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Biomass Research Station |
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Introduction
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Azadirachta indica A. Juss. (Neem) of family Meliaceae is native to the Indian
subcontinent. Neem, today, is grown in many Asian countries and in tropical regions
of the western hemisphere. Leaves and seeds of this tree have traditionally been
used for centuries for treatment of human ailments and control of pests. Azadirachtin
(C35H44O16), a tetranortriterpenoid from neem kernel
has been rated as the most potent naturally occurring insect feeding deterrent and
has generated wide academic and industrial interests. Azadirachtins function as
natural insect control agents because of their antifeedant as well as insect growth
regulatory properties. Several active metabolites in various plant parts of neem
bestow it unique properties as health protective, promotive, curative properties
apart from it being a pesticide par excellence, universally accepted biofertilizers
and a major constituent of village and household dispensaries3. |
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Figure 1: Neem tree with high seed output
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Background
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NBRI was involved in neem research since early 1970s when Dr. Mitra wrote perhaps
the first book on Neem detailing various chemical constituents, several of which
were unknown at that time. However, neem was not further explored in the following
years. Neem plantation trials were laid at Biomass Research Center of the institute
in 1988 but the investigations were not intensive. Dr. H. M. Behl proposed a national
program on neem in mid 1990s with the encouragement of (late) Professor S. K. Sinha
the then Chairman of the Research Council of Institute could not proceed due to
lack of sufficient funding. In 1999 National network programme involving 11 institutions
in country was launched in India with NBRI as the nodal Institution and Dr. H. M.
Behl as the coordinator of the same. Further development of technologies and products
was initiated in 2000, based on research findings under this programme. Currently,
NBRI maintains a large number of accessioned germplasm with reference material.
Also products and technologies have been developed for commercial exploitation.
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R&D Program and major achievements
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Neem exhibits a large variability as s it grows in a variety of habitats and in
almost all states of the country. There is often a mention in literature about variations
in its major metabolites such as azadirachtin content in neem from different countries
or different regions within a country. Most of the studies using chemical markers
and molecular biology tools had been contradictory.
The Biomass Biology group undertook extensive surveys from throughout the country,
more extensively from Bihar and Uttar Pradesh and screened the germplasm for phenological
and phytochemical traits. Other colleagues (Dr. Vijay Laxmi Goel and Dr. Nandita
Singh) provided valuable inputs in collection, accessioning and evaluation of the
germplasm)1.
Various provenances of neem were screened for oil percentage, azadirachtin A and
B, salanin and nimbin contents. The metabolites were extracted using conventional
methods and analyzed by High Pressure Liquid Chromatography (HPLC). Earlier Sidhu
and Behl (1996) have developed an effective HPLC protocol for detection of azadirachtin
A, B and F5. Nimbin and salanin were quantified using RP-analytical HPLC.
A large variability was observed in azadirachtin content (Figure 2) Azadirachtin
A varied from 556.9 to 3030.8 mg kg-1 with an average of 1327.4 mg kg-1
amongst various provenances. Certain individual trees (IC268646) within a provenance
had only negligible amount of (68.5 mg kg-1) azadirachtin A. Even within
a provenance there was a great variability in azadirachtin A content. No correlation
between azadirachtin A and B could be established6.
It was observed that ratio of A to B may be from as low as 1.12 to a high of 98.1.
The studies revealed that azadirachtin A is usually 13 to 16 times higher than azadirachtin
B but wide individual differences were observed among natural population.
Average of maximum and minimum temperatures, relative humidity and rainfall at different
provenances were monitored throughout the year during the study.
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Figure 2: Variability in azadirachtin A and B
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Low oil percentages were observed from regions under stress of aridity, salinity
or alkalinity. Jodhpur, Jaisalmer, Ajmer in the dry zone with low humidity and high
summer temperature, and Kanyakumari with a stress of salinity had less than 35 %
oil. These areas also had a low ratio of azadirachtin A to B. Jaisalmer, for example
is an extreme arid site. It had 34.5 % oil and A:B ratio was 2.56. Kanyakumari,
located at the southern tip of landmass, was in the similar range. These are only
trends since there were exceptions and no significant correlation between oil and
azadirachtin content was observed. Kanyakumari that had least variability in temperature
and humidity throughout the year also had least variability in azadirachtin A, azadirachtin
B content and their ratios. No such conclusion could be drawn at other areas where
there is a large variability in daily and seasonal climatic conditions.
Elite individuals with exceptionally higher salanin and nimbin content were identified
(Figure 3). There were wide variations in seed area, oil content, nimbin and salanin
concentrations among different provenances and also between individual trees of
a particular provenance7. Nimbin concentration ranged from 18.2 to 636.8
mg kg-1 whereas salanin was in the range of 45.4 to 1830.3 mg kg-1
kernel weight. Analysis of variance for seed area, oil content, nimbin and salanin
among various neem provenances showed significant differences in oil content, nimbin
and salanin. Correlation coefficients among the parameters investigated showed nimbin
and salanin as significantly correlated. Nimbin and salanin content also varied
between individual trees of a particular provenance.
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Figure 3: Variability in salanin and nimbin content
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The study shows that nimbin or salanin synthesis did not appear to be influenced
by environmental conditions as individual trees from the same agro climatic zone
showed different trends. It can be concluded that there are individual genetic differences
among neem trees.
Hasty conclusions of proposing a particular country or provenance yielding high
azadirachtin may be risky. Findings based on small sample size or conclusions based
on a multitude of factors can be misleading. With nearly 70 countries in the world
interested to raise neem plantations, it is crucial that issues regarding variability
in neem are understood. It can be concluded from present study that there are individual
genetic differences among neem trees and synthesis of azadirachtin is not dependent
upon temperature, humidity or rainfall.
Genetic variability for azadirachtin (Tables 1, 2) may affect the final products.
Large variability provides ample opportunity for selection and further improvement.
It may not be feasible at this stage to select a particular individual with desired
traits as no selections have been made in neem so far. A systematic study for tree
improvement with a population of mother trees with desired traits should be undertaken
by laying half-sib progeny trials and further selections by clonal propagations2.
The role of genetic make up needs further research. |
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Table 1: Seed area, oil content and azadirachtins (A & B) among neem accessions
from different agro climatic zones of India
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Provenance |
Seed Area (mm2) |
Aza (A) mg/kg Kernel wt. |
Aza (B) mg/kg Kernel wt. |
Total A+B |
A:B ratio |
Oil (%) |
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1 |
Orai1
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83.5 |
556.9 |
225.7 |
782.6 |
2.47 |
44.7 |
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2 |
Chapra2
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98.1 |
678.6 |
111.5 |
790.2 |
6.13 |
39.7 |
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3 |
Auraiya1
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83.7 |
804.2 |
68.1 |
872.3 |
33.07 |
37.1 |
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4 |
Kanyakumari3
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88.9 |
829.9 |
379.4 |
1209.3 |
2.95 |
35.0 |
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5 |
Bijnor1
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73.4 |
834.8 |
177.1 |
1011.9 |
4.71 |
46.2 |
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6 |
Kannauj1
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102.1 |
880.2 |
312.3 |
1192.5 |
3.32 |
36.4 |
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7 |
Rampur 1
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72.3 |
929.7 |
88.9 |
1018.7 |
10.48 |
37.3 |
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8 |
Gazipur1
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98.8 |
942.6 |
66.2 |
1008.9 |
14.31 |
42.7 |
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9 |
Ajmer4
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86.3 |
955.5 |
316.2 |
1271.7 |
3.02 |
33.2 |
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10 |
Bhagalpur2
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100.3 |
980.6 |
101.9 |
1082.5 |
9.62 |
43.6 |
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11 |
Barauni2
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90.9 |
982.7 |
147.4 |
1130.1 |
7.25 |
41.8 |
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12 |
Darbhanga2
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89.9 |
1002.5 |
119.7 |
1122.1 |
8.42 |
45.5 |
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13 |
Jodhpur4
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68.8 |
1003.1 |
103.2 |
1106.3 |
9.71 |
32.9 |
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14 |
Moradabad1
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101.3 |
1013.3 |
214.7 |
1228.1 |
6.41 |
50.1 |
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15 |
Farrukhabad1
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112.3 |
1022.5 |
94.0 |
1116.5 |
10.87 |
42.5 |
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16 |
Mathura1
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90.5 |
1071.8 |
58.7 |
1130.5 |
31.41 |
45.3 |
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17 |
Kheri1
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91.6 |
1082.6 |
173.2 |
1255.7 |
5.88 |
43.1 |
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18 |
Jaisalmer4
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75.2 |
1092.2 |
426.2 |
1518.4 |
2.56 |
34.5 |
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19 |
Hathras1
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82.4 |
1126.9 |
134.4 |
1261.2 |
8.40 |
42.4 |
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20 |
Ranchi7
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92.5 |
1159.8 |
190.1 |
1349.9 |
6.09 |
41.2 |
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21 |
Muzzafarpur2
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90.1 |
1169.4 |
141.5 |
1310.9 |
8.26 |
41.6 |
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22 |
Bulandshahar1
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97.0 |
1205.1 |
139.4 |
1344.5 |
9.79 |
49.7 |
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23 |
Haridwar5
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83.3 |
1231.0 |
182.8 |
1413.8 |
7.17 |
46.1 |
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24 |
Hazipur2
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101.1 |
1254.4 |
96.6 |
1351.0 |
13.04 |
42.9 |
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25 |
Patna2
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86.2 |
1272.6 |
228.3 |
1500.9 |
5.58 |
41.1 |
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26 |
Lucknow1
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105.2 |
1334.2 |
139.4 |
1473.6 |
9.66 |
45.2 |
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27 |
Badaun1
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97.8 |
1388.1 |
143.9 |
1532.0 |
9.64 |
35.2 |
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28 |
Dehradun5
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89.3 |
1459.7 |
326.6 |
1786.3 |
4.72 |
45.3 |
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29 |
JP Nagar1 |
74.9 |
1505.2 |
590.6 |
2095.9 |
6.03 |
37.1 |
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30 |
Varanasi1
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104.8 |
1531.7 |
69.7 |
1601.5 |
24.54 |
48.3 |
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31 |
Muzzafarnagar1
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88.8 |
1534.5 |
149.7 |
1684.2 |
10.27 |
40.1 |
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32 |
Meerut1
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101.2 |
1573.8 |
105.2 |
1679.0 |
22.63 |
40.8 |
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33 |
Agra1
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76.2 |
1576.2 |
261.9 |
1838.0 |
8.16 |
43.9 |
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34 |
Siwan2
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81.1 |
1629.0 |
96.0 |
1725.0 |
17.0 |
42.8 |
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35 |
Kanpur1
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80.5 |
1671.4 |
68.9 |
1740.4 |
23.14 |
40.8 |
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36 |
Pratapgarh1
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78.9 |
1705.9 |
100.8 |
1806.7 |
17.12 |
38.6 |
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37 |
Jaipur4
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92.6 |
1706.4 |
251.5 |
1957.9 |
6.78 |
33.5 |
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38 |
Samastipur2
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84.5 |
1916.5 |
95.0 |
2011.5 |
20.19 |
45.0 |
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39 |
Fatehpur1
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97.0 |
1941.2 |
43.1 |
1984.3 |
98.15 |
51.6 |
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40 |
Rohtak6 |
86.6 |
2032.9 |
243.9 |
2276.8 |
8.33 |
37.9 |
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41 |
Mau1
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92.6 |
2058.2 |
100.3 |
2158.5 |
20.53 |
43.2 |
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42 |
Sri Ganganagar4
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89.0 |
2400.0 |
172.3 |
2572.3 |
13.92 |
37.3 |
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43 |
Shahjahanpur1
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110.9 |
3030.8 |
134.1 |
3165.0 |
24.32 |
40.6 |
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CD at 1% |
61.10 |
1023.04 |
326.10 |
1126.3 |
33.87 |
4.45 |
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CD at 5% |
45.94 |
769.20 |
245.19 |
846.8 |
25.47 |
3.34 |
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1 = Uttar Pradesh; 2 = Bihar; 3 = Tamil Nadu; 4= Rajasthan; 5 = Uttranchal; 6 =
Haryana; 7 = Jharkhand. |
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Table 2: Analysis of variance between seed area, oil content, azadirachtin A, B;
A+B and A:B ratio among different provenances |
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Source |
Degree of Freedom |
SS |
MS |
F value |
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Seed area (mm2) |
42 |
11446.1 |
317.9 |
0.4 NS |
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Oil (%) |
42 |
1393.97 |
38.7 |
9.2** |
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Azadirachtin A |
42 |
2.31 x 107 |
642225.1 |
2.9* |
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Azadirachtin B |
42 |
1168336 |
32453.7 |
1.4 NS |
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Total Azadirachtin (A+B) |
42 |
2.32 x 107 |
647116.4 |
2.4* |
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A:B ratio |
42 |
28426.2 |
789.6 |
3.2* |
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Results are of two-way ANOVA (seed parameters, oil content, azadirachtin A, azadirachtin
B, total azadirachtin (A+B), A/B ratio, and various provenances); total=36; * =P<0.01;
** =P<0.001; NS =not significant. |
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Dental Cure Recipe from Neem |
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The authors developed a unique recipe for dental cure. It filed a patent (Non toxic
Dental care herbal formulation for preventing dental plaque and gingivitis Patent
Application No. 569 NF 2002)4.
Neem extracts reduce gingival bleeding and inflammation. These are also useful against
plaque formation. Neem extracts and the prouct developed with synergistic effect
from the herbs eliminated anaerobic micro-organisms (Peptococcus, Fusobacterium
and Bacteroides) and aerobic species (Streptococcus viridans, Bacilli,
E. coli and Streptococcus epidermidis) within a few days after application.
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Skin care, pet care and other products |
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The group has developed several products like skin care, pet care, pesticidal formulations,
fertilizers etc. from various parts of neem involving state of art technologies.
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International Conferences
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Dr. H M Behl was the Chief Editor of Proceedings of the World Neem Conference (Neem
99) held at Vancouver, British Columbia, Canada. The proceedings were published
by the Neem Foundation, India. He was also the co-editor of proceedings entitled
“Collection, Processing and Commercial Utilization of Neem”, published by the the
Ministry of Industries, Govt. of India in 2000.
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Acknowledgments
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The study was a part of National Neem Network activities. The network was sponsored
by the NOVOD Board, Ministry of Agriculture, Govt. of India.
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Literature cited
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- Behl, H.M. (2000). Collection, Processing and Commercial
Utilization of Neem. Published by the Fragrance & Flavour Development Centre,
Ministry of Industries, Govt. of India, 2000; Co-editor: A. Lahri.
- Behl, H.M. (2000). Technical manual for High Tech Protected
Environment Nursery Cultivation, National Botanical Research Institute publication,
Lucknow, India.
- Behl, H.M. (2002). Proc. World Neem Conference (Neem 99),
Vancouver, British Columbia, Canada, Published by Neem Foundation, India.
- Behl, H.M., Saimbi, C.S., Srivastava, N. and Kant, S. (2000).
In: Collection, Processing and Commercial Utilization of Neem (Eds. A. Lahri, H.
M. Behl), Fragrance & Flavour Development Centre, Govt. of India, pp. 239-242.
- Sidhu, O.P. and Behl, H.M. (1996). Current Sci.
70(12): 1084-1086.
- Sidhu O.P., Kumar, V. and Behl, H.M. (2003). Industrial
Crops & Products, 2003.
- Sidhu O.P., Kumar, V. and Behl, H.M. (2003). J. Agr. Food
Chemistry, 51(4):910-915.
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