Neem Leaf BioMass And Leaf Extracts

Neem leaves have been known for centuries to contain compounds that are repellent to insects, thus serving to protect neem foliage from insect feeding, but they also contain anti-fungal compounds which can be evidenced by either direct incorporation of the leaves into soil as organic amendments (green manure) or by using extracts of them. The concentration of bioactive compounds in the leaves is probably lower than in the fruits, but still of interest for agricultural purposes.

Neem Leaf Biomass

It has been demonstrated that the direct incorporation of either fresh or dried leaves as a green manure can result in the population decline of several soilborne pathogens. A significant reduction in will incidence, caused by Phytophthora Capsici, was achieved in belelvine (Piper betle) when dried neem leaves were applied near the base of the vine. Population of Pythium Aphanidermatum were reduced following incorporation of green neem leaves (5% w/w) into infested soil (SINGH and PANDEY, 1966). Similarly, the application of chopped neem leaf amendment reduced the total fungal population in tomato rhizosphere soil. However, neem as a green manure prolonged the vialbility of sclerotia of Curriculum Sasazaki over controls while increasing the total microbial population in the trested Soil.

Seed Cake

In greenhouse experiments NAIDU et al. (1980) treated rice clip-inoculated with a virullent strain of Xanthomonas Oryzae with uncoated urea, coal-tar-coated urea, and nem cake with coal-tar-coated urea and comapred the results of the untreated control. There was no reduction in leison formation. Compared with the control, all various had higher values for leison length, the symptom parameter for X. oryzae, and higher values were obtained for higher additional doses of nitrogen and split dosages with a base dose. The highest rates were for neem cake with coal-tar-coated urea and an additional split dosage of 200kg N/ha which also led to the tallest plants to the experiment. Undoubtedly the total amount of fertilizer played an important role, as did the influence of neem cake on nitirification - its inhibition or recardation in the soil. The influence was studied in many experiments. MISRA and CHHOKAR explained the inhibition of denitrification by the inhibition of Nitrosomonas and Nitrobacter spp.

Another strategy to save nitrogen is by the use of seeds treated with Rhizobium spp Neem cake did not affect module formation, except at high doses, which had some effect on Rhizobium spp but not on grain on master yield. (The authors did not say, what the does was but it has been more than 7 ppm on the soil weight basis).

In experiments to evaluate the inhibitory effect an Fusarium Udum of soil incubated with neem cake in vitro B. Thuringiensis was frequently detected in the lylic zones of fungus growth. The microbial activity in decomposing neem cake, especially of bacteria, was described by PETHAMBARAM the increase in the population of bacteria in neem cake amended soils was mentioned by JEYARAJAN et al.

ESWARAMURTHY et al.(1993) reported reduction of symptoms in rice caused by X. Compestria pv. oryzae and a considerable increased in yield when neem cake extract was sprayed twice at a one-weel interval. Repetitions of the treatment also increased yields compared with control.

In laboratory experiments CHANDRAMOHAN and MOSES (1996) demonstrated that extracts of neem cake were effective against X. Compestria pv. cltl with extracts being better than awueous extracts.

Seed Oil

As early as 1917, the antibiotic in vivo effectiveness of margosates obtained from neem oil was proved by CHATTERJI and RAY in combating syphilis and other skin diseases. CHOPRA et al tested oil from neem seeds against a number of bacteria causing diseases in humans and animals. The oils were added directly to the substrates in vitro at different dilutions, Neem oil was very efficient against a human strain of the same bacterium showed some growth at the dilution of 22,000. P.pestis and S. aurex showed some growth already at a dilution of 5,000, while S.schotemelleri and K. pneumoniac showed normal growth at this concentration.

PATEL and TRIVEDI tested neem oil in a cup-plate method measuring the inhibition zones. Unfortunately, they did not use an antibiotic standard and compared only the inhibition of different oils with that of the emulsifier used, which have no inhibition. Neem oil showed the weakest inhibition when compared with oils of karanja, malkanguni and darudi. No inhibitions by neem oil was found against Pseudomonas Pyocyanea and Proteus Vulgaris. When compared with darudi oil, which in most cases ws the best inhibitor, neem oil gave only 63% inhibition for two strains of S. aureus genes var. aureus and M.pyogemer, B. subtilis, S. paratyphi and E. coli. Against another strain of S. aureus neem oil gave 65% inhibition; for C. diphtheriac, 67%; for S. schotmuelleri 74% and for S. typhosa, 78%. The maximum inhibition zone for darud and neem oil was 32 and 20 mm, respectively, achieved with S. aureus, the minimum was 21 and 14mm, respectively with C, dlphtheriae. However, in serial dilution tests darwal oil was less effective then neem oil in reaching the inhibiting concentration for S. aureus with 0.4% and 0.3% and for S. typhosa with 0.8% and 0.4%.

The antibacterial activity of neem oil in vitro was tested also by RAO et al. using many strains of Pseudomonas Aeruginosa(55), S.aureus, E. coli, Proteus spp, and Klebsiella Pneumoniae (?)(5). Inhibition with pure neem oil detected to the extent of 85%, 94%, 94%, 100% and 80% of the strains respectively. The effect was good for 11%, 12%, 21%, 8% and 60%, medium for 42%, 59%, 45%, 46% and 20% of the strains, and a week reaction was observed in 33%, 22%, 21%, 42% and 0% respectively. However inhibition was lost quickly, when the oil was diluted. In I/64 dilution none of the Pseudomonas Aeruginosa strains only 20% of S. aureus, E. coli ans Proteuis spp and 25% of K. aerogenes strains were still sensitive. SINGH et al. described in 1974 an antibiotical effort of neem oil on E. Coli.

Neem oil extracted with petrol either in a Soxhlet appratus showed in vitro inhibition of Bacillus pumilus, B.substiz, S. typhasa, S. paratyphi and S.aureus using filter disc test, but it is difficult to quantity of activity as a results of controls were given.

Commercial Products

Little inforamtion is available on the effect of commercial neem products on viruses. AH-MED(1988) reports that 'Wellgro' a product of ILTD is used by farmers to control TMV in the West Godavari district. The effectiveness of the treatement seemed to be as good as spraying neem leaf extracts, but no detailed data or quantities were given. Other commercial products are used in the control of vector as vector repellents, like 'RD-Repelin' which is a mixture of extracts of neem(A.Indica), karamja(Pongamia Pinnata), castor (Ricinus Communis), Mahua (Madhuca Indica) and sesame(Sesamum Indicum). In addition to a repellent effect against the pea aphid. Acyrthosiphon Pisum, which did not prevent virus transmission, a delayed symptom expression for zucchini yellow mosaic virus(ZYMV) was observed in 81% of the test plants. The mixture of ingrediants does not, however allows us to contribute this finding to a particular plant extract. No influence on virus replication by 'RD-Repelin' was found, because the ELISA-readings of untreated and treated plants did not differ significantly.

ADDING NEEM TO SOIL

The use of neem cake as an amendment to agricultural soils is a well known practice in India, mainly to exploit the fertilizer properites of this material but also in combination with synthetic chemicals for plant protection. The influence on viruses has not been studied yet in details. Some experiments with neem cake(NC), neem seed kernels(NSK) and dried neem leaves(NL), in the field and under greenhouse conditions yielded very different results, depending on the virus and host plant used. In field experiments, when neem was added to 1% of the upper 10cm layer none of the neem products used was able to prevent infection of zucchini with watermelon mosaic virus. In a second experiment under much higher infection pressure, the results in the neem plots were much better than in the control, and a delay in symptoms expression was significant for NSK powder and to some extent for NL powder. In a greenhouse experiment with artificial inoculation, in which the zucchi plants were planted in prickout pans with soil containing 1% neem, the inhibition was low but marked: only 80% of the zucchini plants treated with NS, 85% with NC and 82% with NL, but 100% of the control plants became infected, as determined by symptom expression.

When hups were used as test plants with the same method, a reduction of the ELISA reading for hop mosaic and hop latent catlaviruses was obtained in the plots treated with crushed NSK and NL, but the differences from the control were not significant. Another system used for testing this type of application was Prunus Necrotic ringspot virusand cucumber planted in 10cm pots of five plants each. In this artificial system the use of neem did not prevent virus infection after artificial inoculation, but there was a reduction in the number of plants successfully inoculated and with increasing amounts of neem added(0.5%, 1%, 2,5%) there was a significant reduction in symptom expression. This corelated with the average vales for the ELISA-readings. The efficacy increased from Soxlet extracted NC to NC to NSK, but 5% NSK induced also phytotoxic reactions, measurable as reduced growth and fresh weight.

Pure Ingrediants

VERMA(1974) demonstrated inhibition of PVX in vivro and in vivi using C, Amaranticolor as a single lesion host. When allowed to preincubatable for 15-20 min, 44% and 63% inhibition of lesion formation was obtained using nimbin and nimbidin, respectively. With pretreatment of the test plan by spraying nimbin and nimbidin even better results were obtained, when the treatment was carried out 12 h or more before in oculation. The inhibitory effect of the treatments p.i. also increased with more time after the infection process and reached 43%(nimbin) and 35%(nimbidin) when carried out 24 h p.i. Unfortunately, VERMA did not mention which of the three concentrations used in the first test used to perform the pre- and post- inculation tests. A pre inculation root treatment for 24 h did not lead a reasonable inhibition when tomatoes were used, but with C. amaranticolor, local lesion formation was reduces using a mixture of nimbidin and nimbin at 250 ppm. Higher concentrations mostly  resulted in phytotaxics reactions of both test plant species. Experiments by the author using aza resulted in high or low inhibition, but it was never significally different from with the methonal control, indicating the aza has no significant antiviral properties that the methonolic solution seems to be an inducer of resistance. Similar results were obtained with methonolic extracts of neem representing an earlier stage by the methonal control, but in some experiments higher concentrations of extracts tended to have stronger effects.

SEED OIL

Seed Oil

In field experiments conducted in Tamil Nadu (India) 1% neem oil sprays reduced significantly ring mosaic disease of peanut caused by ToSWV. The effect of oil application seemed to be twofold; the incidence of the viral vector Frankliniella Schultzei was reduced and an antiviral activity was diagonosed under laboratory conditions. Unfortunaetly the numbers did not give the details of these laboratory experiments that "antiviral activity was measured by symptoms, When Vigna Sinensis was used as the indicator plant, neem oil completly inhibited lesion formation. A reduction in urd bean leaf crinkle infection in Vigna Mungo was demonstated by SABITHA and JEYARAJAN with sprays of neem oil(1% 5%) but it did not exceed 40% and it cannot be excluded that this effect was only due to an inhibition of the virus transmission i.e the vectors. As no further details were given, it remains unclear which of the crinkle viruses of the urd bean found in India is meant the ULCV transmitted by Henosepilachna Dodecastigma the applied transmissible virus or the whistlefly-transmissited virus. All three vector groups can be affected by neem, as discussed later. In experiments with ilarviruses and cucumbers, topical application of neem oil had no effect on the number of infected plants or severity of symptoms ROYCHOUDARY and JAIN gave a diffferent interpretation for their results with mechanical inoculations of different host species with TMV, saying that virus multiplication is inhibited by neem oil.

Seed Kernel Extracts

AQUEOUS EXTRACTS

Aqueous extracts of neem seed kernels, when preincubated with TMV in laboratory and greehouse tests, gave varying results. Strong inhibition in one test was followed by increased numbers of lesions in the next. In general concentrated virus inocula led to less inhibition than more diluted inocula. Using systemise hosts like N.labocum cv.Java, a delay in symptom development was observed, along with much better growth of the neem-treated plants. Using apple mosaic ilavirus and cucumber as a model system, topical applications of aqueous extracts pre and post infection did not lead to a reduction of the symptoms.


METHONOLIC EXTRACTS

Methonolic extracts, as discussed below for Azadirachtin(a2a), bear the problem of methonal itself, either as a denaturating component or as a resistance inducer. Preinoculation applications of methonolic extracts inhibition when the results were compared with the methonal control, but the data differed widely among the various concentrations of neem and there was no correlations with the inhibition values. When the results of the pretreated halves of the leaves were comapred with the untreated ones, there were always fewer Jesions but the difference was not always significant and again there was no correlation with the neem concentration. When no methanol control was used, positive results were always reached, with the above mentioned limitations. When the extract were used only for co-inoculation, a significant reduction - with values above 90% - was obtained again when compared with pure virus control coly. Also the leaf halves inoculated with the virus alone showed nearby the same reduction as the other half with relatede virus. However, in this experiment, no menthol control was used.

BARK AND ROOT EXTRACTS

MURTY described a 87% reduction of TMV infection of N. gluntinoza using an aqueous extract of the bark of the neem tree. In experiments by the author, a methonolic extract from neem bark from Senegal resuspended with the microvolume of methonol and diluted with buffer, gave similar results when preincubated with the virus. Using the half leaf method, the number of lesions also decreased on the control-half of the leaves inoculated without extract, indicating a systemic effect when cv. Xanthi NN used. This effect was less significant using cv. White Burley as test plant. The bark extracts from different sources behaved very similarly. In an additional test, the bark from two different sources in Senegal was compared. With 250 ppm of extract, 57% and 62% inhibition respectively, was obeserved. Inner bank and wood was less effective, with only 43% inhibition. Compared with a methyl tert, buthylether extract of dried leaves from Senegal which gave 74% inhibition, all other bank and wood preparation were weaker. This activity was even weaker when Datura Stramonium was used as the test plant instead of tobacco; the inhibition reached , only 34% and 27% for the outer bank, 36% for the inner bark and wood, and 33% for the left extract. Symptom inhibition was significant only for one of the bark-samples when Nicotiana Benthamiana was used as systematic test plant. When the preincubation period was extended to 5 d, the inner bark and wood extract gave a weak inhibition of <10% and the outer bark, and the leaf extracts showed even an increase in lesions formation on the half-leaf inoculated withn the pretreated inoculum.

In general, there was a great variability in the values obtained for individual leaves when pretreating virus with extracts. Even in control experiments using the half-leaf method, differences of up to 49% could be observed when both parts were inoculated with the same pretreated inoculum, an effect never observed with the virus alone. This indicates that experiments with a low number of replications will occasionally lead to erratic results. Inhibition - with lower than with whole plants - was obtained using leaf discs and pretreated inoculum. When an inhibitor-seaked filter paper was placed on leaf discs. Again the effect could not be enchanced by using more inhibitor-discs. These experiment were performed with N. tabacum. When Nicotiana rustica was used, no inhibition was observed and with D.Stramonium only 30% inhibition was achieved under the same conditions.

Chapter 1 - Growth and yield of fruit

The neem tree generally grows rapidly under favourable conditions. The pH value and the waster table in the soil, rainfall, irrigation, nutrients, inter and intraspecific competition, mycorhiza, temperature and genetic factors all play a role. Little is knowna about the impact of the last, but environmental influences seems to play an important role in many regions of the tree's distribution range.

In very dry areas, in high altitudes at colder temperaturesand in high rainfall areas, the growth may be happened to secure extent . For instance, as already started, an attempt to grow neem in the western part of the Amazonian basin in Eastern Ecuador failed due to high humanity and precipication. In Vili Levir, neem grows well in the dry and warm western parts whereas only a few trees, with low yields exists in the welter and cooler eastern parts. At high altitudes, relatievely low temperatures may be the main reason fine growth and lower yield. Low rainfall may be another reason for slow growth. Apporximately 130m annual rainfall enables A. Indica to survive but its growth is then very slow. Drought may even lead to details of tree in plantations already 3 - 5 years after planting, due to intraspecific competitions for water, especially if spacing between trees is less than 3m. Much faster growth occurs with 8 - 1.2 grew to a height of 6 -7 m within 2 1/2 years in an area with low rain fall and on soil with high salinity. The leaves of these trees were dark green and locked very healthy. 

Chapter 1 - Ecology

The neem tree is famous for its drought resistance. Normally it grows well in areas with sub-arid to sub-humid conditions, with an annual rainfall between ca 400 and 1,200 mm. It can also grow in regions with an annual rainfall below 40mm, but in such cases it depends largely on the ground water. Under such conditions very low annual rainfall is sufficient for its survival. During severe, long-lasting droughts the leaves are shed, sometimes completely. Neem tree exist also in regions with up to ca 2,500 mm annual rainfall, provided the soil on which they grow is well drained, for instance on hills. The yield of fruits is normally low under such conditions, owing to flower- and fruit-shelding during the rainy seasons. Some attempts some years ago to grow neem in areas with high rainfall of ca. 3,000 - 4,000 mm/a, as in eastern Ecuador in the Amazonian basin and on Tonga Island, failed completely.

Neem can grow in many different type of soil, but it seems to develop best on well drained, deep sandy soils. In the Sudan and in India, it thrives on black cotton soils. In Nigeria A. indica develops well on other soils with high clay content but not with a high proportion of sand. In other parts of Africa(Benin) neem trees are found on red ferrolas and in the Caribean on calcareous soils. In soils with a high content of fine sand or silt, nutrient defeciences, lead to an obvious chlorosis of the leaves. Chlorosis is common for instance in parts of Haiti, in Westeren Senegal and West of Thies and in south western Madagascar south and north of Toliara. A indica exists on stony shallow soil, on soil with waterless subsoil and also on solis with a hard calcareous or clay pan near the surface. On very poor stands it does not grow well in groups or plantations owing to high intraspecific competiotion for water and soil nutrients. Under conditions neem plantation in Africa, have often died out after some years. FISHWICK emphasizes that soil water availability appears to the most critical factor, and this factor is combination of soil texture and local ground water supplies. On the other hand neem can also grow on alkaline or saline soil. In the Dominican republic, for instance, it grows well on former sugar cane fields which were abndoned owing to high salinity of the soil.

Neem as a typical stropical/substorpical plant, exists at annual mean temperatures between 21 and 32'c. It can tolerate high to very high temperatures, for instance northest ans central Africa where temperatures in the shade can reach ca. 50'C during the summer months. Temperatures below 4'c, and frost, are unfavourable and may result in shedding of leaves and occasionally even death of young plants. In the sub-Himalayan zone, where temperatures may fall below 0'c in winter, neem seems to be better adpted to cold conditions but young plants have to be protected by screens.

The neem tree is usually found on plains and low-lying hilly country. It thrives at altritudes upto 700-800 m and ocassionaly 1000m above the sea level. Higher altitudes are, as a rule, much less favourable.

Chapter 1 - Botanical characteristics

Neem

Azadirachta Indica is a fast growing plant that usually reaches a height of 15-20m, and under very favourable conditions upto approximately 30-35. As a rule it is an evergreen tree, but under extreme circumstances, such as extended dry periods, it may shed most or nearly all of its leaves. The branches spread widely. The fairly dense crown is roundish or oval and may reach a diameter 15-20m in old, free standing specimens.

The trunk is relatively short, stright and may reach a girth of 1.5 - 3.5 m. The bark is hard, fissured or scaly planet and whitish-gray to reddish-brown. The sap wood is graylish-white and the heart wood reddish when first exposed to the air, be coming reddish-brown after exposure. The root system consists of a strong taproot and well developed lateral roots. The lateral surface roots may reach over 18m. Vesicular-arbuscular Mycorrhiza is associated with the rootlets; HABTE et al, categorized neem as a highly VAM dependant plant species.

The unpaired, pinnate leaves are 20-40 cm long and the medium to dark green leaflets which number upto 31, are approximatelty 3-8 cm long. The terminal leaflet is often missing. The petioles are short. Very young leaves are reddish to purplish in color. The shape of mature leaflets is more or less asymmetric and their margins are denate with the exception of the base of their basiscopal half, which is normally very strongly reduced and cuneate. The leaves contain 12.40 - 18.27% crude protein, 11.40 - 23.08% crude fiber, 43.32 - 66.60% N-free extract 2.27 - 6.24 ether extract, 7.73 - 18.37% total ash, 0.89 - 3.96% calcium and 9.10 - 0.30% phophorous.

Natural hybrids between indica and siamensis, found in Thailand on places where both species grow together have an intermediate position regarding the shape and consistency of the leaflets.

The white, fragrant flowers are arranged in axillary normally more or less dropping panicles which are up to 25 cm long. The inflorescences, which branch up to third degree, bear approximately 150, and occasionally up yo 250 flowers. An individual flower is 5-6mm long and 8-11 mm wide. The five petals are 5-5.5mm long and 2mm wide. Protandric bisexual flowers and male flowers exist on the same individual . Certain solitary growing neem trees seem to be unable to self-fertilization as observed in Nicaragua. Honeybees are important pollitaris in Africa, Asia and Australia in stringless bees in tropical America. The ovary is trilcoluar. There are ten glabrous anthers which are inserted at the base of the flowers. The nectary is annular and fused at the bases of the ovary.

The glabrous fruits are olive-like drupen which vary in shape from elongate oval to nearly roundish and when ripe are 1.4-2.8 * 1.0-1.5 cm. They are green when young and yellowish - green to yellow, rarely reddish when mature. The fruit skin is thin and bitter-sweep pulp is yellowish-white and very fibrous. The mesocarp is 0.3 - 0.5 cm thick. The white, hard 'shell' of the seed encloses one, rarely two and very rarely three eloganted seeds having a brown testa. The seeds measure 0.9-2.2 * 0.5-0.8 cm, and the 'seed kernals' are 0.8- 1.6 * 0.4-0.5 cm.

The number of chromosomes is : 2n=28, 30; n=14. The anatomy of the leaf, bark and wood as well as palynology and embryology are described in detail by TEWARI and are therefore not discussed here.

A. Indica may reach an age of more than 200 years.

Neem - Chapter I - The Tree and the Characteristics

Adrien Henri Laurent De Jussieu described in 1830 the neem tree as Azadirachta Indica. Its taxonomic position is as follows :

• Order Rutales
• Suborder Rutineae
• Family Maliaceae
• Subfamily Melioideae
• Tribe Melieae
• Genus Azadirachta
• Species A. indica

There are many Common names for the neem tree, especially in those countries where it has been establised for a long time. Some examples are listed in Table.

Arabic	Azad-darakhul-hind
Assamese	Nim
Bahasa Malaysia	Baypay, dawoon-nambu
Bali	Intaran
Bengali	Nim, Neemgachh, Neem
Burmese	Tamaka, thin, thinboro tamakha, bowtamaka, tama, tamabin
English	Neem, Indian Lilac, Margosa tree, nim, crackjack, paradise tree, white cedar, chinaberry
French	Azadirac de 1'Inde, margosier, margousier
German	Grossblaettiger zedrach, Indischer zedrach
Gujarati	Limba, Limbado, Leemgo, Danu-jhada, Kohalu limdo, Kohumba, Libado, Leemdo, Nimuri
Hindi	Nim, Nimb, Bal-nimb, neem, nind.
Kannada	Bevinamara, bevu, hebbevu, kiri-bevu, kai-bevu, kadbevina-mara, nimb, olle, val-venu, venu, vevu kol Nim
Konkani	Nim
Kumaoni	Betain
Madura	Mempheuh
Malayalam	Aryaveppu, veppu, Aryaveshnu, Rajavedhu, vepe
Marathi	Kadukhajur, limba, limb, nimbay, Bakayan, Balanti-limb, bal-nimb, limbacha-jhada, kadu-limba, nimuri.

Production Flow chart for Coco Peat

CoCo Peat Production:

Coco Peat is an Organic Natural Renewable Green Grow medium and is a sure alternative to all the peat products available in the market. It is 100% Organic product derived from Indian Coconut husks.

The selected coconut husks of best quality are crushed and busted in special machineries, where the fiber and the residual coir pith is separated by sieving.  The long and medium fibers are dried and compacted into bales or spun into ropes as per the need of the end clients.

The Coir pith, a granulated spongy product which is obtained as residue after the fiber extraction from the coconut husk is carried to yards, where they are washed to leach of the excess salt and allowed for a period of time for little composting. Then they are steam sterilized to eliminate any microbial contamination. The sterilized product is carried to huge cement or stone dry yards and allowed to dry These drying yards are fenced for any cross contamination. When the moisture reaches up to 15%, is then screened in double sieve’s where the fine dust is removed and graded as per sieve size and quality standard of the end clients usage. The graded material is conveyed to hydraulic press machines, which presses the material to form 5kg or any other required weight or sizes. The pressed blocks are packed in pallets and stuffed in containers. Quality checking is held in all the chains of production for physical and chemical properties of the product. The process involves no chemical treatment and is 100% natural organic product.

The Quality Control is done at all levels checking various Parameters like EC, Contamination of foreign materials, weeds (by weed grow trays) etc.

Flow Chart: