DESC:FIELD OF THE INVENTION
[001] This invention relates to formulations to improve bud-break in a variety of trees or crops including deciduous fruit trees, grape vines and shrubs, or bushes, to its method of preparation, and to its method of use.

BACKGROUND OF THE INVENTION
[002] “Bud break” in deciduous fruit trees such as apples, plums, nectarines, peaches, grapes and pears, grape vines and shrubs, or bushes is very essential to facilitate their new growth. It is the first stage of the growth cycle which finally yields fruits. These buds appear in summer of previous growth cycle and are covered in scales. During winter dormancy, they turn brown until the vine and shrub begins the process of bud break.
[003] These Deciduous fruit trees, grape vines and shrubs, or bushes normally require enough winter chilling hours to produce a crop of fruit. The amount of chilling required depends upon the kind of fruit and the cultivar. Problems can arise in tropical and semi-tropical climates when there is no chilling, or in desert climate zones where there is insufficient chilling. The result of these problems can be growth abnormalities such as delayed and uneven blossoming, poor leaf cover, and insufficient or inconsistent fruit-set and reduced fruit size can occur. At the end of a vegetation period, such plants cease growing after bearing fruit and form buds. This budding enables a meristem, which has undifferentiated leaf and blossom structures, to survive under the unfavorable environmental conditions of winter because buds, in the latent stage, are much more resistant to frost and low temperatures than active tips of vegetation. Without sufficient chilling, the buds can be devoid of a commercially acceptable yield of fruit.
[004] Many horticultural regions do not experience cold winters, and changes in local climatic conditions are increasingly exacerbating the problem of poor winter chill. It is possible to overcome the effects of poor winter chill and in some plants or to augment the benefits of good winter chill by chemical treatment of plants in the latter part of dormancy.
[005] In the past, several measures have been followed like bending and twisting of canes to increase bud break which gradually became impractical owing to the amount of labor involved. Moreover, these operations sometimes resulted in damage to the sprouted buds.
[006] Another method to overcome the effects of poor winter chill or to augment the benefits of good winter chill is by chemical treatment of plants in the latter part of dormancy. The simplest chemical “bud-breaker agents” are 4, 6-dinitro-o-cresol, now deregulated in most countries, and hydrogen cyanamide (HC), which is still widely used in some countries. Unfortunately, HC is a very toxic chemical and was excluded from Annex I to Directive 91/414/EEC in October 2008, and its continued use in non-European countries is under threat. The chief concerns are damage to non-target crops and human exposure, which can produce contact dermatitis, respiratory and gastrointestinal tract irritation, headaches, and liver damage when exposure is followed by alcohol consumption. In addition, in some crops HC produces too strong a response and can produce undesirable side effects. For example when applied to yellow-fleshed kiwifruit varieties such Hort16A (Actinidia chinensis Planch. var. chinensis ‘Hort16A’) and Go1d3 (Actinidia chinensis Planch, var. chinensis‘Zesy002’), HC produces too many flowers and increases the frequency of unwanted lateral flowers.
[007] In view of the above, safer bud-break formulations with more refined activity in plants are clearly needed for overcoming the aforementioned problems.

DESCRIPTION OF THE INVENTION
[008] The present invention is directed towards a formulation for bud breaking which is safe to use and ameliorates the disadvantages of the earlier known bud breaking agent. The said formulation also provides safe bud breakings two days in advance as compared to hydrogen cyanamide and also results in high yield.
[009] In one aspect, the present invention provides formulation for bud breaking, the formulation comprising at least one compound of formula (I),

Wherein,
R= -C(O)-Me, -C(O)-(CH)mCH3, -C(O)-CF3, -C(O)-CHF2, -C(O)- CH2F, -C(O)-CCl3, -C(O)-CHCl2, C(O)-CH2Cl, CBr3, C(O)-CHBr2, C(O)-CH2Br, -C(O)-CH2OCH3, -C(O)-CH2CH2OCH3, H, -CH2-Me, -C H2-(CH)mCH3, -CH2-CF3, -CH2-CHF2, -CH2-CH2F, -CH2-CCl3, -CH2-CHCl2, CH2-CH2Cl, CBr3, CH2-CHBr2, CH2-CH2Br,- CH2-CH2OCH3, , , , , , ;
n=1,2 or 3;
m=0, 1, 2 or 3;
X and Y= H, amino acids selected from S-acetyl cysteine, Glu, Cys, Asp, Gly, Arg, His, Ala, Leu, Ile, Val, Lys, Val, Pro, Met, Gln.
[010] In an embodiment of the invention, the specific compounds of formula (I) includes compounds where R= H, Me or-C(O)-Me, n = 1 or 2, Y = OH or NH-CH2-C(O)OH, and /or X = -C(O)-Me, -C(O)- -CH2--CH2-CH(NH2)-C(O)OH.
[011] In another embodiment the specific compounds of formula (I) of the composition of the present invention include following compounds 1, 2, 3 & 4.

[012] In an aspect, the formulation may further comprise an agrochemically acceptable additive. The additive may be a surfactant selected from, but not limited to, Amine ethoxylate, EO/PO block copolymer, Ether sulfate, Fatty alcohol alkoxylate, Fatty alcohol ethoxylate, Methyl oleyl taurate, Olefin sulfonate, Sulfosuccinate.
[013] In an embodiment of the invention, the compound of formula (I) and the surfactant is in a ratio in the range of 0.5 – 2 : 0.5 – 2. A preferred ratio of the compound of formula (I) and the surfactant is 1:1.
[014] In another aspect, the formulation is in the form of Wettable powder, soluble concentrate, water soluble granule, water dispersible granule, emulsifiable concentrate, oil dispersion, gel.
[015] In an embodiment of the invention, the formulation may further comprise conventional bud break agents selected from Hydrogen Cyanamide, Rape seed Oil Methyl ester; Potassium Nitrate (KNO3), Salicylic acid, Thidiazuron, Benzyl adenine, Fresh extract of Garlic, Allicin, Choline salt, Thibarbituric acid, 6-chloro-2-mercapto benzoic acid, Vitamin B6, Gallic acid, Rosolic acid, Glucosinolate (95% Allyl isothiocyanate), N-Acetly L Cysteine, N-Acetyl L Methionine, Sodium 4 amino salicylate dihydrate and/or L Glutathione (GSH-reduced form).
[016] The invention also relates to a process for preparation of a formulation comprising the steps of preparing an aqueous solution of compound of formula (I) and adding required additives to the same.
[017] The invention also relates to a process of inducing bud break comprising the steps
i. Dissolving a formulation comprising compound of formula I in water; and
ii. Application of the aqueous solution formulation comprising compound of formula I at the prescribed time after pruning.

[018] In an embodiment, the prescribed time is preferably on the day immediately following the day of pruning but necessarily within 48 hrs. after pruning.
[019] In an embodiment, the application of the formulation is by using a suitable cotton cloth or sponge with good water absorption capacity as a carrier for the formulation and swabbing on the four apical buds of each cane starting from the lower one (4th bud) and towards the apical one.
[020] The invention further relates to use of a formulation comprising compound of formula (I) for inducing bud break. In an aspect, the formulation may further comprise conventional bud break agents.
[021] Advantageously, the present invention provides a formulation for bud break, the formulation comprising at least one compound of formula (I) as defined. This formulation is safe for use and also avoids toxic chemicals conventionally used for bud break. Moreover, the shelf life of the formulation is very high compared to the shelf-life of conventionally used chemicals such as Hydrogen Cyanamide. It is found that the use of this formulation results in early bud break and in high yields.

EXAMPLES
[022] The following examples are illustrative of the invention but not limitative of the scope thereof:
[023] Example 1
[024] Soluble Liquid Formulation-SL
[025] 1.05 g of L-?Glu-S-Ac-L-Cys-Gly-OH powder (Compound No. 4) + 1.05 g of Castor oil Ethoxylate surfactant + 0.10g of Ascorbic acid was dissolved in 60 ml. distilled water. This solution is stirred to get clear SL formulation. The above SL formulation was dissolved in one litre water to give the final working solution.

[026] Example 2
[027] Soluble Powder Formulation- SP
[028] 1.05 g of L-?Glu-S-Ac-L-Cys-Gly-OH powder (Compound No. 4) + 1.05 g of SLS (Sodium lauryl sulfate) surfactant + 0.10g of Ascorbic acid + 2.8g of anhydrous Ammonium sulfate are mixed together thoroughly to get homogeneous fine powder (SP). This 5.0g of SP formulation was dissolved in one litre water to give the final working solution.

[029] Example 3
[030] Soluble Powder Formulation- SP
[031] 1.05 g of L-?Glu-S-Ac-L-Cys-Gly-OH powder (Compound No. 4) + 1.05 g of SLS (Sodium lauryl sulfate) surfactant + 0.10g of Ascorbic acid + 2.8g of anhydrous Sodium sulfate are mixed together thoroughly to get homogeneous fine powder (SP). This 5.0g of SP formulation was dissolved in one litre water to give the final working solution.

Application time:
pH of this solution was adjusted so that the final working solution was in the slightly acidic range of 6 to 6.5. Next the canes were individually swabbed starting from the bud on the lower end of the cane. Only the top four buds were swabbed using cotton or cloth of good water absorbing capacity which was dipped in the solution.

Field Trial data for Percent Bud break:
Season 2016-17:
Four field trials were undertaken on different grape varieties at multiple locations. The individual formulations were applied to ten vines each which were pruned the previous day. This applied to all the four trials undertaken. Application was made by swabbing the apical four buds of all canes for each of the vine. Observations for bud break were recorded at 14 DAA (Days after application) for number of buds sprouted of the four buds applied for ten individual canes of each of the ten vines and their total was taken into account to calculate the % bud break.
Bud Break was recorded 14 DAA for 400 buds in all per treatment @ 40 buds/vine.

Table 1
Variety: Sharad Seedless; Location: Nasik

Percent Bud Break for individual treatment (Total No of buds sprouted / 400 buds)
Vine No. Treatment
Vitamin B6
@ 200 ppm N Acetyl L Cysteine
@ 5 mM L Glutathione @ 5 mM N-Acetyl L Methionine@ 5 mM Formulation of the Compound 4 @ 5 mM H2CN2
2.5%
(Farmer’s
Practice)
1 24 17 32 25 38 36
2 22 16 33 27 36 33
3 23 17 31 29 38 34
4 22 14 31 25 39 35
5 23 16 33 24 38 33
6 21 17 31 28 39 34
7 22 18 30 25 37 35
8 20 15 32 28 40 37
9 21 16 34 24 39 36
10 21 15 32 23 38 34
Total
Buds Sprouted 219 161 319 258 382 347
Percent Bud Break 55% 40% 80% 65% 96% 87%

Inference:

1. Vitamin B6 @ 200 ppm, N Acetyl L Cysteine @ 5 mM and L Glutathione @ 5 mM resulted in 55%, 40% and 80% bud break resp.

2. Formulation of the Compound 4 @ 5 mM resulted in 96% bud break.

3. It can be concluded from the above data that formulation of the Compound 4 @ 5 mM was the best compound for inducing higher bud break.

4. Moreover, current formulation of the Compound 4 @ 5 mM results in the highest percent bud break of 96% which is higher than 87% for that of 2.5 % H2CN2 (Farmer’s Practice).

Table 2
Variety: Sonaka; Location: Sangli

Percent Bud Break for individual treatment (Total No of buds sprouted / 400 buds)
Vine No. Treatment
Vitamin B6
@ 200 ppm N Acetyl L Cysteine
@ 5 mM L Glutathione @ 5 mM N-Acetyl L Methionine@ 5 mM Formulation of the Compound 4 @ 5 mM H2CN2
2.5%
(Farmer’s
Practice)
1 22 19 32 26 39 34
2 23 16 30 25 37 35
3 25 17 34 30 37 35
4 22 19 31 28 38 34
5 21 18 32 25 39 34
6 24 18 33 29 37 33
7 20 19 34 28 38 36
8 19 20 31 27 39 35
9 23 18 34 25 38 35
10 24 18 34 24 37 34
Total
Buds Sprouted 223 182 325 267 379 345
Percent Bud Break 56% 46% 81% 67% 95% 86%

Inference:

1. Vitamin B6 @ 200 ppm, N Acetyl L Cysteine @ 5 mM and L Glutathione @ 5 mM resulted in 56%, 46% and 81% bud break resp.

2. Formulation of the Compound 4 @ 5 mM resulted in 95% bud break.

3. It can be concluded from the above data that formulation of the Compound 4 @ 5 mM was the best compound for inducing higher bud break.

4. Moreover, formulation of the Compound 4 @ 5 mM results in the highest percent bud break of 95% which is higher than 86% for that of 2.5 % H2CN2 (Farmer’s Practice).

Table 3
Variety: Nanasaheb Purple; Location: Nasik
Percent Bud Break for individual treatment (Total No of buds sprouted / 400 buds)
Vine No. Treatment
Vitamin B6
@ 200 ppm N Acetyl L Cysteine
@ 5 mM L Glutathione @ 5 mM N-Acetyl L Methionine@ 5 mM Formulation of the Compound 4 @ 5 mM H2CN2
2.5%
(Farmer’s
Practice)
1 21 22 34 24 38 33
2 24 18 33 23 38 32
3 22 16 32 28 39 33
4 24 17 30 27 37 35
5 25 19 31 26 36 33
6 22 18 32 25 38 34
7 23 17 33 24 37 35
8 21 22 32 23 38 33
9 22 19 33 27 36 32
10 23 19 34 25 37 33
Total
Buds Sprouted 227 187 324 252 374 333
Percent Bud Break 57% 47% 81% 63% 94% 83%

Inference:

1. Vitamin B6 @ 200 ppm, N Acetyl L Cysteine @ 5 mM and L Glutathione @ 5 mM resulted in 57%, 47% and 81% bud break resp.

2. Formulation of the Compound 4 @ 5 mM resulted in 94% bud break.

3. It can be concluded from the above data that Formulation of the Compound 4 @ 5 mM was the best compound for inducing higher bud break.

4. Moreover, formulation of the Compound 4 @ 5 mM results in the highest percent bud break of 94% which is higher than 83% for that of 2.5 % H2CN2 (Farmer’s Practice).

Table 4
Variety: Thompson Seedless; Location: Nasik
Percent Bud Break for individual treatment (Total No of buds sprouted / 400 buds)
Vine No. Treatment
Vitamin B6
@ 200 ppm N Acetyl L Cysteine
@ 5 mM L Glutathione @ 5 mM N-Acetyl L Methionine@ 5 mM Formulation of the Compound 4 @ 5 mM H2CN2
2.5%
(Farmer’s
Practice)
1 20 24 35 26 37 32
2 25 19 36 24 36 34
3 23 17 32 26 38 35
4 21 16 33 26 36 33
5 22 18 30 24 37 34
6 24 17 34 27 39 33
7 21 19 35 23 38 36
8 23 21 33 25 37 34
9 24 21 34 26 38 33
10 22 19 35 24 39 34
Total
Buds Sprouted 225 191 337 251 375 338
Percent Bud Break 56% 48% 84% 63% 94% 85%

Inference:

1. Vitamin B6 @ 200 ppm, N Acetyl L Cysteine @ 5 mM and L Glutathione @ 5 mM resulted in 56%, 48% and 84% bud break resp.

2. Formulation of the Compound 4 @ 5 mM resulted in 94% bud break.

3. It can be concluded from the above data that Formulation of the Compound 4 @ 5 mM was the best compound for inducing higher bud break.

4. Moreover, Formulation of the Compound 4 @ 5 mM results in the highest percent bud break of 94% which is higher than 85% for that of 2.5 % H2CN2 (Farmer’s Practice).

Conclusions based on four multi location field trials in Season 2016-17:
1. Formulation of the Compound 4 @ 5 mM results in the highest percent bud break in all the four field trials despite having different grape varieties and was also found to be better than Farmer’s practice of using H2CN2 @ 2.5%.
2. There was no adverse impact on bud break despite application being done in temperature as high as 31 degree centigrade.

Percent Bud break data for Season 2017-18:
Three field trials were undertaken on different grape varieties at multiple locations. The individual formulations were applied to ten vines each which were pruned the previous day. This applied to all the three trials undertaken. Application was made by swabbing the apical four buds of all canes for each of the vine. Observations for bud break were recorded at weekly intervals viz.7 DAA and 14 DAA for number of buds sprouted of the four buds applied for ten individual canes of each vine and their total was taken into account to calculate the % bud break.
Bud Break was recorded at 7 DAA and 14 DAA for 400 buds in all per treatment @ 40 buds/vine.
Based on the observations at 7 DAA / 14 DAA, it can be concluded that the composition comprising formulation of the Compound 4 @ 5 mM resulted in higher and faster bud break than H2CN2
Table 5
Trial No. Treatment
H2CN2 Formulation of the Compound 4 @ 5 mM
% BB 7 DAA % BB 14 DAA % BB 7 DAA % BB 14 DAA
1 30.8 88.0 53.3 92.0
2 35.8 93.0 52.3 94.5
3 41.3 94.0 62.8 95.5
Mean 36.0 91.7 56.3 94.0
Percent Bud break data for Season 2018-19:
Three field trials were undertaken on different grape varieties at multiple locations. The individual formulations were applied to ten vines each which were pruned the previous day. This applied to all the three trials undertaken. Application was made by swabbing the apical four buds of all canes for each of the vine. Observations for bud break were recorded at weekly intervals viz.7 DAA and 14 DAA for number of buds sprouted of the four buds applied for ten individual canes of each vine and their total was taken into account to calculate the % bud break.
Bud Break was recorded at 7 DAA and 14 DAA for 400 buds in all per treatment @ 40 buds/vine.
Based on the observations at 7 DAA / 14 DAA, it can be concluded that the composition comprising formulation of the Compound 4 @ 5 mM resulted in faster bud break than H2CN2.
Table 6
Trial No. Treatment
H2CN2 Formulation of the Compound 4 @ 5 mM
% BB 7 DAA % BB 14 DAA % BB 7 DAA % BB 14 DAA
1 56.3 94.0 75.0 95.0
2 64.5 91.0 84.5 94.0
3 71.5 97.0 88.0 97.5
Mean 64.0 94.0 82.5 95.5
Percent Bud break data for Season 2019-20:
Three field trials were undertaken on different grape varieties at multiple locations. The individual formulations were applied to ten vines each which were pruned the previous day. This applied to all the three trials undertaken. Application was made by swabbing the apical four buds of all canes for each of the vine. Observations for bud break were recorded at weekly intervals viz.7 DAA and 14 DAA for number of buds sprouted of the four buds applied for ten individual canes of each vine and their total was taken into account to calculate the % bud break.
Bud Break was recorded at 7 DAA and 14 DAA for 400 buds in all per treatment @ 40 buds/vine.
Based on the observations at 7 DAA, it can be concluded that the composition comprising formulation of the Compound 4 @ 5 mM resulted in higher and faster bud break than H2CN2.
Table 7
Trial No. Treatment
H2CN2 Formulation of the Compound 4 @ 5 mM
% BB 7 DAA % BB 14 DAA % BB 7 DAA % BB 14 DAA
1 57.0 83.0 63.0 94.0
2 62.0 87.0 71.0 93.0
3 61.0 90.0 68.0 91.0
Mean 60.0 86.7 67.3 92.7
Harvest data for Season 2018-19:
Field Trial was initiated in October 2018 on Flame Seedless variety and grapes were harvested in March 2019. In this field trial, the harvest data (Number of Grape bunches and their weight) was recorded for five vines each treated with H2CN2 and formulation of the Compound 4 @ 5 mM respectively.
Table 8
Number and Bunch Weight per five vines harvested
Treatment Wt.(g) Bunch No. Single
Bunch Wt. % rise in Wt. % rise (Bunch Nos.)
H2CN2
(Farmer’s Practice) 21200 105 201.9 --- ---
Formulation of the Compound 4 @ 5 mM 27000 132 204.5 27.4% 25.7%

Conclusion:
Usage of formulation of the Compound 4 @ 5 mM resulted in 27% rise in yield and 26% rise in the no. of grape bunches per five vines harvested viz-a-viz those treated with H2CN2.

Harvest data for Season 2019-20:
Field Trial was initiated in October 2019 on Thompson Seedless variety and grapes were harvested in March 2020. In this field trial, the harvest data (Number of Grape bunches and their weight) was recorded for five vines each treated with H2CN2 and of Formulation of the Compound 4 @ 5 mM respectively.
Table 9
Number and Bunch Weight per five vines harvested
Treatment Wt.(g) Bunch No. Single
Bunch Wt. % rise in Wt. % rise (Bunch Nos.)
H2CN2 31400 110 285.5 --- ---
Formulation of the Compound 4 @ 5 mM 43000 145 296.6 36.9 31.8

Conclusion:
Usage of formulation of the Compound 4 @ 5 mM resulted in 37% rise in yield and 32% rise in the no. of grape bunches per five vines harvested viz-a-viz those treated with H2CN2.
[032] The foregoing description of the invention has been set merely to illustrate the invention and is not intended to be limiting. Since the modifications of the disclosed embodiments incorporating the spirit and substance of the invention may occur to the person skilled in the art, the invention should be construed to include everything within the scope of the disclosure.
,CLAIMS:We Claim:
1. A formulation for bud breaking comprising at least one compound of formula (I)

Wherein,
R= -C(O)-Me, -C(O)-(CH)mCH3, -C(O)-CF3, -C(O)-CHF2, -C(O)- CH2F, -C(O)-CCl3, -C(O)-CHCl2, C(O)-CH2Cl, CBr3, C(O)-CHBr2, C(O)-CH2Br, -C(O)-CH2OCH3, -C(O)-CH2CH2OCH3, H, -CH2-Me, -C H2-(CH)mCH3, -CH2-CF3, -CH2-CHF2, -CH2-CH2F, -CH2-CCl3, -CH2-CHCl2, CH2-CH2Cl, CBr3, CH2-CHBr2, CH2-CH2Br,- CH2-CH2OCH3, , , , , , ;
n=1,2 or 3;
m=0, 1, 2 or 3;
X and Y= H, amino acids selected from S-acetyl cysteine, Glu, Cys, Asp, Gly, Arg, His, Ala, Leu, Ile, Val, Lys, Val, Pro, Met, Gln.

2. The formulation as claimed in claim 1, wherein R= H, Me or-C(O)-Me.

3. The formulation as claimed in claim 1, wherein n = 1 or 2.
4. The formulation as claimed in claim 1, wherein Y = OH or NH-CH2-C(O)OH.

5. The formulation as claimed in claim 1, wherein X = -C(O)-Me, -C(O)- -CH2--CH2-CH(NH2)-C(O)OH.

6. The formulation as claimed in claim 1, wherein the compound of formula (I) is

7. The formulation as claimed in claims 1-6 wherein the formulation comprises acceptable agrochemical additives.

8. The formulation as claimed in claim 7, wherein the additive is a surfactant.

9. The formulation as claimed in claims 1-7, wherein the compound of formula (I) and the surfactant is in a ratio in the range of 0.5 – 2 : 0.5 – 2.

10. A method of inducing bud break comprising the steps of dissolving the formulation as claimed in claim 1 in water; and application of the formulation at the prescribed time after pruning.

11. Use of a formulation comprising at least one compound of formula (I) as claimed in claims 1-9 for inducing bud-break.