, the
369
management of the fertilization for banana crop can be directed to obtain not only high yield
370
but also in view of higher levels of functional ingredients. PrataAnã banana stands out as a
371
source of dry matter and high resistant starch content. On the other hand, Grande Naine is the
372
most productive, but with low levels of starch and dry mass.
373 374 375 376 377 378 379
Conflict of interest The authors declare no conflict of interest. Author contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript.
17 380
Acknowledgement
381
The authors thank CNPQ for financially supporting this work (Processes 303373/2014-8,
382
302827/2017-0, 304455/2017-2).
383 384
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Phosphate fertilization changes the characteristics of ‘Maçã’ banana starch, Int J Biol
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Macromol 112 (2018) 1138-1145.
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22 489
Figure captions
490 491
Fig. 1. Maximum, minimum, average temperatures (ºC) and rainfall (mm) in the months of
492
November/2012 to January/2016 in the experimental area.
493 494
Fig. 2. Ranges of bunches mass (A), total mass of fingers (B) and finger mass (c) of banana
495
genotypes in function of phosphate fertilizer rates. The polynomial trend lines show the
496
phosphate fertilizer rate effects on each banana genotype. Different letters next to markers
497
within each phosphate fertilizer rate indicate a significant difference between banana
498
genotypes according to Tukey’s test, at 5% of probability. MP indicates the maximum point
499
of polynomial regression.
500 501
Fig. 3. Ranges of dry matter (A), total starch (B), resistant starch (C) and phosphorus (D)
502
contents in fruits of banana genotypes in function of phosphate fertilizer rates. The
503
polynomial trend lines show the phosphate fertilizer rate effects on each banana genotype.
504
Different letters next to markers within each phosphate fertilizer rate indicate a significant
505
difference between banana genotypes according to Tukey’s test, at 5% of probability. MP
506
indicates the maximum point of polynomial regression.
507 508
Fig. 4. Scores plot (A) and loadings plot (B) of principal component analysis performed on
509
physicochemical characteristics of the fruits of different banana cultivars. Trait
510
abbreviation: FH18, FHIA 18 cultivar; GN, Grand Naine cultivar; NNC, Nanicão cultivar;
511
PA, Prata Anã cultivar; BchM, bunch mass; TMFg, total mass of fingers; FgM, finger mass;
512
DMt, dry matter; TStc, total starch; RStc, resistant starch; P, phosphorus.
23 514
S0141-8130(19)37156-9 https://doi.org/10.1016/j.ijbiomac.2019.10.217 BIOMAC 13718
To appear in:
International Journal of Biological Macromolecules
Received Date: Accepted Date:
4 September 2019 24 October 2019
Please cite this article as: M. Leonel, A. Carolina Batista Bolfarini, M. Jocimar Rodrigues da Silva, J. Mirellys Azevêdo Souza, S. Leonel, Banana fruits with high content of resistant starch: effect of genotypes and phosphorus fertilization, International Journal of Biological Macromolecules (2019), doi: https://doi.org/ 10.1016/j.ijbiomac.2019.10.217
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© 2019 Published by Elsevier B.V.
1 1
Banana fruits with high content of resistant starch: effect of genotypes and phosphorus
2
fertilization
3 4
Magali Leonela,*,[email protected], Ana Carolina Batista Bolfarinib, Marlon Jocimar Rodrigues
5
da Silvab, Jackson Mirellys Azevêdo Souzab, Sarita Leonelb
6 7
aCenter
8
Botucatu, São Paulo 18610-307, Brazil
9
bSchool
for Tropical Roots and Starches (CERAT), São Paulo State University (UNESP),
of Agriculture, São Paulo State University (FCA/UNESP),
10
Botucatu, São Paulo 18610-307, Brazil
11
*Corresponding
12
Highlights
13
Phosphorus plays an important role in starch accumulation in bananas;
14
Response to phosphate fertilizer is genotype dependent
15
Phosphate fertilization increases resistant starch content in green banana fruits
16
author.
2 17 18
Abstract
19
Banana fertilization practices aim to increase agricultural yield and the effects of these
20
practices on fruit components have not been well explored. This study aimed to evaluate the
21
effects of phosphate fertilizer levels on production parameters and accumulation of dry
22
matter, phosphorus, starch and resistant starch in green fruits. Four banana genotypes were
23
cultivated under the same cultural practices, with varying levels of phosphate fertilizers
24
during three production cycles. Bunches were harvested and evaluated for total fruit mass,
25
average finger mass and yield. Fresh green fruit pulps were analyzed for dry matter,
26
phosphorus, starch and resistant starch content. The results showed that the effects of
27
phosphate fertilizer were genotype dependent and that the increase of P2O5 rates applied in
28
banana fertilization promoted an increase in fruit production with higher levels of phosphorus,
29
starch and resistant starch.
30
Keywords: Musa spp; mineral nutrition; resistant starch
31 32
3 33
1. Introduction
34
Banana, as a climacteric fruit, has a very short period of post-harvest
35
commercialization, leading to considerable economic losses, which has been increasing the
36
use of green bananas as a source of functional ingredients [1, 2].
37
In most of the Brazilian regions there are favorable soil conditions for banana
38
production, but not always the chemical and physical properties of the soils are the most
39
adequate, reflecting in low quality and fruit productivity. Thus, fertility and soil physical
40
structure are fundamental factors [3].
41
Phosphorus (P) is an essential nutrient for plants and it is important in breathing,
42
energy production and transformation processes by the plant, as well as in cell division. This
43
nutrient is quickly mobilized in plants and, when there is a deficiency, is translocated from
44
older tissues to active meristematic regions [4].
45
The adequate supply of P is therefore essential from the early stages of plant growth
46
and the application of adequate P doses enables root development, besides increasing water
47
and nutrients uptake, which can contribute to fruiting [5,6].
48
Banana crop demands a high amount of fertilizers, not only because of the high
49
accumulation and the export of nutrients, but also because it is commonly grown on soils with
50
lower fertility. The phosphorus rates recommended in banana cultivation vary from 30 to 120
51
g of P2O5 per planting pit, depending on the available P content and soil texture. However,
52
banana absorbs most of the required P between three and nine months after planting, and
53
reduces to 80% in the reproductive phase [7].
54
In addition to the phosphorus functions already mentioned, this nutrient play important
55
roles in starch biosynthesis and in functional properties of the starches. Photosynthesis and
56
sucrose synthesis-related genes have been shown to change their expression following
57
phosphorus deficiency. Phosphate esters are covalently bound to starch granules, mainly in
4 58
amylopectin molecule, providing specific properties for starches and thus making them more
59
suitable for different industrial applications [5, 8].
60
Starch is the main component of unripe bananas, which undergoes important changes
61
during ripening. The average starch content drops from 70 to 80% in the pre-climacteric
62
period (prior to starch breakdown) to less than 1% at the end of the climacteric period [2].
63
Starch occurs naturally in the form of granules in plant cells and is mainly composed
64
of two polymers: amylopectin and amylose. Amylose is a linear polymer composed of
65
glucopyranose units linked through α-D- (1 → 4) glycosidic linkages, while amylopectin is a
66
highly branched polymer of high molecular weight [9].
67
Based on in vitro digestibility starch may be categorized into rapidly digestible starch
68
(RDS), slowly digestible starch (SDS), and resistant starch (RS). Resistant starch (RS) has
69
been defined as the starch fraction which escapes digestion in the small intestine of healthy
70
individuals [10].
71
Resistant starch occurs because of its inaccessibility to amylase. Depending on the
72
nature of inaccessibility, there are five classes: RS1 is found in grains and the granule may be
73
encased inside cells or in a strong protein matrix; RS2 consists of raw starch granules that
74
resist amylase digestion; RS3 is retrograded starch; RS4 is chemically modified starch, and
75
RS5 is amylose complexed with lipid [11, 12, 13].
76
The use of green banana pulp as a source of resistant starch involves the study of
77
several interfering factors, such as the particular characteristics of each banana genotype, as
78
well as the interference of the growing conditions on the productivity and the intrinsic
79
characteristics of the starch.
80
Considering the growing market for functional foods and the importance of banana in
81
the multiple aspects involved in food security, this study evaluated the effect of levels of
82
phosphate fertilizers applied to the cultivation of four banana genotypes on parameters of
5 83
productivity, dry matter accumulation, phosphorus, total starch and resistant starch in fruit in
84
order to provide useful information for the expansion of production and marketing of green
85
banana pulp.
86 87
2. Material and Methods
88
2.1. Banana genotypes
89
The banana genotype Prata Anã is from the genomic group AAB. It is a very vigorous
90
plant and does not require shoring. It presents size of 2.0 to 3.5 m, good tillering, vegetative
91
cycle of 280 days and average yield of 15 t/ha. It presents small bunches (14 kg), with 100
92
fruits/bunch and fruits of 13 cm with 110g of weight. It is tolerant to cold, susceptible to
93
Yellow and Black Sigatoka, and to bacterial wilt (Moko), moderately susceptible to fusarium
94
wilt (Panama disease), and moderately resistant to nematodes and banana root borer [14].
95
The cultivar Fhia 18, a hybrid of PrataAnã, is tetraploid (AAAB) and was developed
96
by the Honduran Agricultural Research Foundation [15]. (FHIA, 2018). The plant presents
97
medium size, a vegetative cycle of 353 days, good tillering and average yield of 20 t/ha. Its
98
fruits are straight, thick with medium size. This cultivar is resistant to black Sigatoka and
99
Panama disease, moderately resistant to yellow Sigatoka and susceptible to Moko and banana
100
root borer [14].
101
The genotypes 'Nanicão-IAC-2001' and 'Grand Naine' are of the genomic group (AAA)
102
and show similar characteristics regarding the physical fruits characteristics. These genotypes
103
are susceptible to yellow and black Sigatoka and resistant to Panama disease [14].
104 105
2.2. Banana growth
106
The cultivation of the four banana genotypes was conducted in Red Latosol at
107
Experimental Farm from School of Agriculture (FCA) that belongs to UNESP, located in the
6 108
city of São Manuel, state of São Paulo (22°44'28"S 48°34'37"W, at an altitude of 740m). The
109
climate is humid subtropical (Cfa), that is, temperate hot (mesothermic), with concentrated
110
rains from November to April (summer) and average annual rainfall of 1376.70 mm; the
111
city´s average temperature of the warmest month exceeds 22°C [16]. Figure 1 show the
112
temperature and rainfall data in the experimental area during the study.
113
Soil samples were collected at a depth 0-20cm to determine their chemical properties:
114
pH in CaCl2, 5.5; M.O., 12 g dm-3; P resin, 16 mg dm-3; H+Al, 15 mmolc dm-3; K, 1 mmolc
115
dm-3; Ca, 13 mmolc dm-3; Mg, 5 mmolc dm-3; SB, 19 mmolc dm-3; CTC, 34 mmolc dm-3;
116
V%, 57; S, 1 mg dm-3.
117
In November 2012, it was held the transplanting of micro propagated bananas plantlets
118
adopting a spacing of 4 m between rows and 2.5 m between plants, totaling 1000 plant ha-1. It
119
was opened planting holes in the dimensions of 60 cm in diameter by 60 cm in depth and
120
were applied 10 L of cattle manure and half of the recommended doses of phosphorus for
121
planting. The other half was applied 80 days after planting [17]. Also 0.5 kg of dolomitic
122
limestone was added to raise the base saturation index to 60% and the magnesium content to 9
123
mmolc dm-3 [18].
124
The experiment was conducted during three production cycles. The first production
125
cycle happened from November 2012 to April 2014; while the second from August 2014 to
126
February 2015; and the third from June 2015 to January 2016.
127
Fertilization of production with phosphorus (40 kg P2O5 - 100%) was standardized
128
according to soil chemical analyzes and expected yield of less than 20 t ha-1 [17]. Therefore,
129
P fertilization doses were: no application (control); 20 kg P2O5 ha-1 year-1; 30 kg P2O5 ha-1
130
year-1; 40 kg P2O5 ha-1 year-1; 50 kg P2O5 ha-1 year-1; and 60 kg P2O5 ha-1 year-1. Furthermore,
131
triple superphosphate (46% P2O5) was used as the source of phosphorus. The applications
132
realized in the form of semicircle (100 cm radius) in front of the youngest shoot. These
7 133
production fertilizers were carried out in January 2014, August 2014 and June 2015 for the
134
first, second and third cycles of production, respectively.
135
Supplemental fertilizations were 600 g of ammonium sulphate and 550 g of potassium
136
chloride per plant, divided in the course of the cycle, 20% at 35 days after planting (DAP),
137
50% at 80 DAP and 30% at 130 DAP [18]. The fertilizers were applied in circles of 100 cm in
138
diameter around the plant. At the time of thinning it was carried out the fertilization with 10 g
139
of boric acid in the open hole in the rhizome of the mother plant.
140
In the course of the experiment were performed: irrigation, weed control, thinning and
141
removal of dried leaves, fertilization, pest control and disease, elimination of banana heart,
142
removal of pistils and cutting pseudo stem after harvesting.
143 144
2.3. Production analysis
145
Fruits at maturity stage 1 (fully green peel) of five plants of each genotype in each
146
treatment were harvested. In order to standardize the maturation stage of the banana cultivars,
147
the ratio (ºBrix/titratable acidity) of the fruits was also determined.
148
The following variables were evaluated: mass of bunch (kg), total mass of fruits (kg),
149
average mass of fingers (g) and yield (t ha-1). The yield was calculated considering the weight
150
of the bunch in a stand of 1000 plants ha-1.
151 152
2.4. Fruits characteristics
153
The green fruits of each banana genotype, grown under the different conditions of
154
phosphate fertilization, were evaluated for phosphorus [19], dry matter [20], total starch and
155
resistant starch in the three cycles.
156
For the analysis of starch fractions in banana pulp samples of immature fresh fruits
157
(0.5 g) were disintegrated with liquid nitrogen and then treated with successive washes of
8 158
80% alcohol, 50% alcohol and water to remove sugar soluble and other soluble substrates
159
[21]. Determinations of resistant starch and total starch contents followed the methodology
160
described by Gõni et al. [22].
161 162
2.5. Experimental design and statistical analysis
163
The experimental design was completely randomized systematized in subdivided
164
plots, with the plots represented by the five rates of phosphate fertilization and the subplots by
165
the four banana cultivars, with 10 plants per experimental plot and 6 replicates in blocks,
166
totaling 60 plants of each genotype.
167
In order to study only the effect of phosphate fertilization and genotypes, the crop
168
cycles factor was eliminated. For this, we calculated the average of the results for each
169
production cycle, then being carried out the analysis of these averages.
170
The data were submitted to the test of homogeneity and normality, analysis of variance
171
(test F) and, when significant, the means of treatment of the P2O5 rates for each genotype were
172
expressed in regressions for the effects of rates of fertilizer for all variables. The criterion for
173
choosing the model was the significance of the F test up to 5% of probability and the highest
174
value of the determination coefficient (R2). Aiming to have a better visualization of the
175
physicochemical profile of the fruits of the four banana cultivars, regardless of the applied
176
phosphorus doses, the principal component analysis (PCA) was performed.
177 178
3. Results and discussion
179
There was a significant interaction between the banana cultivars and the phosphorus
180
doses for all variables analyzed (Figures 1 and 2). Thus, the effects of the two factors were
181
jointly analyzed.
182
9 183
3.1. Production analysis
184
The effect of phosphate fertilizer levels on the bunch mass, total fruit mass and fruit
185
mass was very similar in all banana cultivars studied. For all of them, the quadratic regression
186
model was better fitted, and it was verified that the highest values of the mentioned
187
production characteristics were observed when 35 kg of P2O5 ha-1 were applied. Fertilizer
188
doses lower and higher than this provided lower results (Figure 2).
189
The quadratic effect in response to increasing phosphorus fertilizer levels on this
190
parameter also were observed in other studies [23,24]. However, Silva and Rodrigues [7]
191
evaluating five rates of triple superphosphate (0, 50, 100, 200 and 300 g of P2O5) applied
192
during cultivation of ‘PrataAnã’ (AAB) in a clayey Red Latosol with low phosphorus
193
availability reported a linear increase in bunch mass in function of the P2O5 rates.
194
Despite the variations in the regression models, the results observed by these authors
195
corroborate with those obtained in this study, showing that the inadequate supply of nutrients
196
due to the use of low fertility soils has been one of the main causes of obtaining low yields in
197
banana plants. The perennial vegetative and reproductive growth of this plant requires the
198
adequate supply of nutrients throughout the crop cycle. The phosphorus absorbed by the plant
199
participates in several metabolic processes, including energy transfer, synthesis of nucleic
200
acids and starch, respiration, membrane synthesis and stability, activation and deactivation of
201
enzymes, redox reactions and carbohydrate metabolism [25,26], actions directly related to the
202
increase of productivity.
203
Significant differences in all productive characteristics were observed among the
204
cultivars studied. Regardless of the fertilizer rate, the highest values of bunch mass, total fruit
205
mass and fruit mass were obtained in the cultivar Grand Nine, followed by the cultivars
206
Nanicão and FHIA 18. On the other hand, the cultivar Prata Anã showed the lowest values for
207
production parameters. The differences observed between the banana genotypes for the
10 208
phosphate fertilizer could be due to their mechanisms of adaptation to the growing conditions,
209
as well as due to their genetic characteristics. It is known that Cavendish bananas (Grande
210
Naine and Nanicão), are generally more productive than Prata (Prata Anã and FHIA 18).
211
Plants growing at different P concentrations develop adaptive mechanisms including
212
changes in morphology and architecture of the root system as well as in physiological
213
characteristics of roots. A more developed and extensive root system allows greater
214
absorption of less mobile ions, such as phosphate [27, 28]. Thus, lower levels of phosphate
215
fertilization are required to increase the parameters of productivity.
216
Besides this, the increase in production parameters observed in this study may be
217
related to the important effects of phosphorus on photosynthesis. Photosynthesis and sucrose
218
synthesis-related genes have been shown to change their expression following P deficiency,
219
indicating their involvement in the observed increase in growth [5].
220
Another point is that in the adequate levels in the soil the phosphorus stimulates root
221
growth and development, allowing greater efficiency in water utilization and ionic absorption
222
of other elements. However, elevation of salinity and toxicity provided by high concentrations
223
of phosphate fertilizer reduce root growth and, consequently, affect nutrient uptake, which
224
may justify the decrease in production parameters with the use of fertilizer rates higher than
225
40 kg of P2O5 ha-1 year-1.
226
So, results obtained by adequate levels of phosphorus fertilizer in banana culture is so
227
important because as discussed by Zhu et al. [29] in their revision about phosphorus in
228
agricultural soils, there are a number of adverse environmental impacts associated with the
229
excessive use of inorganic phosphate fertilizers. These include increasing risk of P loss from
230
soils with elevated phosphorus concentrations, which may lead to eutrophication in water
231
bodies and the depletion of finite resources of high grade phosphate rocks.
11 232
In view of the above, it is evident the need to study the real need for phosphate
233
fertilization by the different genotypes. It was observed a better productive performance of all
234
genotypes at doses below the recommended level (40 kg P2O5 kg-1 year-1), especially in the
235
cultivar Grand Naine, in which the highest bunch weight was obtained with 27.2 kg P2O5 kg-1
236
year-1, i.e. only 68% of the recommended dose.
237 238
3.2. Fruits characteristics
239
The analysis of the levels of dry matter, phosphorus, total starch and resistant starch in
240
the fruits of the four banana genotypes showed differences among the cultivars within each
241
level of phosphate fertilization and the effect of the P2O5 rates used on these components in
242
the unripe fruits (Figure 3).
243
When we analyzed the effects of phosphorus doses on the physicochemical
244
characteristics of banana fruits, it was possible to observe that the quadratic regression models
245
were the best fit for all cultivars studied. The data analysis showed that the increase of
246
phosphate fertilizer rates applied in banana cultivation promoted an increasing in dry matter,
247
phosphorus, total and resistant starch contents. However, in the higher concentrations there
248
was a decreasing in the contents of these components. As all these parameters are directly
249
related to the metabolism of the plants and the phosphorus is intrinsically linked to it, it is
250
important to emphasize that in the extreme conditions of low or high availability of
251
phosphorus several control mechanisms are activated in the plant, reflecting in the
252
accumulation of dry matter, in the starch content and its structure characteristics, as well as, in
253
the accumulation of this mineral in the fruit.
254
When the supply of phosphorus is limited, plants grow more roots, increase the rate of
255
uptake by roots from the soil, retranslocate Pi from older leaves, and deplete the vacuolar
256
stores of Pi. In addition, mycorrhizal fungi may more extensively colonize the roots.
12 257
Conversely, when plants have an adequate supply of Pi and are absorbing it at rates that
258
exceed demand, a number of processes act to prevent the accumulation of toxic Pi
259
concentrations [30].
260
In general, the highest contents of dry matter in the fruits were obtained when the
261
bananas were fertilized with 40 kg of P2O5 ha-1. Using this dose, we observed that the fruits of
262
the cultivars Nanicão and Prata Anã averaged 42.16% of dry matter, differing significantly
263
from fruits of the cultivars Grand Naine and FHIA 18, which, with the same dose of the
264
fertilizer, presented about 38% dry matter (Fig. 2A).
265
The effect of phosphorus fertilizer rates on fingers dry matter showed that the
266
adequate availability of this nutrient in the soil influenced the production and allocation of
267
assimilates in the fruits. On the other hand, the lower phosphorus availability in soil with the
268
use of the half levels of fertilizer recommendation (20 kg of P2O5 ha-1 year-1)may have
269
decreased the dry matter content by reducing the radiation intercepted during the crop cycle
270
(IR), as cited by Jenkins and Ali [31] in their study with potatoes.
271
The levels of phosphorus in banana fruits ranged from 31.62 to 42.45 mg 100 g-1, with
272
differences between cultivars and fertilizer rates (Fig. 2B). The highest accumulations of
273
phosphorus in the fruits occurred with fertilization doses of 40 Kg ha-1 for the Grand Nine,
274
around 43 Kg ha-1 for the Nanicão and FHIA 18, and 47 Kg ha-1 for Prata Anã. Within the
275
evaluated dose of 40 kg ha-1, there was no significant difference in phosphorus content in the
276
Nanicão and Grand Naine cultivars, with a mean of 41.7 mg 100 g-1. However, these cultivars
277
differed significantly from the cultivars Prata Anã and FHIA 18, which showed on average
278
35.2 mg 100 g-1 of phosphorus in fruits.
279
For all cultivars, the increase of the fertilizer rates led to an increasing of the
280
phosphorus content, however, in the higher levels it was not observed an increasing in the
13 281
phosphorus levels in fruits pulps. Applying higher rates of phosphate fertilizer may result in
282
considerable immobilization, leaving part of the phosphate inaccessible to the plants [32].
283
The differences in phosphorus content in fruits, as well as the response to fertilization
284
among cultivars may be related to the root system. In a more developed and deeper root
285
system there is an increase in the contact area between the roots of the banana and the soil,
286
which favors the absorption of the P by the plant.
287
This increase in the phosphorus content in the fruits pulps is very important since in
288
the composition of a diet the phosphorus comes mainly from the animal protein sources.
289
Phosphorus is an essential nutrient in human health and has a variety of physiological roles.
290
The increase of its content in the banana fruits leads to nutritional benefits, since phosphorus
291
has the function of buffering acidic or alkaline systems, assisting in the maintenance of pH, in
292
the temporary storage of energy derived from macronutrient metabolism, in the form of ATP,
293
being responsible for the activation through the phosphorylation of several enzymatic
294
cascades [33].
295
The increase in the doses of phosphate fertilization led to a limited increase in the
296
content of starch in the banana fruits. For the cultivars Grand Naine and Prata Anã, the
297
highest content of starch in the fruits was obtained when the fertilization was carried out with
298
37 kg of P2O5 ha-1. On the other hand, fruits of the cultivars Nanicão and FHIA 18 showed the
299
maximum values for starch content with higher doses of the fertilizer, that is, with 44 kg of
300
P2O5 ha-1, with reduction in the starch content in fruits in higher doses of fertilizer.
301
These results evidenced the important participation of phosphorus in the biosynthesis
302
of starch in plants. AGPase is a key regulatory enzyme that catalyzes the rate limiting step of
303
starch biosynthesis and it is extremely sensitive to allosteric regulation, with glycerate-3-
304
phosphate (3-PGA) acting as an activator and Pi as an inhibitor [34].
14 305
Zhao et al. [26] in their study with the effects of nitrogen and phosphorus on
306
transcriptional regulation of genes encoding key enzymes of starch metabolism in duckweed
307
described that the activities of AGPase and soluble starch synthesis enzyme (SSS EC
308
2.4.1.21) were positively correlated with changes in starch content.
309
The global starch market is poised to register in the period of 2015 to 2023 a
310
Compound Annual Growth Rate (CAGR) of 5.85%. The market is competitive and driven by
311
an increase in the trend of “health and wellness” and growing consumer demand for all
312
natural ingredients [35]. Thus, the possibility of increasing the resistant starch content by
313
agricultural management in banana cultivation may allow Brazil a commercial differential.
314
The levels of resistant starch in the fresh pulps of unripe bananas ranged from 17.0 to
315
29.8 g 100 g-1. Studies related that banana starches have B-type crystalline pattern and
316
starches with this type of crystallinity tend to be more resistant to pancreatic amylases. This
317
increased strength is due to branches and associations of amylopectin and amylose [14, 36,
318
37].
319
Phosphate fertilization rates had effect on this fraction of the starch for all genotypes.
320
The maximum values for resistant starch contents were obtained with fertilizations at the
321
doses of 40 kg of P2O5 ha-1 for Prata Anã, around 42 kg of P2O5 ha-1 for the cultivars Nanicão
322
and FHIA 18 and 54 kg of P2O5 ha-1 to Grand Naine. The changes in phosphorus levels in
323
unripe fruits pulp can have promoted changed in phosphorus linked in starch structure with
324
can have led to a disruption of the hydrogen bonds of the amylopectin double helix reducing
325
the crystalline region of the starches and interfering in the resistant starch content in the
326
banana fruits.
327
Interference of phosphorus in starch digestibility has been evaluated and some studies
328
related that as amylolytic enzymes are incapable of bypassing the phosphorylated glucosyl
15 329
residue, phosphoryl-oligosaccharides are released from the digestion of starch with amylase
330
[38, 39].
331
The effect of phosphate fertilizer on the characteristics of banana starch was evaluated
332
by Mesquita et al. [40] and these authors observed effect of phosphate fertilization on the
333
cristallinity of the banana starch (genotype Maçã). The increase in phosphorus fertilizer led to
334
a decreasing of cristallinity of the starch and resistant starch content.
335 336
3.3. Principal components analysis
337
The effect of the different doses of phosphorus was clearly evidenced by Figures 2 and
338
3, which also showed the effects of the banana cultivars in each dose of the evaluated
339
fertilizer. However, in order to better understand the physicochemical profile of the fruits of
340
the four banana cultivars, independent of the doses of phosphate fertilizer, the principal
341
components analysis was applied, and the results showed that only the first two components
342
(PC1 and PC2) explained 93.89% of total variability (Figure 4).
343
Analyzing PC1, which explained 65.26% of the data, we observed that the fruits of the
344
cultivar Prata Anã were clearly separated from the other cultivars, mainly from Grand Naine,
345
which was positioned on the opposite side in PC1 (Figure 4A). According to PC1 loadings,
346
except for the phosphorus content, all other variables strongly contributed to this separation
347
(Figure 4B). PC1 scores and loadings indicate that the cultivar Prata Anã showed better
348
quality fruits, with a higher content of dry matter and higher concentrations of starch, total
349
and resistant. However, this cultivar was less productive, presenting lower values of bunch
350
mass and fruit, individual and total. An opposite result was observed in the cultivar Grand
351
Naine, which presented high productive indexes, however with fruits with low starch
352
contents. In addition, the analysis showed that the cultivar Nanicão was similar to Grand
16 353
Naine, but this cultivar, together with FHIA 18, showed intermediate values because they
354
were closer to the central axis of PC1.
355
The PC2, which explained 28.63% of the variability of the experiment, was evidenced
356
mainly by the phosphorus content in the fruits, as can be visualized in PC2 loadings, showing
357
the importance of this element in the distinction of the banana cultivars.
358
The analysis of PC2 scores and loadings especially suggests a separation of the
359
cultivars Nanicão and FHIA 18, since the highest levels of phosphorus were obtained in
360
bananas of the cultivar Nanicão and smaller in the fruits of FHIA 18. Thus, it can also be
361
observed that this element was observed in intermediate concentrations in the fruits of the
362
cultivars Prata Anã and Grand Naine.
363 364
4. Conclusion
365
Results obtained in this study showed that, through the management of phosphate
366
fertilization, it is possible to increase the agricultural production of green banana fruits with
367
higher levels of phosphorus and resistant starch, allowing a substantial increase in the
368
commercial value of banana pulp by the growing market in the food industry. Therefore, the
369
management of the fertilization for banana crop can be directed to obtain not only high yield
370
but also in view of higher levels of functional ingredients. PrataAnã banana stands out as a
371
source of dry matter and high resistant starch content. On the other hand, Grande Naine is the
372
most productive, but with low levels of starch and dry mass.
373 374 375 376 377 378 379
Conflict of interest The authors declare no conflict of interest. Author contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript.
17 380
Acknowledgement
381
The authors thank CNPQ for financially supporting this work (Processes 303373/2014-8,
382
302827/2017-0, 304455/2017-2).
383 384
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Figure captions
490 491
Fig. 1. Maximum, minimum, average temperatures (ºC) and rainfall (mm) in the months of
492
November/2012 to January/2016 in the experimental area.
493 494
Fig. 2. Ranges of bunches mass (A), total mass of fingers (B) and finger mass (c) of banana
495
genotypes in function of phosphate fertilizer rates. The polynomial trend lines show the
496
phosphate fertilizer rate effects on each banana genotype. Different letters next to markers
497
within each phosphate fertilizer rate indicate a significant difference between banana
498
genotypes according to Tukey’s test, at 5% of probability. MP indicates the maximum point
499
of polynomial regression.
500 501
Fig. 3. Ranges of dry matter (A), total starch (B), resistant starch (C) and phosphorus (D)
502
contents in fruits of banana genotypes in function of phosphate fertilizer rates. The
503
polynomial trend lines show the phosphate fertilizer rate effects on each banana genotype.
504
Different letters next to markers within each phosphate fertilizer rate indicate a significant
505
difference between banana genotypes according to Tukey’s test, at 5% of probability. MP
506
indicates the maximum point of polynomial regression.
507 508
Fig. 4. Scores plot (A) and loadings plot (B) of principal component analysis performed on
509
physicochemical characteristics of the fruits of different banana cultivars. Trait
510
abbreviation: FH18, FHIA 18 cultivar; GN, Grand Naine cultivar; NNC, Nanicão cultivar;
511
PA, Prata Anã cultivar; BchM, bunch mass; TMFg, total mass of fingers; FgM, finger mass;
512
DMt, dry matter; TStc, total starch; RStc, resistant starch; P, phosphorus.
23 514