量測發酵液之NADH光電感測器

Measuring the fermentation broth of NADH photoelectric sensor

Presenter: Shih-Chieh ChaoAdviser: Dr. Hung-Chi Yang

Chairman: Dr. Hung-Chi YangDate:05.08.2013

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OutlineBackgroundIntroductionPurposeMethods & MaterialsExperiment Future worksReference

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BackgroundFluorescent luminous phenomenon is a light cooling. When

some kind of substance at room temperature by a certain wavelength of the incident light (usually ultraviolet or X-ray) radiation, absorb light energy into excited states and decay and issued by the outgoing light.

Human tissue containing many fluorescent substances, including collagen, NADH, FAD, rhodopsin, will be excited by UV fluorescence.

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BackgroundHuman tissue in the cancer process, will produce a change

in the structure, such as: the cells become less dense connective tissue collagen tissue to reduce; metabolism of cancer cells than normal cells, so that the increased NADH; caused by increased blood flow absorbed rhodopsin rise. Therefore, the fluorescence spectrum of the cancer tissue and normal tissue will be different, and performance in the fluorescence spectral peak position and strength of the ebb and flow.

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IntroductionNADH( 煙 醯 胺 腺 嘌 呤 二 核 苷 酸 ) NADH is a

widespread variety of animals, plants and humans in living cells, natural substances , and is a very important compound, was not discovered until 1905, usually with many enzymes in the body co-participate in the reaction, usually called "coenzyme“.

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IntroductionCell growth and energy metabolism in the cells are required

to NADH, food into energy process requires NADH participate, and in the cell to produce energy in the process, NADH plays the important role of the portable electronic, in addition to NADH is also a very important the antioxidants can protect cells from damage of the harmful substances.

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IntroductionNADH Excitation light of 340 ± 20nm.

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Introduction

UV-A UV-B UV-C

Wavelength 400 ～ 315nm 315 ～ 280nm 280 ～ 200nm

Nociceptive weak medium strong

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IntroductionThe most important function of the Fermenter, is to make

the microorganisms in the growth conditions it needs to reach the desired proliferation.

Therefore the design and production of the fermentation tank you need to have a lot of considerations.

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IntroductionNeed to maintain long-aseptic operation.Microbial metabolism of aeration and agitation, but can not

give microbial mechanical shock stirring intensity.Need to configure the device temperature system Imperial.Need to be able to control the pH.Need to be able to sample.Must be appropriate size.Internally to maintain a smooth inner surface, not have

welded junction.By fermentation of a variety of different, you need to install

the necessary equipment incidental.

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Introduction

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PurposeThis study focused on :

The sensor can be fully inserted fermenterContinuous and real-time performance measurement

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Detector

Broth+

fungusLED

MCU Control

PC

Purpose

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340 nm

450 nm

Purpose

30F4011

UART

PWM

TIMER

A/D

LED*4

DETPC

RS232

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Methods & MaterialsSolidwork

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Methods & MaterialsType 304—the most common grade; the classic 18/8 stainless steel.

Outside of the US it is commonly known as "A2 stainless steel”.Type 316—the second most common grade (after 304); for food and

surgical stainless steel uses; alloy addition of molybdenum prevents specific forms of corrosion. It is also known as marine grade stainless steel due to its increased resistance to chloride corrosion compared to type 304. 316 is often used for building nuclear reprocessing plants.

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Methods & Materials

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Methods & Materials

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LED

Detector

Methods & Materials

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Methods & Materials

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10mm

44mm

20mm

44mm

40mm

10mm

20mm12mm

8mm

Methods & Materials

40mm

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Methods & Materials

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Methods & Materials

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30mm

14mm

26mm

18mm14m

m26mm30mm

Methods & Materials

26mm24

Methods & MaterialsExcitation light ： NSHU551B

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Methods & MaterialsKupo ： UV Transmission

300 400 500 600 nm

stut

0102030405060708090

100%T

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Methods & MaterialsNADH emit light of 450±20nm ， so the selection of the

received light 320-730nm.Hamamatsu ： S1133

spectral response range 320~730nm

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Methods & MaterialsKupo ： C11715

300 400 500 600 nm

stut

0102030405060708090

100%T

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Methods & MaterialsKupo ： UV Blocking

300 400 500 600 nm

stut

0102030405060708090

100%T

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300 400 500 600 nm

stut

0102030405060708090

100%T

Methods & MaterialsUV Blocking+C11715

300 400 500 600 nm

stut

0102030405060708090

100%T

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Experiment

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Experiment

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Experiment

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Experiment

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ExperimentFluorescein, 90+% L13251Formula: ： C20H12O5 Molecular weight :332.32Orange powder

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Experiment

5mL AOH(47.5%) 5mL fluoresein(1mM)

0.5mM

5ml AOH(47.5%) 5mL fluoresein(0.5mM)

0.25mM

5ml AOH(47.5%) 5mL fluoresein(0.25mM)

0.125mM

5ml AOH(47.5%) 5mL fluoresein(0.125mM)

0.0625mM

Deployment method, beginning first with a 1-to-1 water diluted to 47.5% alcohol with 95% alcohol(AOH)

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Experiment

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Experiment

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ExperimentSigma N8129 ：

Reduced coenzyme Yellowish white powderStored at -20 ℃Excitation light = 340 nm 。Emit light = 460 nm 。

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Experiment Fluorescence Spectrometer F-7000

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Future worksCircuit design30F4011Labview

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Reference[1] Y-C Liu, F-S Wang, W-C Lee, “ On-line monitoring and controlling system for fermentation processes. “

Biochemical Engineering Journal 7 (2001) 17–25.[2] Monici M. (2005). "Cell and tissue autofluorescence research and diagnostic

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RR, Van Dam J, Feld MS (2002-02-01). "NAD(P)H and collagen as in vivo quantitative fluorescent biomarkers of epithelial precancerous changes". Cancer Res. 62 (3): 682–687. PMID 11830520

[4] Windholz, Martha (1983). The Merck Index: an encyclopedia of chemicals, drugs, and biologicals (10th ed.). Rahway NJ, US: Merck. p. 909. ISBN 911910271 Check |isbn= value (help).

[5] Bakker BM, Overkamp KM, van Maris AJ, et al. (2001). "Stoichiometry and compartmentation of NADH metabolism in Saccharomyces cerevisiae". FEMS Microbiol. Rev. 25 (1): 15–37.

[6] Nicholls DG; Ferguson SJ (2002). Bioenergetics 3 (1st ed.). Academic Press. ISBN 0-12-518121-3.[7] Sistare FD, Haynes RC (15 October 1985). "The interaction between the cytosolic pyridine nucleotide redox potential

and gluconeogenesis from lactate/pyruvate in isolated rat hepatocytes. Implications for investigations of hormone action". J. Biol. Chem. 260 (23): 12748–53. PMID 4044607. 

[8] Freitag A, Bock E (1990). "Energy conservation in Nitrobacter". FEMS MicrobiologyLetters 66 (1–3): 157–62. doi:10.1111/j.1574-6968.1990.tb03989.x.

[9] Starkenburg SR, Chain PS, Sayavedra-Soto LA, et al. (2006). "Genome Sequence of the Chemolithoautotrophic Nitrite-Oxidizing Bacterium Nitrobacter winogradskyi Nb-255". Appl. Environ. Microbiol. 72 (3): 2050–63. doi:10.1128/AEM.72.3.2050-2063.2006. PMC 1393235. PMID 16517654.

[10] Dawson, R. Ben (1985). Data for biochemical research (3rd ed.). Oxford: Clarendon Press. p. 122. ISBN 0-19-855358-7.

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