• Field of the invention
This invention relates to a novel plant extract useful for the control of mosquitoes and a process for the preparation of said extract. The invention particularly relates to a novel plant extract containing Lectins having potent anti- mosquito activity. The extract of the present invention is expected to be active against the control of the mosquito belonging to the genus Aedes . The extract is prepared from the plant Sapindus emarginatus (belonging to the Family : Sapindaceae)
• Background
Mosquitoes are small insects which include some 3200 species and subspecies belonging to 37 genera, all contained in the family Culicidae. Mosquitoes have a world-wide distribution; they occur throughout the tropical and temperate regions and extend their range northwards into the Arctic Circle. The only areas from which they are absent are Antarctica, and a few islands. They are found even on elevations at 5500 m and down mines at depths of 1250 m below sea level (Service, 2000).
Of all the insects that transmit diseases (vectors), mosquitoes represent the greatest menace. Indeed, WHO has declared the mosquito "Pubic Enemy Number One", because the mosquitoes serve as vectors of various dreadful diseases (WHO, 1996). Such human diseases include malaria, filaria (example: elephantiasis), Japanese encephaUtis (brain fever), and dengue. The most important disease transmitting and nuisance causing mosquitoes belong to the genera Anopheles, Culex, Aedes, Psorophora, Haemagogus and Sabethes. In India, the various species of Anopheles, Culex and Aedes are important as carriers of pathogens or parasites capable of causing several diseases in human beings. In tropical countries Duke India, a considerable portion of the National Health Budget is being spent on the control of especially mosquito-borne diseases.
• Strategies to control mosquito
Many control strategies for mosquitoes have been suggested and followed since the ancient times. Among them, chemical insecticides are the most important components of integrated vector control. The discovery of dichloro-diphenyl trichloroethane (DDT) was a break-through in the mosquito control programmes. DDT was highly effective for killing indoor resting mosquitoes, when it was sprayed on the walls and roofs of houses. However, because of their persistence in the environment and accumulation in food-chains, DDT and other organ chlorine insecticides

eventually became undesirable for use to kill larvae (larvicides) of mosquitoes and other insects. Besides, pyrethroid such as permethrin, and deltamethrin are being used as larvicides, indoor residual spraying to kill adult mosquitoes, and for treatment of mosquito nets to keep away adult mosquitoes. It is remarkable that the use of all these chemical compounds induce resistance in mosquitoes as well as development of cross resistance to other chemical compounds, thereby limiting the number of effective alternatives suitable for mosquito control by chemical methods (Mazzarri & Georghiou, 1995).
The extensive application of synthetic chemical insecticides has also caused irreparable hazards to the environment and humans. The World Health Organization has estimated that about 3 million people are affected annually by acute poisonings caused by pesticides. Moreover, additional problems like lack of understanding of their proper use, non-availability of suitable application equipments and inadequate storage conditions have intensified the risk out of indiscriminate use of chemical insecticides especially in developing countries. Furthermore, routine and indiscriminate application of such chemical insecticides has been shown to cause a high negative effect on non-target beneficial organisms (Shah, 1975).
• Biocidal potential of plants
The problems of high cost of chemical insecticides and development of resistance invariably by vector mosquito species to the synthetic insecticides have revived interest in exploring the potentials of plants it contouring insect vectors.
• Prior art : Biomolecules from plants with antimosquito properties
Plants are a rich source of bioactive compounds and they appear to synthesize a variety of primary and secondary metabolites to serve as defense chemicals or molecules that are active against insect attack. More than 2500 plants were identified with insecticidal properties against insect pests and vectors. Phytochemicals with insecticidal property can be extracted from either whole plants or specific parts of the plant, depending on the distribution of specific activity of the desired biomolecules. These biomolecules, in particular, may kill the mosquitoes, or reduce / inhibit feeding, egg laying, growth and development of mosquitoes (Grainge & Ahmed, 1988). Therefore, such biomolecules offer unique advantages over synthetic insecticides, as they are

effective, easily biodegradable (eco-friendly), inexpensive, and with a remote chance for the development of resistance by target mosquito species.
One of the earliest reports of the use of plant extracts against mosquito larvae is credited to Campbell et al (1933) who found that plant alkaloids from the Russian weed Anabasis aphylla killed mosquito larvae. Among many known phytochemicals, the widely studied biological insecticide with potent anti mosquito activity is the azadirachtin, one of the active ingredients isolated from neera seed, Azadirachata indica which originated in Indian subcontinent (Schmutterer & Doll, 1993). Azadirachtin is a secondary metabolite (tetranotriterpenoid compound) and acts as contact and systemic poison. Azadirachtin in the neem seed extract has been proved to be the main agent for combating mosquitoe larvae, since it not only repels them but also disrupts their growth and reproduction (Su & MuUa, 1999). However, it is notable that azadirachtin does has an important disadvantage as it degrades rapidly upon exposure to sunlight, resulting in great loses of its anti-mosquito activity (Johnson et al 2003).
In view of serious limitations perceptible with the neem product (azadirachtin), there is a need to search for new source of botanicals in order to develop environmentally safe, stable biodegradable, low cost, and indigenous method for mosquito control strategy. Such biomolecules, when detected, should allow their use with minimum care by individual and communities in specific situations.
• Lectins as potential biocides
Lectins are proteins/glycoproteins with an about to reversibly interact with specific carbohydrate structures present on cell surfaces, extracellular matrices, or secreted glycoproteins (Sharon & Lis, 2004). When lectins possess two or more carbohydrate-binding sites, they can cause aggregation or agglutination of cells, there by they are also called as agglutinins.
The occurrence of lectins in living materials was first discovered by Still mark in 1888 (as cited in Goldstein & Hayes, 1978) in the castor-bean {Ridings communis) extracts which he called as rising. Thereafter, the ubiquitous occurrence of lectins have been reported in a wide range of microbes, plants, and animals (Bald & Gelding, 1975). The lectins have been detected in different parts of plants and animals. In the case of plants, for example, the lectins are detectable in leaves.

barks, fruits, roots, and seeds, with it highest concentration invariably recorded in seed kernels. It is also notable that different living resources do possess lectins with different carbohydrate binding properties.
• Biomedical appKcations of lectins
Purified lectins molecules with unique carbohydrate-binding specificities are being extensively used as valuable tools or probes in many areas of scientific research including agriculture, biotechnology, and biomedical sciences (Bog-Hansen & Freed, 1990; Sharon & Lis, 2004).
• Some of the significant biomedical appUcations of lectins are :
1. Blood group determination
2. Identification of bacteria
3. Glycoconjugates purification and cell fractionation
4. Isolation and characterization of polysaccharides and glycocnjugates in cancer research
5. Early detection of cell membrane alterations
6. Surface carbohydrate changes that accompany neoplastic process
7. Immunological studies involving misogynic stimulation or non-mitogenic activation of peripheral lymphocytes, induction of cytokine Production and chromosomal analysis for diagnosis of genetic disorders or chronic diseases characterized by immunodeficiency.
• Biocidal potential of lectins
It is notable that the native functions of plant or animal lectins are not fully understood. However, many functions have been envisaged for plant lectins. Among the various roles that have been attributed to plant lectins is that of defence against pathogens and phytophagous insects (Peumans & Van Dammed, 1995; Jansen et a/., 1976). This protective activity is in accordance with the observation that most plant lectins are not targeted against plant carbohydrates, but preferentially bind foreign glycans (Peumans et al, 2000). Several plant lectins have been found to be effective in reducing insect survival, development and fecundity of a wide range of insect orders including Homoptera (Down et a/., 1996), Lepidoptera (Fitches et al, 1997), Hymenoptera (Bell et al, 1999) and Diptera (Yeasmin et al, 2001). Little is known

about the mode of insecticidal action of plant lectins. In animal systems, their detrimental effects are attributed mainly to binding of the lectin to the surface of the intestinal epithelial cells. Such binding appears to lead to interference with the digestive, protective or secretary functions of the intestine and eventually the target animals die. The biochemical satiety of lectins as well as their ability to bind specific carbohydrates seems to be prerequisites for their effective function.
The aforementioned scientific information strongly indicate that plant lectins can be used as potential biocides, particularly as eco-friendly insecticides. But, it is pertinent to note that the efficacy of plant lectins against mosquito has not been investigated so far.
• Description of the plant material used in the present invention
We have carried our extensive research work on various plants which has resulted in our finding that the plant Sapindus emarginatus {belonging to the Family : Sapindaceae) contains Lectins which can be isolated and can be used for controlling Mosquitoes . This is the first time that the presence of Lectins in the plant Sapindus emarginatus is being reported.
• The Common names of this fruit are the following

The plant Sapindus emarginatus belonging to the Family : Sapindaceae is a common tree, often of large size, chiefly known for its fruits which are in universal use as a substitute for soap. Bark gray with rough scales, wood yellow, hard, but little used. This plant is widely distributed in North India, Deccan and Carnatic, extending to the East slopes of the Nilgiris and Palani and the Hills of Tirunelveh, in Tamil Nadu , in deciduous and dry evergreen forests, frequent on the coast as at Sriharikota in Nellore (Gabble, 1967).

In Thailand traditional medicine, the pericarps (fruit part) of Sapindus emarginatus is used as an antipruritic as well as natural surfactant (Kanchanapoom et al. (2001). In a preliminary study, antifertility effect of the periapt has also been reported by Ahamed & Garg (1998). Kanchanapoom et al. (2001) have isolated three new acetylated triterpene saponins together with hederagenin and five known triterpene saponins, as well as one known sweet acyclic sesquiterpene glycoside, mukurozioside II b. The biological activities of these compounds have not been determined.
The main objective of the present invention is to provide a novel extract of the plant Sapindus emarginatus belonging to the Family : Sapindaceae which is useful for contouring mosquitoes
Another objective of the present invention is to provide a process for the preparation of novel extract of the plant Sapindus emarginatus belonging to the Family : Sapindaceae which is useful for controlling mosquitoes
In the present invention Kernels of the seeds of the dried fruit of the plant Sapindus emarginatus belonging to the Family: Sapindaceae are used .
Accordingly the present invention provides a novel extract of the plant Sapindus emarginatus belonging to the Family Sapindaceae containing Lectins having anti-mosquito activity especially against Aedes Egypt.
According to another feature of the present invention, there is provided a process for the preparation of novel extract of the plant Sapindus emarginatus belonging to the Family : Sapindaceae) containing lectins having anti-mosquito activity especially against Aedes aegypti, which comprises
(i) drying the soap nut fruits of the plant Sapindus emarginatus and removing the seed
kernels and powdering, (ii) adding physiological saunas (0.9% NaCl solution) or distilled water and mixing thoroughly (iii) Keeping the resulting mixture over night at a temperature in the range of 10 to 15°C,
preferably at 10°C

(iv) Stirring the mixture using a magnetic stirrer for a period in the range of 2 to 5hrs,
preferably for 3 hours.. (v) Ultrasonic ting the resulting slurry (vi) Centrifuging the extract (vii) Filtering the resulting clear supernatant and
(viii) Storing the extract
The Stirring in step (iv ) may be effected preferably using a magnetic stirrer for a period preferably for 3 hours at room temperature (26±2’C). The resulting slurry is ultrasonicated, preferably at 20 kHz/ 100 watts; 4 x 30 sec, 2°C in Labsonic 2000 ultrasonicator. The kernel extract may be centrifuged at 8000 x g, 4*'C for a period of 30 minutes. The resulting clear supernatant may be filtered using Whatman No.l filter paper and the filtrate stored at 10°C until use. From this stock filtrate solution, different desired concentrations of the extract were prepared using unchlorinated tap water for testing its anti-mosquito activity.
In a preferred embodiment of the invention the dried soap nut kernel may be soaked in physiological saline or double distilled water
The swollen and soft kernel is ground well in a mixer grinder and stirred well on a magnetic stirrer for a period preferably 3 hours at room temperature .The ultrasonication may be effected using 20 kHz/ 100 watts; 4 x 30 sec, 2°C in Labsonic 2000 ultrasonicator . The kernel extract is centrifuged preferably using 8000 x g, 4°C for a period of 30 minutes. The resulting clear supernatant is filtered using Whatman No.l filter paper and stored at 10°C until use. From this stock filtrate solution, different desired concentrations of the extract were prepared using unchlorinated tap water for testing its anti-mosquito activity.
According to yet another feature of the present invention, there is provided a process for the preparation of novel extract of the plant Sapindus emarginatus belonging to the Family : Sapindaceae) containing lectins having anti-mosquito activity especially sigednsi Aedes aegypti, which comprises
(i) dissolving the soap nut kernel powder in a solvent selected from petroleum ether, chloroform, methanol or acetone and mixing it well

(ii) incubating the solution for a period in the range of 12 to 24 hrs, preferably for 16 hours at a temperature 26±2’C.
(iii) Decanting the resulting supernatant (Supernatant I) completely
(iv) Adding the same solvents used in step (i) to the resulting sedimented residue
(v) Processing the undissolved material separately as in step (ix)
(vi) Mixing the supernatant obtained in step (iv) with supernatant (I) obtained in step (iii)
(vii) Filtering the supernatant to remove suspended particles,
(viii) air-drying, dissolving the dry residue in acetone and filtering the acetone fraction
(ix) air- drying the undissolved material recovered in step (iv)
(x) adding physiological saline (0.9% NaCl solution) and mixing thoroughly
(xi) Keeping the resulting mixture over night at a temperature in the range of 10 to 15*’C, preferably at
lO’C. (xii) Stirring the mixture using a magnetic stirrer for a period in the range of 2 to 5 hrs with a preferably
for 3 hours (xiii) Ultrasonicating the resulting slurry (xiv) Centrifuging the extract (xv) Filtering the resulting clear supernatant and (xvi) Storing the extract at 10°C
The soap nut kernel powder is added to one of the four organic solvents selected from petroleum ether, chloroform, methanol or acetone and mixed well. Incubation may be done for 16 hours at 26±2*’C. The resulting supernatant (Supernatant I) is completely decanted. Then the same solvent is added to the sedimented residue and the extraction is performed for the second time following the procedure described above. The supernatant obtained after second extraction is pooled with supernatant I, filtered using prefereably Whatman No.l filter paper to remove suspended particles, and the filtrate transferred to glass beaker. After air-drying, the weight of the residue obtained after extraction with each solvent is recorded, and then finally dissolved in acetone. This acetone fraction was filtered, as described above, air-dried to determine its dry weight of the compounds extracted. From

• Test / assay condition adopted to confirm the activity of the extract of the present invention
Determination of the anti-mosquito activity of the extract of the invention
The anti-mosquito activity of the extracts prepared by the process of the present invention was comparatively analysed by bioassays performed using n instars larvae of Aedes aegypti, and the experimental procedures followed in this study are given below :
Bioassay
The field collected eggs oi Aedes aegypti were maintained at 26±2*’C with a photoperiod of 12h light and 12 h dark and 80±10% relative humidity. The larvae hatched from the eggs while maintaining in tap water were fed with powdered dog biscuits and yeast at 3:1 ratio, and II instar larvae were used for the study. The mortality of II instar larvae in the presence of known concentrations of different preparations of 5. emarginatus seed kernel extract was determined.
Batches of 10 larvae were introduced into the medium (50 ml of unchlorinated tap water) containing a particular concentration of each preparation of extract (Table 1). Controls received appropriate volume of 0.9% saline, distilled water or acetone in 50 ml of unchlorinated tap water. Mortality of the larvae was recorded after 48 hrs exposure. Larvae were considered to be dead if appendages did not move when prodded with a wooden dowel. No mortality was observed in any control groups. The experiment was performed with four replicates at a time for each concentration of particular preparation of the extract of the present invention. Each experiment was conducted for three times under the identical laboratory conditions. Percentage of mortality was calculated using the formula given below :


the filtrate of acetone fraction, different desired concentrations of this extract were prepared using unchlorinated tap water for testing its anti-mosquito activity.
The undissolved residue obtained as sediment after extraction with each organic solvent was air dried and its dry weight was recorded. Each residue was suspended in 30-35 ml of physiological sahne, mixed well and left over night at lO’'C. Then the sample was stirred well on a magnetic stirrer for 3 hours at room temperature and ultrasonicated (20 kHz/ 100 watts; 4 x 30 sec, l’'C) in Labsonic 2000 Ultrasonicator (B.Braun, Germany). The resulting slurry was centrifuged (8000 x g, 4''C for 30 minutes), the clear supernatant was filtered using Whatman No.l filter paper and the filtrate stored at lO’C until use. From this stock filtrate solution, different desired concentrations of the extract were prepared using unchlorinated tap water for testing its anti-mosquito activity.
• Biomedical importance of this mosquito species :
Aedes aegypti is one of the most important mosquito vector for lethal human diseases, namely, dengue and dengue haemarrhagic fever. Over 80 million people per year worldwide and about 10,000 people per year in India have been reported to suffer from such diseases transmitted by the mosquito Aedes aegypti (WHO, 2(K)0). It is found in the major urban and rural areas of India. This mosquito has been often found to breed profusely in rain water collections in discarded tyros, tins etc., and in water storage containers like cisterns, barrels and pots. They have also been found to breed occasionally in wells. Dengue outbreaks are often associated with irregular potable water supply, growing urban population, availabiUty of numerous breeding sites, rapid transportation involving the movement of both infected people and infected mosquito vectors.

The results of the susceptibihty of second instar mosquito larvae to different preparations of extracts from soap nut kernel S.emarginatus are presented in Table 1. The relative efficacy of each preparation of the extracts was assessed based on its performance to produce 100% larval mortality at its tested concentration. Out of twelve different extract preparations tested in bioassays, nine extracts killed all the larvae (100% mortality) within 48h exposure period at their tested concentrations. These effective extracts include two aqueous preparations from dry powder, all the four organic solvent extracts, and three saline extracts from organic solvent (petroleum ether, chloroform or acetone) undissolved residues, and their effective concentrations ranged from 2.5 to 6 mg dry weight / ml.
Two types of extracts from each method of extraction effective at relatively low concentrations were chosen for the standardization of bioassay conditions (Table 2).
Standardization of bioassav conditions
In order to standardize bioassay condition for conducting toxicity experiment, two different assay conditions were employed to determine toxicity of the six empirically chosen extracts of seed kernel of soap nut Sapindus emarginatus (Table 2).
Bioassay procedure
The mortality of II instar larvae of A. aegypti was determined in the presence of known concentrations of six different preparations of extract from seed kernel of S. emarginatus. The second instar larvae were maintained in two different volume of unchlorinated tap water, and the experiment was performed with four reeducates at a time for each concentration of particular preparation of the extract, as described below;
(i) Larger volume (50 ml)
Batches of 10 larvae were introduced into the medium (50 ml tap water) containing a particular concentration of each preparation of the extract (Table 2). Controls received appropriate volume of 0.9% sauna, distilled water, or acetone in 50 ml of tap water.

(ii) Smaller volume (3 ml)
Two larvae were introduced into the assay medium (3 ml tap water) containing a particular concentration of each preparation of the extract (Table 2). Controls received appropriate volume of 0.9% saline, distilled water, or acetone in 3 ml of tap water.
In both assay conditions. mortuary of the larvae was recorded after 48h exposure. Larvae were considered to be dead if appendages did not move when prodded with a wooden dowel. No mortuary was observed in any control groups. Percentage of mortality was calculated using the formula given below.

Based on the efficacy of six extracts tested (lowest effective concentration versus 100% larval mortality), three extracts were selected for further experimental study. These include physiological saline extract, acetone fraction of methanol extract and methanol insoluble residue re-extracted with physiological saline. These empirically selected three different preparations of the extracts were tested for their relative toxicity on various developmental stages of mosquito, namely, egg hatchability, survival of larvae and pupae of A. aegypti.
Determination of the anti-mosquito activity of three empirically selected preparations of soap nut kernel extract
The anti-mosquito activity of three selected preparations of the extract (as given above in detail) was comparatively analysed by bioassays performed using eggs, larvae of various instars and pupae of Aedes aegypti, and the experimental procedures followed in this study are given below :
A. Assay of homicidal effect
Effect of each one of the three selected preparations of extract of the present invention on the hatchability of A. aegypti eggs was determined as explained below. Bioassays were performed in four replicates at a time by placing 10 mosquito eggs in 50 ml of unchlorinated tap water containing a specific concentration of each extract preparation. Controls received appropriate volume of 0.9% saline or acetone in 50 ml of tap water. All containers were maintained at room temperature (26±2’C) with naturally prevailing photoperiod in the laboratory. The hatchability of the eggs was recorded periodically at 24 h intervals until 96 h from the initial time of the experiment. This time point was fixed since the completion of embryogenesis occurs within 96 h (Judson & Gujarat, 1967). The test medium was replaced after 48 h with fresh medium containing the same extract at the testing concentration. All the test media were carefully examined once in 24 h for the number of hatching eggs which was determined based on the appearance of I instar larvae, and this indicated the successful egg hatchability. Percentage egg hatchability was calculated using the formula given below :


B. Assay of parricidal and peptide effects
Bio-assays were carried out to test the efficacy of three selected preparations of the extracts used in the present invention on various developmental stages of mosquito larvae (instars I, II, in and IV) and pupae of A. aegypti. Batches of 10 larvae of each instar or pupae were introduced into test medium (50 ml unchlorinated tap water) containing a specific concentration of each extract preparation. Controls received appropriate volume of 0.9% saline or acetone in 50 ml of unchlorinated tap water. Mortality of the test organisms was recorded after 24 hrs and 48 hrs exposure. Larvae were considered to be dead if appendages did not move when prodded with a wooden dowel. Experiment was performed with four replicates at a time for each concentration of a particular preparation of the extract. Each experiment was conducted for three times under the identical laboratory conditions. Percentage of mortality of larvae or pupae was calculated using the formula given below.

From these findings, it is inferred that the toxic components from the different preparations of the extract of the present invention would have diffused into the eggs and affected vital physiological and biochemical processes associated with the development of the mosquito embryos inside the eggs. Such deleterious effects could have in turn inhibited the hatching of the larvae from the hard-shelled eggs. It is also envisaged that the extract components could have similarly produced lethal effects on the various stages of mosquito larvae and pupae.
CONCLUSIONS
The above investigations clearly demonstrated the toxic nature of the seed kernel of soap nut of S. emarginatus against various developmental stages of mosquito A. aegypti. All the three different extract preparations of soap nut, namely, physiological saline extract, acetone fraction of methanol extract and methanol insoluble residue re-extracted with saline exhibited strong homicidal, larvicidal and suicidal activity against the mosquito A, aegypti at various minimal effective concentrations.
It is, therefore, notable that the toxic component of the seed kernel extract was extractable with both polar and non-polar solvents. Thus these findings reveal that the active principle responsible for the anti-mosquito activity is not solely associated with a singular type of biochemical component of the soap nut seed kernel.

Overall, the findings of the present investigation unambiguously demonstrated that the kernel extract of soap nut possesses remarkable toxic activity against medically important vector mosquito namely A. aegypti. Further work is in progress to isolate the active agent(s) responsible for the activity from the extract and to evaluate its/ their activity
• Advantages of the invention
1. The extract of the present invention is useful for the control of vector mosquitoes especially Aedes aegypti.
2. The lectin which is the active agent which is being isolated is expected to be active at concentrations at least 10 times lesser than that of the crude extract recorded.
3. Since the active principle is ex tractable using water - based medium it would be advantageous for its application in the field conditions in an eco-friendly manner.
4. The plant material used is easily available throughout the year in the market and inexpensive (1 kg soap nut costs Rs.30/=).

We Claim
1. A novel extract of the plant Sapindus emarginatus belonging to the Family Sapindaceae containing
Lectins having anti-mosquito activity especially against Aides aegypti.
2. A process for the preparation of novel extract of the plant Sapindus emarginatus belonging to the
Family : Sapindaceae) containing lectins having anti-mosquito activity especially against Aedes aegypti, which comprises
(i) drying the soap nut fruits of the plant Sapindus emarginatus and removing the seed kernels and powdering,
(ii) adding physiological sauna (0.9% NaCl solution) or distilled water and mixing thoroughly
(iii) Keeping the resulting mixture over night at a temperature in the range of 10 to IS'‘C, preferably at 10°C.
(iv) Stirring the mixture using a magnetic stirrer for a period in the range of 2 to 5hrs, preferably for 3 hours..
(v) Ultrasonic ting the resulting slurry
(vi) Centrifuging the extract
(vii) Filtering the resulting clear supernatant and
(viii) Storing the extract at
3. A process as claimed in claim 2 wherein the dried soap nut kernel is soaked in physiological saline or double distilled water
4. A process as claimed in claims 2 & 3 wherein the Stirring in step (iv ) is effected preferably using a
magnetic stirrer for a period preferably for 3 hours at room temperature (26±2

5. A process as claimed in claims 2 to 4 wherein the resulting slurry is ultrasonicated , preferably at 20
kHz/ 100 watts; 4 x 30 sec, 2'‘C) in Labsonic 2000 ultrasonicator and the kernel extract is centrifuged taboo x g, 4°C for a period of 30 minutes..
6. A process as claimed in claim 5 wherein the ultrasonication is effected using 20 kHz/ 100 watts; 4 x
30 sec, 2°C) in Labsonic 2000 ultrasonicator and centrifuged preferably using 8000 x g, 4’C for a period of 30 minutes.
7. A process for the preparation of novel extract of the plant Sapindus emarginatus belonging to the
Family : Sapindaceae) containing lectins having anti-mosquito activity especially against
Aedes aegypti, which comprises
(i) dissolving the soap nut kernel powder in a solvent selected from petroleum ether, chloroform, methanol or acetone and mixing it well
(ii) incubating the solution for a period in the range of 12 to 24 hrs , preferably for 16 hours at a temperature 26±2''C.
(iii) Decanting the resulting supernatant (Supernatant I) completely
(iv) Adding the same solvents used in step (i) to the resulting sedimented residue
(v) Processing the undissolved material separately as in step (ix)
(vi) Mixing the supernatant obtained in step (iv) with supernatant (I) obtained in step (iii)
(vii) Filtering the supernatant to remove suspended particles,
(viii) air-drying, dissolving the dry residue in acetone and filtering the acetone fraction
(ix) air- drying the undissolved material recovered in step (iv)
(x) adding physiological saline (0.9% NaCl solution) and mixing thoroughly
(xi) Keeping the resulting mixture over night at a temperature in the range of 10 to 15°C, preferably at lOX.
(xii) Stilting the mixture using a magnetic stirrer for a period in the range of 2 to 5 hrs with a preferably for 3 hours
(xiii) Ultrasonicating the resulting slurry

(xiv) Centrifuging the extract
(xv) Filtering the resulting clear supernatant and
(xvi) Storing the extract at lOO’C
8. A process as claimed in claim 7 wherein the Incubation is done for 16 hours at 26±2*'C.
9. A novel extract of the plant Sapindus emarginatus belonging to the Family : Sapindaceae containing
Lectins having anti-mosquito activity especially against Aedes aegypti substantially as herein described
10. A process for the preparation of a novel extract of the plant Sapindus emarginatus belonging to the
Family : Sapindaceae containing Lectins having anti-mosquito activity especially against
Aedes aegypti substantially as herein described