JASCO

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J-810 Application Data Vol. I
The history of the measurement of optically
active molecules goes back a long way and
maintained a primitive state until 1812 when Biot
discovered the optical characteristics of quartz.
VantHoff and LeBel concluded (1874) that the
optical activity of matter is caused by
stereoisomerism of molecules. One hundred years
later (1960), Djetassi applied Optical Rotatory
Dispersion (ORD) to determine the absolute
configuration of steroids. This was the
beginning of the use of chiroptical spectroscopy
in the study of stereochemistry. Only one year
after Djerassis work was published (1961) JASCO,
realizing the need for chiroptical spectroscopy
instrumentation introduced the first commercial
automatic recording spectropolarimeter. Since
that time JASCO has supplied over 2000 units of
the worlds finest CD equipment. Without JASCO,
the advance of stereoisomerism research would
probably not have occurred as fast as it did.
Chiroptical spectroscopy instruments began with
the introduction of the Automatic recording
spectropolarimeter in 1961, the ORD/CD 1964, the
J-600 computer controlled CD, The J-710/720, The
J-715, and now JASCO offers the J-810 the latest
efforts to lead the fields of Chiroptical
spectroscopy.
JASCO J-810 Spectropolarimeter
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J-810 Application Data
Standard measurement
JASCO J-810 Spectropolarimeter
JASCO proudly announces the new model J-810
Spectropolarimeter, our latest Circular
Dichro-ism and UV / Visible absorbance
research-grade chiroptical spectrometer.
Following our suc-cessful J-715, the new J-810
offers a host of new hardware and software
features to save you time and deliver even better
results. Instrument control and data processing
are handled effort-lessly by our user-friendly
and innovative Spectra Manager Software running
on the Windows multi-tasking PC platform.
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J-810 Application Data
G value software A G value is CD(delta Abs)
divided by Abs. G values are known as a symmetric
factors and serves as an optical purity index.
Data is represented as G values for results
divided by the UV spectrum that measured the CD
spectrum measured by the J-820 using an
ultraviolet visible light spectrometer.
JASCO J-810 Spectropolarimeter
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J-810 Application Data
CD and Fluorescence simultaneous
measurement J-810 reads up to four channels of
data as a standard feature. Standard
specifications include CD/HT and CD/DC mode, but
it is also easy to implement a CD/fluorescent
simultaneous measurement system and read and
display temperature, pH and various polarity
signals.The data is a measurement example of
CD/HT mode(standard) for lysozyme and a
CD/fluorescent simultaneous measurement
system(optional).
JASCO J-810 Spectropolarimeter
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J-810 Application Data
Wide coverage from vacuum ultraviolet to
near-infrared region Assures a CD spectrum with
superior S/N all the way to the vacuum
ultraviolet region of 170 or less. This increases
the amount of information for secondary
structures such as proteins. The near-infrared
region is effective for such things as colored
proteins that include prosthetic groups such as
chiral transition metal complexes and
hemoproteins.
JASCO J-810 Spectropolarimeter
D-Camphor
Copper gluconate
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J-810 Application Data
Microcell The use of an optional micro cell
enables CD measurement at a low volume of
100micro litter. These options use a cylindrical
cell with low distortion. Rectangular micro cells
are also available.
JASCO J-810 Spectropolarimeter
MA-406/407 Micro cell attachment
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J-810 Application Data
Handles measurement of low-stray light, highly
absorbent samples In spectrometers with high
stray light, the CD spectrum is largely distorted
by high absorbance. The J-810 is equipped with an
extremely low stray light (0.0003 or
less)monochromator, enabling them to obtain a
highly precise CD spectrum even under high
absorbance. This data confirms that the CD
spectrum of NH4-d-10Camphorsulfonate does not
distort even when placing an Abs3 sample (an
alkali aqueous solution of potassium dichromate,
273 nm, OD3) into the optical path.
JASCO J-810 Spectropolarimeter
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J-810 Application Data
CD spectrum of lysozyme The ultraviolet CD
appearing in the middle near 280 nm on the right
side is caused by tryptophan residue and tyrosine
residue being buried within the tertiary
structures, thereby showing that lysozyme is
indeed lysozyme. The CD in this wavelength area
is sensitive to minute changes in substrates and
pH among other things, and is crucial even for
checking the correlation between protein
structure and activity. On the other hand, the
ultraviolet external CD below 250 nm on the left
side results in a peptide bond (amide group) as a
chromophore, reflecting secondary structures such
as alpha-helixes and beta sheets. CD is widely
applied as a means that can most easily observe
secondary structure.
JASCO J-810 Spectropolarimeter
Molecular model of lysozyme
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J-810 Application Data
Protein secondary structure analysis software We
estimate the helix and beta content () by using
a reference spectrum (Dr. Yang) for the secondary
structure to find the curves for protein CD. This
analysis technique is known as the reference
spectrum technique, but it can respond to
reference spectra that have different number of
secondary structure components (4 components is
standard) (2 to 8 components).
JASCO J-810 Spectropolarimeter
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J-810 Application Data
Temperature variation CD spectrum
JASCO J-810 Spectropolarimeter
CD spectra were obtained at a preset temperature,
and then displayed by overlay. Three-dimensional
display is also possible. Temperature control
range of -10 to 110degree Celsiua (used in a
range that is not boiling or freezing). Protein
higher-order structures (2 to 4 order structures)
change due to temperature, and this is reflected
in the CD spectrum.
Temperature scan data was obtained for CD in a
fixed wavelength by means of a constant
temperature rise program. This data was monitored
at wavelength of 220 nm, and it indicates that
secondary structure greatly changes near 70
degree Celsius.
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J-810 Application Data
Thermal degeneration analysis program
Employing a thermal denaturation program (new
optional software), and analyzing temperature
scan curves can obtain thermodynamic parameters
such as transition temperature, delta H, and
delta S.
JASCO J-810 Spectropolarimeter
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J-810 Application Data
Auto-titration
The auto-titration attachment performs syringe
control and CD data reads for titration using a
special program. The two types of data is
acidmetry data for poly-D-glutamate. Fig. 1
Titration curve of CD at fixed wavelength. Since
titration curve data is acquired as a text file,
it is output after being processed by Excel or
other programs. Fig. 2 Titration/wavelength
scan CD data. Wavelength scans are acquired in
the J-820 standard spectrum format.
JASCO J-810 Spectropolarimeter
Fig. 1
Fig. 2
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J-810 Application Data
Stopped-Flow CD Measurement
Stopped-flow CD for proteins is effectivefor
applications such as pH jumps and guanidine
denaturation and unwinding measurement. Fig. 1
Pre- and post- denaturation CD spectra due to pH
jump in myoglobin. Fig. 2 pH-jump T-scan data
at 222nm wavelength of myoglobin. We mixed 40 ul
of myoglobin neutral solution with 400 ul of
myoglobin acidic (pH3.31) solution (110), and
then observed the data. (8 integrations)
JASCO J-810 Spectropolarimeter
Fig. 2
Fig. 1
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J-810 Application Data
FDCD Measurement
The method of detecting CD using fluorescent
light is known as fluorescentdetection circular
dichroism (FDCD). When there is both a
fluorescent chromophoric group and
non-fluorescent chromophoric group within a
molecule, the CD of the former is often not
detected because it is hidden by the signal from
the latter that produces an even stronger CD. In
such cases, FDCD is extremely effective because
it can selectively capture the CD of the
fluorescent chromophoric group. The figure is an
example of selectively measuring FDCD in
tryptophan residue that was hidden by the normal
CD of Human serum albumin. Since proteins have
high fluorescent polarization, the polarizer is
inserted horizontally to prevent the appearance
of artifacts.
JASCO J-810 Spectropolarimeter
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J-810 Application Data
FDCD Measurement (High sensitivity measurement )
The FDCD attachment comes in either a
highlysensitive type or a standard type suited
for samples with a high degree ofpolarization
(the analyzer is an option) such as proteins.The
latter type also comes in a Peltier temperature
control type. The highsensitivity type is suited
to microdetection, while the standard type
issuited to chirality detection in fluorescent
probes embedded withinpolymers. The standard type
can also be used in wavelengthscan measurement of
fluorescent polarization. The figure is an
example of High-sensitivity measurement using
FDCD-405.
JASCO J-810 Spectropolarimeter
FDCD-405
Sample 1(S),2(S)-t-Cyclohexanediol bis
(6-methoxv-2-naphthoate) Concentrarion 212 ppb
(Solvent CH3CN, cell 10mm) (Dr. Nakanishi of
Columbia University)
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J-810 Application Data
Applications to Natural Organic Chemistry and
PharmaceuticalScience
CD and ORD are indispensable to thedetermination
and identification of natural organic compound
chirality.Chirality is identified by CD and ORD
signals. Effective empirical rules andtheoretical
rules are employed to that end. CD is limited to
the absorption band wavelength region of a
sample,but ORD is also effective for samples
without chromophoric groups such assaccharides
because it appears in all wavelength regions.
JASCO J-810 Spectropolarimeter
ORDE-402
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J-810 Application Data
ORD Attachment
Glucose and CD and UV lie on the wavelength side
shorter than 200 nm(lower). The upper right is an
example of mutarotation measurement of glucose
ORD. ORD is not only effective for samples in
which CD does not come out,but it also offers a
powerful means of identifying chiral compounds
along the lines of CD. There are two ORD
attachments. The ORDM-401 attachment, which
employs the optical zero method is suited to
precise photometry with anangle range /-45degree
wide. TheORDM-402 attachment for photoelectric
photometry has narrow angle range of /-1degree,
but its scanning speed is as fast as CD. CD
spectrum of glucose aqueous solution. There is a
CD peak at a wavelength shorter than 185 nm, but
measurement is not easy due to the absorption of
the water in the solution.
JASCO J-810 Spectropolarimeter
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J-810 Application Data
Application to Metal Complex Chemistry
CD and ORD are also widely used for chiral metal
complexes. In addition, they are used to measure
the metal complexes of polymers. Note that the
J-805 can handle short wavelength measurement
less than 400 nm, but it does not cover 400 to
800 nm. ORD and CD are also widely employed
inchiral metal complex compounds. The data to the
right is ameasurement example for
atris-ethylenediamine cobalt complex.
JASCO J-810 Spectropolarimeter
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J-810 Application Data
MCD Applications
Circular dichroism evoked by amagnetic field is
known and magnetic circular dichroism. CD
reflects thechirality of molecules, while MCD
reflects the electronstate (spin in particular)
of molecules and atoms.
JASCO J-810 Spectropolarimeter