Studies On Agar Forming Mechanism Of Aquilaria Sinensis (Lour.) Gilg

Aquilaria sinensis （Lour.） Gilg was a paticular tree for producing agarwood used ashigh-quality incense and traditional chinese medicine in china. At abroad, there were somestudies on agar formating mechanism of Aquilaria agallocha, Aquilaria malaccensis, Aquilariacrassna, etc.. In china, studies targeted on Aquilaria sinensis （Lour.） Gilg have been carried outsince1970s. In this paper, studies on agar formation mechanism started with relation betweenAquilaria sinensis （Lour.） Gilg and the involved fungi from the standpoint of plant physiology,which including separation, purification and identification on involved fungi, fungi effect onchemical composition and chemical property of the wood tissue, physiological and metabolicchanges in the interaction between Aquilaria sinensis （Lour.） Gilg and the fungi. The mainresults about agar formation were as follows:1. After wounding Aquilaria sinensis （Lour.） Gilg on the trunk, more pathogenic fungiwould appear, including Lasiodiplodia theobromae, Fusarium solani, Curvularia spp. andPestalotiopsis Microspora, etc.. The above pathogenic fungi were not found in the healthytrunk. It indicated that these pathogenic fungi were from outside through wound, and agarformation by physical wounding was the interaction results of various pathogenic fungiinfection.2. In the studies, it was found that fungal species respectively decreased in the order：no-agar wood�'little-agar wood�'agar wood. It indicated that the formation of agar materialhad inhibiting effect on certain fungi, and then other fungi would replace them, so the agarformation process was not the result of some specific fungi infecting from first to last.Fusarium proliferatum and Hypocrea jecorina were found in the two test sample: little-agarwood, agar wood.3. After instilling with the liquid inoculum of Rhizomucor variabilis, Fusariumproliferatum and Hypocrea jecorina, fatty acid in the timber of Aquilaria sinensis （Lour.） Gilgdecreased by similar rule, early （especially in the first2months） decreasing rapidly, and then steadily decreasing. The fatty acid consisted of octadecenic-[9]-acid, octadecenic-[10,2]-acidand palmitinic acid, among which, octadecenic-[9]-acid decreased at most, followed byoctadecenic-[10,2]-acid, and palmitic acid decreased at least. So the three fungi all coulddecompose the fatty acid in the timber of Aquilaria sinensis （Lour.） Gilg, the decompositiondegree in the order：Rhizomucor variabilis�'Hypocrea jecorina�'Fusarium proliferatum.4. From the view of the content of benzyl acetone, on the6thmonth, benzyl acetonecontent of the sample instilled with liquid Fusarium proliferatum inoculum was the highest,reaching104.7μg·g^-1, followed by the sample instilled with liquid Hypocrea jecorina inoculum(33.38μg·g^-1), and the sample instilled with liquid Hypocrea jecorina inoculum was very low,only1.94μg·g^-1. It had been shown that the fungus species had great effect on benzyl alcoholcontent, namely, Rhizomucor variabilis had little effect on the formation of benzyl acetone, andFusarium proliferatum the best effect.5. In this study, the effect of liquid inoculant on benzyl acetone content was comparedwith sterile fungal culture solution. The results had shown that there was little differencebetween liquid inoculant and sterile fungal culture solution. It indicated that there was notdirect relationship between agar formation and the fungal mycelium, on the contrary, there wasclose relationship between agar formation and mycelium metabolites.6. After instilled with the culture solution of Fusarium proliferatum, on the6thmonthhot-water extractives content in the agar wood was higher than that in no-agar wood by26.923%, which indicated that the cell wall of the timber tissue had been partially damaged andthe metabolites increased; Benzene-ethanol extractives in the agar wood was higher than that inno-agar wood by184.665%, which showed that a part of carbohydrate had been degradated,terpenoid compounds, aromatic compounds content increased accordingly; Holocellulosecontent in the agar wood was lower than that in no-agar wood by21.737%, pentosan（hemicellulose） also lower by9.986%, and lignose content higher by38.316%, which indicatedthat the culture solution could degradate Holocellulose, but not degradate lignose; Comparedwith the no-agar wood, mineral elements content, except for S（increased by24.85%）, in theagar wood decreased by7.92%⁵5.96%, which showed that the timber tissue was under stress, the S content increasing may be the result of passive absorption for self-protection; In theprocess of agar formation, normal balance between growth and respiration had been striked,which resulted in starch conversing to sugar, so the soluble sugar content in the agarwoodincreased by56.10%and starch content decreased by62.92%.7. Agar material occurred mainly in parenchyma cell of the xylem tracheal and xylem rays.In paraffin sections, no fungi were found where abundant agar engendered, and agar was littlewhere the mycelium was found. Moreover, the fungus found in paraffin sections was notFusarium proliferatum. It had been shown that agar was not metabolites of growth andreproduction of Fusarium proliferatum in timber tissue of Aquilaria sinensis （Lour.） Gilg.8. After Added Aquilaria sinensis （Lour.） Gilg tissue culture seedling with culture solutionof Fusarium proliferatum and Hypocrea jecorina, there was significant difference in cellmembrane lipid oxidation, cell membrane permeability and glucose metabolism between thetwo groups of samples. From the view of cell membrane lipid peroxidation, the MDA contentin leaves of the samples added Hypocrea jecorina culture solution （H-group） increasedsignificantly, while the one of the samples added Fusarium proliferatum culture solution（F-group） increased less, especially in early phase （24h⁴8h）, which showed that Fusariumproliferatum, as a fungus more conducive to agar formation, could cause Aquilaria sinensis（Lour.） Gilg physiologic stress reaction, but the cell membrane lipid oxidation was not asserious as Hypocrea jecorina. From the view of cell membrane permeability changes, REC（Relative Electrical Conductivity） in the leaves of H-group increased steadily, while F-group sREC increased rapidly at first and then increases slowly. which showed that culture solution ofFusarium proliferatum would damage leaves s cell membrane gradually, while culturesolution of Hypocrea jecorina would damage acutely; From the view of soluble sugar, theleaves’ soluble-sugar in the two groups of samples all changed little in24h, and then allincreased rapidly with H-group more quickly, at the48thhour soluble-sugar content in theH-group samples was significantly higher than that in F-group samples, after the72thhour, thesoluble-sugar content in H-group samples declined more than that in F-group samples, whichindicated that Hypocrea jecorina infection caused nutrient transport disorder and resulted in irregular change of glucose metabolism, while Fusarium proliferatum effect on glucosemetabolism relatively stable.9. After Added Aquilaria sinensis （Lour.） Gilg tissue culture seedling with culture solutionof Fusarium proliferatum and Hypocrea jecorina, ROS surge phenomenon all occurred in ashort period （0²4h）. Later, at the96thhour the superoxide radical and hydrogen peroxide inthe H-group samples were higher than that of F-group samples by4.42U·g^-1and26.55U·g^-1.There were significant differences in active-oxygen scavenging enzymes during thephysiologic stress phase caused by Fusarium proliferatum and Hypocrea jecorina. Thedifferences were manifested in two aspects：firstly, the enzymes activity （including SOD, PODand CAT） of F-group samples were higher than that of H-group samples；Secondly, changeregularity of the enzymes activity was different, enzymes activity in F-group samples changedrelative steadily, while the one in H-group changed acutely.10. In this study, the hypothesis “physiological stress causing agar” was brought forward.Based on the hypothesis, a trial about agar revulsant formula, including Fusarium proliferatumculture solution, NaCl and JAs, was carried out according to uniform design. The results hadbeen shown that NaCl had greater effect on agarwood quanlity than JAs; the best agarrevulsant formula was Fusarium proliferatum culture solution added1.8248%NaCl and0.0046%JAs. Instilling the trunk with the agar revulsant formula, one year later, maximizedalcohol-soluble extractive content in the wood would be13.6012%theoretically.