Luận văn thạc sĩ Kỹ thuật hóa học: Nghiên cứu trích ly hoạt chất từ nấm linh chi và thử nghiệm hoạt tính sinh học của cao trích

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(Extraction of biological activities from Ganoderma lucidum

and examination of bioactivities)

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7rQWLӃng Anh: Extraction of biological activities from Ganoderma lucidum

and examination of bioactivities

II NHIӊM VӨ 9¬1ӜI DUNG:

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ÿӇ WuPÿLӅu kiӋQWUtFKO\FKRKjPOѭӧng triterpenoid cao nhҩt

Ĉӗng thӡLFiFQKyP chӭFÿһFWUѭQJFӫDFDRWUtFKKuQKWKiLFҩXWU~FEӅ mһt nҩPYjWKjQKSKҫQFDRWUtFKÿѭӧc khҧRViWEҵng phә hӗng ngoҥi chuyӇQKyD)RXULHU)RXULHU

± transform infrared spectroscopy ± )7,5  NtQK KLӇQ YL ÿLӋn tӱ TXpW VFDQ HOHFWURQmicroscopy ± 6(0 YjVҳc NêNKtNKӕi phә (Gas chromatography ± mass spectrometry

± GC±MS)

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khҧ QăQJ NKiQJ NKXҭn cӫa triterSHQRLG ÿѭӧc thӱ nghiӋP WUrQ Escherichia coli,

Staphylococcus aureus, Pseudomonas aeruginosa Yj Salmonella enterica bҵng

SKѭѫQJSKiSÿRÿѭӡQJNtQKYzQJNKiQJNKXҭn

ABSTRACT

In this thesis, triterpenoid was extracted from Vietnamese Ganoderma lucidum (G

lucidum) by the ultrasonic-assisted enzyme extraction (UAEE) method The

triterpenoid content was determined by ultraviolet±visible spectroscopy (UV±Vis)

Chemical composition and heavy metal content in G lucidum have been investigated, showing that G lucidum has high fiber, carbohydrate and lipid content

In addition, the content of heavy metals such as As, Cd, Hg and Pb was within the allowable limit

The influence of each extraction condition such as enzyme concentration, etanol:material ratio, temperature, time and power on triterpenoid content was investigated by the method of alternating each variable to find a suitable range of variation Then, conducting a simultaneous survey by experimental planning using the Plackett±Burman and Box±Behnken designs The data were processed using Design Expert 11.0 software to find extraction conditions for the highest triterpenoid content

At the same time, the characteristic functional groups of the extract, the morphology

of the surface structure of the fungus and the high composition of the extract were investigated by Fourier transform infrared spectroscopy (Fourier transform infrared spectroscopy ± FTIR), scanning electron microscopy (scan electron microscopy ± SEM), and gas chromatography ± mass spectrometry (GC±MS)

The extract at optimal conditions was tested for biological activity through antitumor activity on three human cancer cell lines (Hep-G2 and MCF-7) and the

antibacterial ability of triterpenoids was tested on Escherichia coli, Staphylococcus

aureus, Pseudomonas aeruginosa, and Salmonella enterica by antibacterial ring

diameter measurement

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ABSTRACT v

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DANH MӨ&+Î1+ ix

DANH MӨC BҦNG xi

DANH MӨ&&È&7Ӯ VIӂT TҲT xii

ĈҺT VҨ1Ĉӄ 1

TӘNG QUAN 2

&+ѬѪ1* 1.1 Tәng quan vӅ nҩm linh chi 2

1.1.1 Giͣi thi͏u chung 2

7KjQKSK̯QKRiK͕c 3

1.2 HoҥWWtQKVLQKKӑc cӫa triterpenoid 8

.KiQJR[\KRi 8

.KiQJXQJWK˱ 9

.KiQJNKX̱n 10

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3K˱˯QJSKiSYLVyQJ 12

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3K˱˯QJSKiSVLrXkP 14

3K˱˯QJSKiSHQ]\PH 17

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7URQJQ˱ͣc 18

1JRjLQ˱ͣc 18

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1.5.1 MͭFWLrX 19

1.5.2 N͡i dung 19

3K˱˯QJSKiSQJKLrQFͱu 20

1.5.4 ĈyQJJySFͯa lu̵QYăQ 28

THӴC NGHIӊM 29

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2.1.2 Thi͇t b͓ 29

Ĉ͓DÿL͋m th͹c hi͏n lu̵QYăQ 30

7KtQJKLӋm 30

2.2.1 Kh̫RViWWKjQKSK̯QKRiK͕c trong ṋm linh chi 30

2.2.2 Kh̫RViWV͹ ̫QKK˱ͧng cͯDFiF\͇u t͙ ÿ͇QKjPO˱ͫng triterpenoid 33

3KkQWtFKÿ̿F WtQKKyD OêWKjQKSK̯n cͯDFDRWUtFKYj F̭X WU~FQJX\rQ li͏XWU˱ͣFYjVDXNKLWUtFKO\ 42

2.2.4 Th͵ nghi͏m ho̩WWtQKVLQKK͕c cͯDFDRWUtFK 43

KӂT QUҦ 9¬%¬1/8ҰN 45

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3.2 ҦQKKѭӣng cӫDFiF\Ӄu tӕ ÿӃQKjPOѭӧng triterpenoid 46

3.2.1 ̪QKK˱ͧng cͯa tͳng y͇u t͙ 46

3.2.2 ̪QKK˱ͧQJÿ͛ng thͥi cͯDFiF\͇u t͙ 52

3.2.3 T͙L˱XKRiÿL͉u ki͏QWUtFKO\WULWHUSHQRLG 55

ĈһFWtQKKRiOêFӫDFDRWUtFKYjFҩXWU~FQJX\rQOLӋu 61

3KkQWtFKFiFQKyPFKͱFWURQJFDRWUtFK 61

7KjQKSK̯n ho̩t ch̭t trong cao WUtFK 62

3.3.3 C̭XWU~FE͉ m̿t cͯDQJX\rQOL͏XWU˱ͣFYjVDXNKLWUtFKO\ 63

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.KiQJXQJWK˱ 65

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7¬,/,ӊU THAM KHҦO 102

PHӨ LӨC 112

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+uQK&{QJWKӭc cҩu tҥo cӫa squalen 5

+uQK&iFWULWHUSHQRLGWKXӝFQKyPĮ-amyrin 6

+uQK&iFWULWerpenoid thuӝFQKyPȕ amyrin 7

+uQK&iFWULWHUSHQRLGWKXӝFQKyPlupeol 7

+uQK&ѫFKӃ NKiQJR[\KyDEҵQJFiFKFKRÿLӋn tӱ 9

+uQKҦQKKѭӣng cӫDWULWHUSHQRLGÿӃQTXiWUuQKWӃ EjRFKӃWWKHRFKXNu 10

+uQK&ѫFKӃ NKiQJNKXҭn cӫa triterpenoid 11

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+uQK0{Wҧ hӋ thӕQJWUtFKO\EҵQJVyQJVLrXkPGҥng bӇ 16

+uQK4XiWUuQKSKiYӥ cҩXWU~FWӃ EjRFӫa enzyme 17

+uQK0{KuQK%R[-Behnken 22

+uQK6ѫÿӗQJX\rQOêPi\ÿRSKә89±Vis 23

+uQKCҩu tҥo thiӃt bӏ quang phә FTIR 24

+uQK6ѫÿӗ QJX\rQOêPi\VҳFNêJKpSNKӕi phә GC-MS 25

+uQK1JX\rQWҳc hoҥWÿӝQJPi\6(0 26

+uQK1JX\rQWҳc thӱ nghiӋm hoҥWWtQKNKiQJXQJWKѭ 26

+uQK4X\WUuQKWUtFK ly triterpenoid tӯ nҩm linh chi bҵQJSKѭѫQJSKiSenzyme kӃt hӧSVLrXkP 34

+uQKPhә UV ± YLVYjÿѭӡng cong chuҭn cӫa axit ursolic ӣ ɉ݉ܽݔ ± 548 nm 41

+uQKĈөc lӛ WUrQEӅ mһWÿƭDWKҥch 44

+uQK+jPOѭӧng kim loҥi nһng trong nҩm linh chi* 46

+uQKҦQKKѭӣQJFӫDQӗQJÿӝHQ]\PHÿӃQKjPOѭӧQJWULWHUSHQRLG 47

+uQKҦQKKѭӣng cӫa tӹ lӋ dung P{LQJX\rQOLӋXÿӃQKjPOѭӧng triterpenoid 48

+uQKҦQKKѭӣng cӫa nhiӋWÿӝ VLrXkPÿӃQKjPOѭӧng triterpenoid 49

+uQKҦQKKѭӣng cӫa thӡLJLDQVLrXkPÿӃQKjPOѭӧng triterpenoid 50

+uQKҦQKKѭӣng cӫDF{QJVXҩWVLrXkPÿӃQKjPOѭӧng triterpenoid 51+uQK(a) BiӇXÿӗ SKkQSKӕi nӱa chuҭQYjE %LӇXÿӗ 3DUHWRFiFÿLӇm A, B, C, D, Yj( 55+uQKҦQKKѭӣQJÿӗng thӡi cӫa nӗQJÿӝ enzyme YjWӹ lӋ HWDQROQJX\rQOLӋu 58+uQKҦQKKѭӣQJÿӗng thӡi cӫDF{QJVXҩWVLrXkPYjQӗQJÿӝ enzyme 59+uQKҦQKKѭӣQJÿӗng thӡi cӫa etanol: tӹ lӋ QJX\rQOLӋXYjF{QJVXҩWVLrXkP 60+uQK3 11: Phә FTIR cӫDFDRWUtFK 61+uQK +uQKҧnh SEM cӫa quҧ thӇ nҩPOLQKFKLDE WUѭӟFYjFG VDXNKLWUtFKO\63

+uQK : 9QJ ӭc chӃ cӫa (a) E coli , (b) S aureus , (c) P aeruginosa Yj G