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Compensative effects of chemical regulation on physiological damage caused by water deficiency during filling stage of wheat

Liusheng Duan, Zhaohu Li, Caihong Guan, Zhixi Zhai and Zhongpei He

China Agriculture University, #2 Yuanmingyuan Xilu, Beijing 100094, P. R. China, Email


The experiment was conducted with winter wheat (Triticum aestivum L.) cv. Jingdong 6 in PVC pots and under rainproof conditions in Beijing from 2000 to 2002. It was shown that the distribution of 3H2O in roots and flag leaf, characteristics of vascular bundle in primary roots and the internode below the spike, roots validity, transpiration rate and stomatal impedance of flag leaf were changed by water stress after flowering. The treatment of foliage spraying at early jointing stage using 450 g/hm2 HK, a plant growth regulator containing 3.3% Paclobutrazol and 16.7% Mepiquat Chloride as active components, were proved to release and compensate the harmful effects of water stress. Both the area of vascular bundle in primary roots and internode below ear were increased by HK, while the roots validity and the ability of water absorbing and transporting were promoted. In flag leaf, stomatal impedance was changed to maintain the transpiration rate and the water use efficiency of single wheat plant was higher. The rate and ratio of 14C-assimilates exporting from flag leaf were increased by the chemical regulation.

Media summary

A new method of applying plant growth regulator to improve water using of wheat plants, to compensate and decrease the harm of water stress.

Key Words

Wheat, Water stress, Plant growth regulator, Compensative effects


The filling stage after flowering is of vital importance for formation and development of wheat grains, so water stress during this stage will decrease the yield and grain quality (Ahmadi and Baker 2001; Shi et al. 1999). Even though the 3H2O tracing method has been applied to study and monitor the water movement effectively in other plants and already used to study the water absorbance of wheat roots and to identify the drought resistance of wheat species (Yang K 1998), the dynamic transport within the wheat plant is not clear yet. Under water deficiency, wheat displays a range of adaptive mechanisms such as reduced stomatal conductance, reduced growth, changes in leaf angle or leaf abscission (Tang 1983Bob et al. 2000). Signal substances like plant hormones have been shown to be involved in these responses to water stress, and the application of plant growth regulators which can change the levels and functions of plant hormones have the potential to regulate plant growth, development and related physiological processes including water metabolism, and to induce or enhance the resistance of plant to water deficiency (Luo 1992; Lou 2000). In our experiments, we studied the compensative and improving effects of applying plant growth regulators, HK as an example, under water stress in filling stage, and explored its potential to be used as a novel method to increase water use efficiency, crop yield and quality.


Material raising and treatment

The experiment was conducted at China Agriculture University in 2000-2002, using Triticum aestivum L. cv. Jingdong 8 as material. The plants were grown in split and enlaced PVP pipes (1m height and 25 cm diameter). The soil was from a wheat field and there was one plant per pipe. 450 g/ha HK, a registered plant growth regulator in China containing 3.3% Paclobutrazol and 16.7% Mepiquat Chloride as active components, was first diluted by 450kg/ha water and then foliage sprayed at early jointing stage. At early flowering stage watering was limited to create 85%(water abundance) and 60% (water deficiency) of maximum water withholding potential of soil. Split-split design was adopted and 4 duplicates for each treatment and 50 plants for each duplicate. Treatments were abundant water (WC), abundant water with HK (WR), water deficiency (SC) and water deficiency with HK (SR).

3H2O tracing experiment

The plants were washed out on May 20th (middle filling stage) and put into prepared culture liquid with full nutrients described by Tang (1999). The media liquid with same water potential with respective irrigation condition was prepared using the above culture liquid and PEG-6000 according to Michel (1973). 3H2O was diluted into different media liquid as 1.5 μCi/mL and then were used to soak the wheat roots. After 15,30,60,120 minutes of feeding (MAF), plants were removed and the roots washed using plenty of water, and then roots and flag were sampled to measure radioactivity using LKD1216 liquid scintillation counter.

14C tracing experiment and dynamic analysis

14C-CO2 was fed through gas chamber into flag leaf on May 12th (early filling stage) and May 29th (late filling stage). In the 48h after feeding, measured the radioactivity of the flag leaf in vivo every hour using PRS Miniscaler /Ratemeter (US Technical Associate) and simulated the dynamic equation of assimilates exporting. The method was described previously (Duan LS et al.1998).

Investigation of transportation tissues in wheat root and internode below spike

The roots and internode below spike were sampled on May 16 (early filling stage) as 0.5 cm and fixed in 50% FAA, then prepared slices embed in paraffin and inspected under microscope. Measured the diameter of central vessel and pericycle of roots and the diameter of peripheral vessel. The cross-sectional area of vessel was calculated by the formula(s=πab/4, a and b are maximum diameter in two directions). 10 random slices from the same position of different plants were inspected for each treatment.

Measurement of transpiring rate, stomatal resistance, water use efficiency and roots validity

The transpiration rate, stomatal resistance, photosynthesis rate were detected on May 21st(middle filling stage), using Li-6200 photosynthesis systems. Water use efficiency based on single plant was calculated as photosynthesis rate/transpiration rate. Root validity measuring method was described by He ZP(1993).


Water transpiring rat, stomatal resistance of flag leaf and water use efficiency

Under water deficiency, the stomatal resistance was increase and both the transpiration rate and water use efficiency were decreased. Chemical regulation using HK could reduce the stomatal resistance under water deficiency and promote water transpiration (Table 1). Transpiration is not only involved in photosynthesis, but also is the motivity for the plant to absorb and transport water in long distance, so to maintain the transpiration at a proper level maybe vital for the plant to keep the balance between water and assimilates metabolism. The water use efficiency based on single plant is a good index for the above balance. On this term, chemical regulation increased water use efficiency by 25.0% and 21.7% under water abundance and deficiency conditions respectively, which implied that the harmoniousness between water and assimilates was improved.

Table 1. Effects of chemical regulation on transpiration rate, stomatal resistance and water use efficiency of single plant under water deficiency *


Transpiration rate

Stomatal resistance

Water use efficiency of single plant (g/g)


5.7 a

96 c

2.3 c


5.3 a

130 d

2.8 a


3.6 c

175 a

2.0 d


4.8 b

165 b

2.5 b

Means within a column followed by different letters are significantly different at p≤0.05 level, Duncan’s Multiple Range. Same as in the following tables.

3H2O distribution in roots and flag leaf

Under water deficiency, the 3H2O content in roots had reached a high level 15 MAF,and then went up again from 30MAF to 120MAF. Under water abundance, both the 3H2O content in roots and in flag leaf showed a peak at 30MAF.It was indicated that wheat plants absorbed water faster and longer but less in total under water deficiency than that under water abundance. Chemical regulation promoted 3H2O accumulation in 15 MAF and 30MAF in roots and increased 3H2O accumulation within 120MAF in flag leaf, which showed a proof for the water absorbing and transporting from roots to flag leaf were enhanced and implied that water use was improved (Table 2).

Micro-structure and vitality of roots.

The roots validity was weaker under water deficiency than that in abundance water, especially for the roots distributed in deeper soil layers. Compared with the non-regulation control, the validity of roots of the HK treatment in 0-20cm, 20-40cm and 40-100cm were 13.6%, 5.4% and 8.3% higher under water deficiency, while 30.2%, 33.0% and 16.6% 33.0% higher under water abundance (Table 3), which illuminate that the functions of roots were augmented by chemical regulation using HK. Table 4 showed the structure changes of primary roots under water stress and chemical regulation. Both the long axis and the short axis of central vessel were longer, which lead to the sectional area of central vessel was enlarged under water deficiency. The similar situation was found for the pericycle. These might be the adaptive responses of wheat plant to water scarcity. The chemical regulation reduced the diameter and cross-sectional area of central vessel while it increased the pericycle area and the peripheral vessel number of primary, which might be a good explanation for the improvement of roots functions and water balance of the whole plant.

Table 2. Effects of chemical regulation on 3H2O contents in wheat roots and flag leaf in different minutes after 3H2O feeding (MAF) through roots under water deficiency



3H2O content in 15 MAF (Bq/g)

3H2O content in 30 MAF (Bq/g)

3H2O content in 60 MAF (Bq/g)

3H2O content in 120 MAF (Bq/g)



322.2 A

361.1 B

216.7 B

166.7 B



288.9 B

433.3 A

227.8 A

161.1 B



122.2 D

55.6 D

133.3 C

233.3 A



194.4 C

111.1 C

100.0 D

161.1 B

Flag leaf


61.1 b

73.3 c

63.9 a

62.8 a



75.0 a

80.6 b

58.3 b

62.2 a



50.0 c

62.2 d

47.8 c

47.2 b



72.2 a

88.9 a

55.6 b

50.0 b

Table 3. Effects of chemical regulation on validity of wheat roots in different soil layers

Tillage treatment

Validity of roots in different soil layer (ug/gh)





27.5 c

39.2 b

30.5 b


35.2 a

42.0 a

40.3 a


25.3 d

28.1 c

32.2 b


30.0 b

30.0 c

34.0 b

Table 4. Effects of chemical regulation on vascular of wheat primary roots under water deficiency


Central vessel


Peripheral vessel number

Long axis (mm)

Short axis (mm)

Area (mm2)

Long axis (mm)

Short axis (mm)

Area (mm2)


0.080 a

0.072 b

0.457 b

0.213 a

0.194 a


8.60 a


0.087 a

0.081 a

0.554 a

0.208 a


3.127 a

8.56 a


0.052 b

0.064 c

0.275 c

0.167 b


2.286 c

7.50 c


0.086 a

0.078 a

0.531 a

0.202 a

0.185 b

2.640 b

8.33 b

14C assimilates export from flag leaf and structure of the internode below spike

The dynamic equation of 14C-assimilates exporting from flag leaf and their characteristic parameters were listed as Table 5. Assimilates exportation from flag leaf was slowed down with filling stage proceeding, and both of the two illuminating parameters for export speed:T1/2 (time for the 14C assimilates to export by 50%) and E12h (exported ratio in the total fed dosage in 12 hours after feeding) . The T1/2 for WC, WR, SC and SR in late filling stage was 6.54h, 4.86h, 6.13h and 2.97h behind that in early filling stage, indicating that water deficiency promotes and chemical regulation accelerates the export of assimilates from the flag leaf. This could be also supported by that E12h.In early filling stage, E12h was raised by 5.36% under water abundance and 5.09% under water deficiency. In later filling stage, E12h rose by 35.64% and 34.40% under abundance and deficiency water condition. The internode diameter, vascular bundles number and sectional area are key factors for the assimilate flow between source and sink organs. As showed in Table 6, both the peripheral vessel bundle number and their sectional area were lower under water stress compared with those under water abundance, which might be the key limitation for water transportation and assimilates reallocation. The chemical regulation had no remarkable effects on vascular bundle number of internode below spike while enlarged vessel bundle area significantly by 18.2% and 39.9% under enough and scarce water condition, which did benefits to improve water status, assimilates transportation and distribution.

Table 5. Effects of chemical regulation on exporting dynamics equation of 14C-assimalates from flag leaf during grain filling under water deficiency



Exporting dynamics equation



E12h (%)



Y=30.34ln(x) -9.2086

0.9795 *





Y=28.414ln(x) -0.8798

0.9853 *





Y=29.839ln(x) -8.7868

0.9764 *





Y=27.464ln(x) +2.209

0.9920 *





Y=4.1316x -6.109

0.9734 *





Y=29.432-ln(x) -14.179

0.9599 *





Y=4.1099x -4.6668

0.9752 *





Y=30.787-ln(x) -16.489

0.9501 *



X for time after feeding (h),Y for exporting ratio of 14C-assimalates(%),* for significant at 0.01 level.

Table 6. Effects of chemical regulation on vascular of wheat internode below spike under water deficiency


Peripheral vessel bundle Number

Long axis length

Short axis length (um)

Vessel bundle area (um2)


21.0 a

10.1 b

9.0 c

122.0 b


22.0 a

14.2 a

11.2 b

126.0 b


17.5 b

13.6 a

11.0 b

117.9 c


17.5 b

15.7 a

13.3 a

164.9 a


Wheat plant responses to water deficiency in series of physiological process including changes in roots and internode structure, roots vitality, leaf transpiration and stomatal resistance, assimilates synthesis and distribution, which involved in water absorbing, transporting, losing and utilizing and showed in the end as water use efficiency. HK foliage spraying, as an example of the chemical regulation method, enlarged the pericycle sectional area and the peripheral vessel number in primary roots and internode below spike, increased roots vitality and promoted 3H2O absorbing and transporting to shoots, enhanced stomata impedance, maintained transpiration rate, suspended the reduction of 3H2O in flag leaf. As the result, the water use efficiency based on single plant was elevated. The above positive and compensative effects implied the encouraging potentiality of employing plant growth regulators to enhance the drought tolerance or resistance of crops and eliminate the loss of yield and quality caused by water shortage.


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