ts-1 physiological and biochemical changes during banana ripening and finger drop

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Postharvest Biology and Technology 39 (2006) 211216

Physiological and biochemical changes during bananaripening and finger drop

Wachiraya Imsabai a, Saichol Ketsa b,, Wouter G. van Doorn c

a Department of Horticulture, Faculty of Agriculture at Kamphaeng Saen, Kasetsart University, Nakhon Pathom 73140, Thailandb Department of Horticulture, Faculty of Agriculture, Kasetsart University, Chatuchak, Bangkok 10900, Thailand

c Wageningen University and Research Centre, P.O. Box 17, 6700 AA Wageningen, The Netherlands

Received 21 February 2005; accepted 2 October 2005

Abstract

Fruit drop of banana is due to breaking at the junction of the pedicel and pulp, and we found no true abscission zone. The breakage seems

therefore due to weakening of the peel. We investigated pectin hydrolysis and some properties at the rupture zone, using Hom Thong (Musa

acuminata, AAA Group) and Namwa (Musa x paradisiaca, ABB Group) fruit, which show massive drop and no drop, respectively. During

the period of finger drop, the water content of the peel in Namwa was similar to that of Hom Thong and thus water content does not account

for the high breakage in Hom Thong. The peel thickness at the rupture area in the two cultivars was not significantly different. During the

period of finger drop, the level of water-soluble pectin in the peel at the rupture area of Hom Thong was higher than that of Namwa,

indicating pectin degradation. CDTA soluble pectin and insoluble pectin was lower in Hom Thong, also indicating more pectin breakdown

in this cultivar. Polygalacturonase activity in the peel at the rupture area of Hom Thong bananas rapidly increased, but not clearly more than

in Namwa bananas. Pectinesterase activity in the peel at the rupture area of Hom Thong was much lower than that of Namwa bananas,

and thus does not account for the breakage. Pectate lyase in the peel of Hom Thong was considerably higher than in that of Namwa. The

present data indicate that the much higher pectate lyase activity in Hom Thong might be responsible, at least partially, for the finger drop.

2005 Elsevier B.V. All rights reserved.

Keywords: Banana; Finger drop; Polygalacturonase; Pectinesterase; Pectate lyase

1. Introduction

Finger drop has been defined by Baldry et al. (1981) as

the physiological softening and weakening which causes the

individual fruit in a hand to separate from thecrown. Hands of

banana with fingers missing cannot be sold to consumers and

individual fruit that have dropped have no pedicel and can-

not be marketed. Finger drop has been reported in a diploidcultivar (Prayurawong, 1999), in the triploid Cavendish AAA

Group (Semple and Thompson, 1988), and in tetraploid culti-

vars (Marriott, 1980). Susceptibility varies widely. For exam-

ple,amongthe triploid cultivars Valery is considerably more

prone to finger drop than Gros Michel (New and Marriott,

1983).

Corresponding author. Tel.: +66 2579 0308; fax: +66 2579 1951.

E-mail address: [email protected] (S. Ketsa).

Banana finger drop is stimulated by high relative humid-

ity and high ripening temperature (Semple and Thompson,

1988), and by ethylene (Paull, 1996). In addition, more

mature hands are apparently more sensitive to finger drop

(Paull, 1996).

Finger drop seems to be due to localized weakening of

peel at the pedicel (New and Marriott, 1983; Semple and

Thompson, 1988). Softening of banana peel, as that in othertissues, has been suggested to be due to depolymerization

of pectic substances in the primary cell wall and the mid-

dle lamella (Seymour, 1993). Banana peel softening during

ripening might therefore involve a number of cell wall hydro-

lases.

Banana pulp ripening has been associated the activities of

both endo-polygalacturonase (endo-PG) and exo-PG (Wade

et al., 1992), with that of pectinesterase (PE, also called

pectinmethylesterase; Wade et al., 1992), and pectate lyase

0925-5214/$ see front matter 2005 Elsevier B.V. All rights reserved.

doi:10.1016/j.postharvbio.2005.10.001


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212 W. Imsabai et al. / Postharvest Biology and Technology 39 (2006) 211216

(PL; Marin-Rodriguez et al., 2003; Payasi and Sanwal, 2003;

Lohani et al., 2004). It is not known if the same hydrolytic

enzymes are active in banana peel. The activity of PL in the

peel was below the detection limit (Marin-Rodriguez et al.,

2003), although a peel PL gene was reportedly expressed

(Dominguez-Puigjaner et al., 1997; Marin-Rodriguez et al.,

2003). Several other hydrolytic enzymes genes might beinvolved in pectin degradation.

It is not known if the activity of any of these hydrolytic

enzymes is associated with bananafinger drop. We here report

on two cultivars, one with high and one with no finger drop.

We determined the weight of the fruit, the firmness of the

fruit, the diameter of the stalk at the abscission zone, the

thickness and the water content of the peel at the abscission

zone, a number of pectin fractions and the activities of PG,

PE and PL.

2. Materials and methods

2.1. Plant material

Banana fruit of the cultivars Hom Thong (Musa acumi-

nata, AAA Group) and Namwa (Musa x paradisiaca, ABB

Group) were harvested at commercial maturity. The fruit

wereplaced in corrugatedcardboardboxes and transported by

refrigerated truck (25 C) to the laboratory within 3 h of har-

vest. In the laboratory, hands were selected for size and color

and cleaned in a solution of 100l l1 chlorine (Clorox). The

hands were then dipped for 23min in 500 mg l1 ethephon

for uniform ripening and then dried at ambient temperature

(2930 C).Ripeningoccurredat 25 Cand8590%RH.The

hands were monitored daily for finger drop, pedicel ruptureforce, resistance to finger drop and enzyme activity. The peel

at the middle of the fruit and at the pedicel in the rupture

area was sampled at intervals. Peel from five hands of each

treatment was pooled and frozen at 80 C until further use.

2.2. Finger drop in hands of banana

The method was modified from Semple and Thompson

(1988). A hand of banana was held at 15 cm above a table

for 10 s, and the number of dislodged fingers was recorded,

and expressed as a percentage of total number of fingers on

the hand.

2.3. Pedicel rupture force

Pedicel rupture force was measured by pressing down a

wedge probe at the pedicel until it separated from the fruit.

The required force was expressed in Newtons (N). Twenty

fruit were measured in each treatment, at each time point.

2.4. Resistance to finger drop

Banana fruit, attached to a hand, was inserted in a hole and

held by a bigclip, connectedto a spring weight. As thepedicel

of banana was pulled, the piston of the spring weight and a

marker moved together. The marker on the spring weight

stopped when the pedicel broke. The force at the moment of

rupture was indicated on the marker. The resistance to finger

drop was expressed in kilograms (kg).

2.5. Peel color and fruit firmness

Peel color was measured in the middle of the fruit, using

a Chroma-meter CR-300 series (Minolta, Japan) with 10 mm

viewing aperture.The instrumentwas calibratedusing a white

reference tile and a values were recorded.

Firmness was measured on one side, in the middle of a

fruit. The peel was left on the fruit. The penetrometer (Chatil-

lon and Sons, Kew Gardens, NY) was equipped with a 5 mm

probe penetrating 5 mm into the fruit.

2.6. Peel water content at the rupture area, peel

thickness, diameter of the pedicel, and fruit fresh weight

Peel at the rupture area (1 cm2) of banana fruit was col-

lected, weighed and oven-dried at 60 C for at least 5 days.

It was weighed daily until the weight did not further change.

The percentage water content was calculated. The diameter

of the pedicel and the thickness of the peel at the rupture

area were measured on day 3. Fingers were separated at the

rupture zone and weighed to determine fresh weight (FW).

2.7. Pectin fractions and enzyme assays

The extraction method for the pectin fractions was mod-

ified from Martin-Cabrejas et al. (1994). Briefly, 10 g ofpeel tissue was extracted for alcohol insoluble solids (AIS),

then 30 mg of AIS were used for pectin extraction. The AIS

was placed in water and shaken at 150 rpm for 16 h and

centrifuged at 15,000 rpm, 4 C for 30 min. After that the

pellets were extracted with water again, shaken at 150 rpm

for 6 h and centrifuged at 15,000 rpm, 4 C for 30 min. The

water-insoluble pellet was suspended in 0.05 M CDTA (in

1 M amidazole, pH 7), twice extracted (16 and 6 h) and cen-

trifuged as described above. The supernatant (CDTA-soluble

pectin) from the two extractions were pooled. The remaining

cell walls were twice extracted with 0.05 M Na2CO3 (16 and

6 h). Finally, the suspension was centrifuged as above. The

pectin in each fraction was assayed as uronic acid units, as

described by Blumenkrantz and Asboe-Hansen (1973).

Extraction and assay methods for PG were as described

by Yoshida et al. (1984) and those for PE as by Hagerman

and Austin (1986). Extraction for PL used the method of

Payasi and Sanwal (2003) and PL activity was determined

by the method ofCollmer et al. (1988), with slight modifica-

tion. Briefly the assay was done in a mixture of 0.5 ml 0.36%

(w/v) polygalacturonic acid in 0.05 M TrisHCl buffer, pH

8.5, 0.3ml 4 mM CaCl2, 0.6 ml enzyme and 0.6 ml water.

The reaction mixture was incubated at 37 C for 3 h. The PL

activity was determined by following absorbance at 232 nm.


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W. Imsabai et al. / Postharvest Biology and Technology 39 (2006) 211216 213

In a control tube, the substrate was added after incubation

and absorbance was read immediately. Protein content in the

enzyme extracts was estimated using the Bradford (1976)

method. Specific activity of the enzyme was expressed as

units per mg protein.

2.8. Statistical analysis

Using a SAS package, the data were treated by analysis

of variance, calculating the least significant difference (LSD)

between means, determined at the 5% level. Data were the

average of three or four replications standard error (S.E.).

All experiments were repeated once or twice at later dates.

3. Results

3.1. Peel color and fruit firmness

In order to characterize the ripening stage of the fruit,

three parameters were determined at intervals. The a value

at the middle of the fruit increased similarly in Namwa

and Hom Thong (Fig. 1A). The increase in a value was

associated with increased visible peel yellowing. Fig. 1B

shows that the firmness of the peel in the middle of the fruit

decreased during the experimental period. Initially, the peel

firmness of Hom Thong was significantly higher than that

of Namwa, whereas on day 3 it was significantly lower than

that of Namwa (Fig. 1B). In addition, no premature senes-

cent spotting was observed during the experimental period,

despite the high RH (results not shown).

Fig. 1. a Value (A), andfruit firmness (B)of Hom Thong () andNamwa

() bananas held at high RH.Timeis days after thepeel hadbecome mostly

yellow. Data are meansS.E. of four replications.

Fig. 2. Finger drop (A), pedicel rupture force (B) and resistance to finger

drop (C) of Hom Thong () and Namwa () bananas held at high RH.

Time isdays after thepeelhad becomemostly yellow. Data aremeansS.E.

of four replications.

3.2. Finger drop, rupture force, resistance to finger

drop, and anatomy

Fingerdrop of Hom Thong bananas started on day2 after

peel yellowing (day 2 after yellowing is time 0 in Figs. 16).

Finger drop increased to 100% within 34 days. During the

six days after the peel had become yellow, Namwa bananasdid not showfinger drop (Fig. 2A) and also did not show drop

thereafter (results not shown).

During the 4 days of the experiment, Namwa bananas

had a higher pedicel rupture force than Hom Thong fruit.

The pedicel rupture force of both banana cultivars decreased

in parallel (Fig. 2B). On the first day of the experiment,

the resistance to finger drop of Namwa and Hom Thong

bananas was both more than 5 kg (Fig. 2C). In Hom Thong

the resistance decreased more than in Namwa (Fig. 2C).

From day 2 onwards the difference was statistically signifi-

cant at the 5% level.

A detailed histochemical study of the peel and the interiorof the pedicel at the rupture area did not reveal an abscission

zone (data not shown).

3.3. Peel thickness, water content, diameter of the

rupture area and weight of the fruit

The water content at the rupture area was initially the

same in both cultivars (Fig. 3). In Hom Thong, it did not

much change during the experimental period. In Namwa, it

decreased after day 2. The Namwa pedicel lost about 5%

water (Fig. 3).


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Fig. 3. Change of water content at the rupture area of the peel of Hom

Thong ()andNamwa()bananasheldathighRH.Timeisdaysafterthe

peel had become mostly yellow. Data are meansS.E. of four replications.

The average diameter of the pedicel of Hom Thong, atthe rupture area, was slightly higher than that of Namwa

(Table 1). The thickness of the peel at the rupture area of

the two cultivars studied was not different (Table 1). The

individual fruit weight of Hom Thong bananas was twice

as high as that of Namwa fruit (Table 1).

3.4. Pectin fractions

Total pectin in various fractions was expressed as uronic

acid levels. Water-soluble pectin in the peel taken from the

rupture area was initially higher in Hom Thong than in

Namwa. From day 2 onward no difference was found(Fig. 4A). The CDTA-soluble pectin level was higher in

Namwa than in Hom Thong, throughout the experimen-

tal period (Fig. 4B). The fraction of Na2CO3-soluble pectin

(water-insoluble pectin) in the peel at the rupture area was

also higher in Namwa than in Hom Thong.

3.5. PG, PE and PL activities

Total PG activity increased in both cultivars, both in the

peel at the rupture area and that taken from the middle of the

fruit. No clear differences were observed between the two

cultivars (Fig. 5A). On day 0 and day 2 the PE activity in the

peel of Namwa was much higher than in Hom Thong. The

activity in Namwa decreased. By day4 of theexperiment no

differences were found between the two cultivars (Fig. 5B).

Fig. 6 shows the PL activity in the peel on day 0 and day

Table 1

Peel diameter and thickness and fresh weight of the fingers of Hom Thong

and Namwa bananas

Cultivars Pedicel diameter (cm)* Thickness (mm)* Fruit FW (g)*

Hom Thong 1.10 0.01 26.0 0.2 164.6 2.9

Namwa 0.87 0.02 25.2 0.1 83.1 0.7

* Data are the means of ten fruit with SE.

Fig. 4. Water-soluble fraction (A), CDTA-solublefraction (B) and Na2CO3-

soluble fraction (C) in the peel at the pedicel of Hom Thong () and

Namwa () bananas held at high RH. Time is days after the peel had

become mostly yellow. Data are means S.E. of three replications.

3. The two first bars compare the peel at the middle of the

fruit in Namwa (first bar) and Hom Thong (second bar).

The activity is much higher in Hom Thong. This difference

persisted on day 3. A similar difference existed between the

peel in the rupture area both on day 0 and day 3 (Fig. 6).

Fig. 5. PG (A) and PE (B) activities in peel at the middle (, ) and at the

pedicel (,) of Hom Thong (, ) and Namwa (,) bananas held

at high RH. Time is days after the peel had become mostly yellow. Data are

meansS.E. of three replications.


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W. Imsabai et al. / Postharvest Biology and Technology 39 (2006) 211216 215

Fig. 6. PL activityin peel at themiddle of andat thepedicelof Hom Thong

(H) and Namwa (N) bananas held at high RH.

4. Discussion

The results show a large difference in finger drop between

the two triploid cultivars studied, with no drop in Namwa

and 100% drop in Hom Thong. This was not due to a dif-

ference in maturity. The two cultivars had the same maturity

and ripened normally. The changes in ripening parameters

were very similar.

We did not observe an abscission zone at the area of

rupture in Hom Thong. This indicates that the rupture

occurs due to physical breakage. As the rupture occurs at

the junction of the pedicel and the fruit flesh, the internal

forces that hold the tissue together at this point might berelatively weak. The junction occurs at the sudden transition

between a rather lignified tissue (the pedicel) and a rather

un-lignified one (the fruit pulp). The fruit seems to be held

together mainly by the peel. If the peel is too weak to hold

the weight of the fruit it will break.

Data on pedicel rupture force and resistance to finger drop

(Fig. 2B and C) indicate that the cultivar which is suscepti-

ble to finger drop has a weaker peel at the area where the

break occurs. This peel weakness could be due to a number

of factors, such as the circumference of the area where the

rupture occurs, the thickness of the peel, finger weight, the

water content of the peel, and pectin degradation.

Between the two cultivars studied, the pedicel diameter

and peel thickness did not considerably differ (Table 1). The

fruit of several other cultivars have a really thin peel, thinner

than the one measured in both cultivars studied here. Still, the

fingers in these cultivars generally do not drop. Similarly, the

fruit of other cultivars have a thick peel, but in these cultivars

there is finger rupture (unpublished data). These observations

support the contention that peel thickness is not of major

importance for the tendency to rupture.

Fruit weight was twice as high in the cultivar with greater

finger drop (Table 1). This factor, therefore, may be impor-

tant for the tendency to show finger drop in Hom Thong.

However, under some growing conditions the fruit of the two

cultivars presently studied is sometimes exactly the same,

but still the fingers of Hom Thong drop whereas those of

Namwa do not (unpublished data). Moreover, when com-

paring finger drop in other cultivars, fruit weight does not

seem very important. For example, Leb Mu Nang (Musa

AA Group) fruit weighs about half that of Namwa but LebMu Nang is prone to finger drop (unpublished data) and

Namwa is not. Finally, resistance to finger drop among sev-

eral banana cultivars did not correlate well with finger length

or finger diameter (Nunes de Jesus et al., 2002), which also

argues against a relation with fruit weight. Fruit weight, there-

fore, does not seem a major factor that determines finger drop.

If a cultivar is prone to finger drop, the drop is promoted

by high RH. Semple and Thompson (1988) suggested that

a low water content in the peel at the rupture area results

in less finger drop. Water content does not explain the dif-

ference between the cultivars presently studied. During the

main period of finger drop in Hom Thong (day 02; Fig. 2),

the water content in the peel at the rupture area was the sameas that in Namwa (Fig. 3). The water content in the peel is

therefore not a major contributing factor.

Softening of the peel might be a cause of finger drop. Soft-

ening might be due to cell wall hydrolases. Indeed, day 0 and

1 of the experiment more water-soluble pectin was found

in the peel at the rupture zone of Hom Thong, compared

with Namwa. This might relate to the finger drop in Hom

Thong, which occurred mainly during day 01 and 12. In

Namwa, the CDTA-soluble pectin fraction in the peel at

the rupture zone was considerably higher than that of Hom

Thong. This fraction represents calcium-bound pectin, cal-

cium forming cross-links between adjacent pectin molecules.Insoluble pectin (the sodium bicarbonate fraction) was also

higher in Namwa than in Hom Thong. Both fractions

might help keep the cells of Namwa more together, thus

more resistant to finger drop.

Among the hydrolases studied, finger drop could appar-

ently not be explained by the activities of total PG or PE

(Fig. 5). There was a correlation with PE, but this was oppo-

site to that expected (Fig. 5B). Finger drop was correlated in

an expected way with PL activity (Fig. 6).

It is concluded that fruit drop in Hom Thong banana is

not due to true abscission but to breaking of the peel at the

junction of the fruit and the hand. No evidence was found for

a decisive role in this rupture of any of the following factors:

fruit weight and thickness, and water content of the peel at

the rupture area. A positive correlation was found with the

activity of PL in the peel at the rupture area. The relative high

PL activity might therefore contribute to finger drop in Hom

Thong fruit.

Acknowledgement

The research was financially supported by the Thailand

Research Fund (TRF).


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