| Dangshan Su Pear is native to China, which is also the dominant pear variety. However in recent years, stone cell content in Dangshan Su pears has been increasing, which results in rough, gritty texture, and seriously damages the flavor and quality of Dangshan Pear. In the study, through observing different periods of pear stone cell by microscopic, to understand the change of stone cell and parenchyma cell. Then research lignin deposition and transportation with ultra-microscopic observing, illustrated mechanism of the stone cell secondary wall formation. And then, analyzing lignin structure with spectroscopy, explore the relationship between the synthesis of lignin and lignin structure. Further more, through high performance liquid chromatography (HPLC) analysis of Intermediate metabolites; try to find out the main routes for the synthesis of lignin. The article is trying to provide the foundation for deep study about pear stone cell’s development mechanism.1. Results of Microscopy and Ultramicroscopic:(1) Stone cell clusters and parenchyma cells are arranged in a mosaic pattern.(2) Stone cell is the result of lignin deposition in the secondary wall deposition of thin wall cell wall thickening of the inhomogeneous formation. Pits in it came into being, when stone cell wall was just thickening. As the thickening of cell wall, it apparently observed that finally pits formed branched holes. With development of pear fruit cells; many pits in stone cell walls gradually shifted from the open state to the close state as the development of stone cell clusters as the development of stone cell cluster. Eventually leads to cell fully lignified.(3) Which deposit in stone cells was started from the outer cell wall, and extended from the secondary wall to the whole cell lumen. Lignin deposition with a style of numerous discrete granules occurred at the corner of the primary cell wall, and then extended to the complex heterogeneous granular intercellular layer. Cellulose microfibrils deposition at the S1, S2layer and its parallel arrangement along the circumference, lignin still deposited with granular unevenly along the direction of micro fibrils deposition from its inside. With the alternative arrangement between them, cellulose micro fibrils of the S2 layer and lignin granules filled up with the whole cell lumen.2. The results of FTR and’H-NMR:(1) Lignin of Dangshan pear fruit is a kind of G-S, and there are more guaiaeyl units than syringl units in its lignin. The value of G/S is increased first and then disincreased. Reached the biggest at63days (2.72) after flowering, which is agreement with the highest of lignin content. The value of G/S in the fruit of Dangshan Su Pear is higher than lignin of wood, and closely to the gramineous plant, so the lignin of Dangshan Su Pear has the lesser hardness.(2) Overall, the value of G/S and S lignin at early developed period (47days) is lower than the developmental period (after81days). S lignin is easier degradation than G lignin, which is the reason of the value of G/S slightly elevated.(3) Low condensation degree of pear fruit lignin. Lignin bond type each period is in β-O-4,β-1,β-5, β-β bonds.β-O-4is the mainly key bond, which plays an important role in lignin structure with the dissolved ability of lignin; the content of β-5is little, which is the condensation type. So Dangshan pear fruit has little condensed lignin. In addition, phenolic hydroxyl groups of lignin are less than30%, which is relation with polymerization. Even more MWL in the pulp of Pyrus bretshneideri contains hydrocarbon protons17.54%-34.50%. Lignin monomer is not high degree of polymerization; the side chain is more. This is the reason of pear fruit can edible in spite of lignin deposition.(4)Lignin structures in different periods are basically the same, but their chemical functional groups with a monomer composition proportion of different times is different. Such as C=O stretching in unconjugated ketene appeared in the early periods of pear development, and disappeared at the last periods. However. Aliphatic C-H stretching in CH3(not in Ome) and phen. O-H is happened at the last development periods. The difference of functional groups has great sense for the synthesis of lignin.3. Analysis with HPLC:(1) The content of coumaric acid is increased first, at63days reached a peak (0.0013μg/g), and then decreased, to about80days, the content basically unchanged. Cinnamic acid’s content was the highest at15days (0.231μg/g), shows decreased first and then increased and then decreased, on the63days (0.127μg/g) and81(0.0687μg/g) days reached a small peak. The content of sinapic acid and ferulic acid was also the highest at15days (0.003μg/g and0.0079μg/g), and then decreased, at63days reached a small peak. Caffeic acid’s content is very low during pear fruit development period, the levels rose rapidly from47days to63days; it has the highest value at63days (0.0353μg/g), to abou 80days, the content basically unchanged.(2) The change of several phenolic acids content shows that Cinnamic acid, ferulic acid, sinapic acid is mainly involved in the early development period of Pear stone cell. However, coumaric acid and Caffeic acid is mainly joined in the lignin synthesis at39days to63days, which is peak of lignin synthesis, and also form of stone cell (47days) and the peak of stone cell clusters (63days).(3) Regression analysis indicated that coumaric acid and lignin synthesis has significant positive correlation. While cinnamon acid is the precursors of coumaric acid. So we can know that lignin from cinnamon acid and coumaric acid in the synthesis of way, and coumaric acid is the main precursors of lignin material. |
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