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00:00:02,660 --> 00:00:06,060
Everyone, thanks for
being here today to him,

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00:00:06,060 --> 00:00:07,410
I report on nerve targeted

3
00:00:07,410 --> 00:00:08,730
for our first synthesis.

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My name is
Cassandra Mathison

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and this project
was a collaboration

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between the LHS
you Gibbs lab

7
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and the strong in organic

8
00:00:15,240 --> 00:00:16,770
chemistry lab at PSU,

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of which I'm grateful
to be apart.

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Before I begin, I want

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to give a special
thanks to Dr.

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00:00:21,270 --> 00:00:22,890
Robert Strong in
my nickname and to

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her and to Dr. summer al.

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Gibbs for allowing
me to assist

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in this exciting project.

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Special thanks also to

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Dr. Les Wang for
his guidance and

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expertise in the
daily activities

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00:00:32,040 --> 00:00:33,775
involved in this research.

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Okay, so let's
get started.

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Nerve tissue damage as

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00:00:38,990 --> 00:00:40,310
a result of
surgical injury is

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00:00:40,310 --> 00:00:41,390
a common negative outcome

24
00:00:41,390 --> 00:00:43,355
during many surgical
interventions.

25
00:00:43,355 --> 00:00:45,080
At nerve, neuropathic pain

26
00:00:45,080 --> 00:00:46,325
is currently
experienced by an

27
00:00:46,325 --> 00:00:47,630
estimated 3% of

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the developed
world population.

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00:00:49,385 --> 00:00:50,810
Due to its
persistence well

30
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beyond healing time,

31
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post-surgical nerve
tissue injury may

32
00:00:53,810 --> 00:00:55,160
potentially have
a life altering

33
00:00:55,160 --> 00:00:56,555
effect on the patient.

34
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Depending on the
site of the injury

35
00:00:58,340 --> 00:00:59,960
and the amount of
cellular damage,

36
00:00:59,960 --> 00:01:01,460
neuropathic pain
resulting from

37
00:01:01,460 --> 00:01:02,510
surgical injury may cause

38
00:01:02,510 --> 00:01:05,555
severe debilitation
and interfered

39
00:01:05,555 --> 00:01:07,625
greatly with
quality of life.

40
00:01:07,625 --> 00:01:09,740
Surgical training
does involve

41
00:01:09,740 --> 00:01:11,570
mapping typical
anatomical planes

42
00:01:11,570 --> 00:01:12,710
in order to plan
avoidance of

43
00:01:12,710 --> 00:01:15,289
major nerves during
surgical intervention.

44
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Individual interior
landscapes

45
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may vary greatly, however,

46
00:01:18,170 --> 00:01:19,685
as a result of
previous trauma,

47
00:01:19,685 --> 00:01:21,980
past surgery or married
other conditions,

48
00:01:21,980 --> 00:01:24,455
when anatomical
structures are atypical,

49
00:01:24,455 --> 00:01:26,165
the risk of nerve injury

50
00:01:26,165 --> 00:01:27,740
during surgery is
additionally height

51
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and then some nerve groups

52
00:01:29,810 --> 00:01:31,445
our site enough that
they prove hard to see,

53
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even when they
are optically

54
00:01:32,914 --> 00:01:35,040
magnify to natural light.

55
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In order to
substantially reduce

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the amount of
surgical injury to

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nerve tissue and

58
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associated chronic
post-operative nerve pain.

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Clinical supports for
greater visualization

60
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during surgical
intervention are

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imperative that
the benefit

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of more highly visible

63
00:01:50,620 --> 00:01:51,730
nerve tissue was clear.

64
00:01:51,730 --> 00:01:52,720
Previous approaches to

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improving visualization

66
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during surgery had been

67
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limited in their
resolution,

68
00:01:56,439 --> 00:01:59,260
specificity and
real-time applications.

69
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Surgery That's
fluorescence guided may

70
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highlight complex nerve
structure patterns

71
00:02:03,160 --> 00:02:04,480
with high contrast against

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background tissues and
with high sensitivity.

73
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This real-time

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00:02:08,290 --> 00:02:09,760
intraoperative
visual assistance

75
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may significantly improve

76
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patient outcomes
by helping

77
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surgical staff
to recognize

78
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delicate nerve
structures and

79
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thus avoid inflicting
surgical injury.

80
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The nature of small
molecule fluorophores

81
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is such that they have

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naturally high
specificity of

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uptake in nerve tissue.

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Additionally, the
low molecular weight

85
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belonging to
small molecule

86
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fluorophores may allow

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them to bypass the
blood-brain barrier for

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successful uptake in

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central nervous
system tissue.

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Additionally,
uptake me, be

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successful in the
blood nerve barrier.

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Dividing the
nerve axons of

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the peripheral
nervous system from

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00:02:42,470 --> 00:02:44,390
the, from the bloodstream.

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These qualities provide

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exciting prospects for

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future intraoperative
applications in

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both the peripheral and

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central nervous systems,

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which is lacking enlarge.

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Contrast agents such as

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00:02:55,970 --> 00:02:58,830
nerve specific
peptides for example.

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00:02:59,680 --> 00:03:01,670
So one challenge to

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00:03:01,670 --> 00:03:02,660
the clinical
application of

105
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such fluorophores
is that they may

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offer low quantum yield.

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That is, the
absorbed photons may

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00:03:07,580 --> 00:03:09,995
not equate to acceptable
light emission.

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This presents obvious
problems when

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considering their use an

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image guided surgery.

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And additionally,
a problems with

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solubility instability
can occur.

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So achieve charge of
this research project

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is to discover
chemical variance and

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the molecules which will
allow them to retain

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nerve specificity while

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correcting those
aforementioned challenges.

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What you see here as
the general structure

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of an oxime
fluorophore with

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associated key oxygen
for our forests provide

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the starting material for

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the chemical manipulations

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conducted within
this research.

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So when you're developing

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a chemical
synthesis scheme,

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It's somewhat similar
to writing a recipe.

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At.

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This allows sequential

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progression from chemical,

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from starting material to

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chemical compound
precursors

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to two final fluorophores,

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which we named LG
WE 842 and 840 for

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both of which should have

136
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near infrared emission

137
00:04:04,985 --> 00:04:08,720
from 650 to 900
nanometers.

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For our purposes, if
we're talking about

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00:04:11,840 --> 00:04:14,570
writing a recipe in
designing our scheme,

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00:04:14,570 --> 00:04:16,639
oxacillin fluorophores

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00:04:16,639 --> 00:04:18,500
would serve as our flower.

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And then I'll GW 842

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00:04:20,750 --> 00:04:23,060
and 44 would be
our eventual cake.

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So to get to the cake,

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You must not only
have the flower,

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but you must also make

147
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special preparations to

148
00:04:28,820 --> 00:04:30,605
some of your ingredients,

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which for our
purposes would

150
00:04:32,060 --> 00:04:33,740
be our precursors

151
00:04:33,740 --> 00:04:35,420
before you can combine

152
00:04:35,420 --> 00:04:36,500
your ingredients to get

153
00:04:36,500 --> 00:04:38,480
your desired end product.

154
00:04:38,480 --> 00:04:41,210
So synthesis of
each precursor

155
00:04:41,210 --> 00:04:42,710
and the scheme
proceedings,

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00:04:42,710 --> 00:04:45,170
synthesis of the final
compounds includes

157
00:04:45,170 --> 00:04:46,490
chemical manipulations of

158
00:04:46,490 --> 00:04:48,485
commercially available
fluorophores.

159
00:04:48,485 --> 00:04:50,900
And then each precursor
is synthesized via

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00:04:50,900 --> 00:04:53,060
a single-step
reactions that involve

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00:04:53,060 --> 00:04:55,580
manipulation of
either five methoxy,

162
00:04:55,580 --> 00:04:57,155
two methyl Annalen,

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00:04:57,155 --> 00:04:59,240
or three amino for
methyl phenyl,

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00:04:59,240 --> 00:05:01,430
or a previously
synthesized precursor

165
00:05:01,430 --> 00:05:02,630
originating from those

166
00:05:02,630 --> 00:05:04,130
aforementioned compounds.

167
00:05:04,130 --> 00:05:05,450
Then as a final step,

168
00:05:05,450 --> 00:05:07,040
precursor compounds
are combined.

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Vietnam condensation
reactions in order to

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build our final
compounds are beautiful.

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Looks like two-year-old
birthday cake.

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Following the synthesis of

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each precursor
are samples of

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the product are
evaluated in

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00:05:21,580 --> 00:05:23,335
the lab via
multi-step method.

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Before progressing
forward with the scheme.

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Thin-layer chromatography
pictured here on

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the side is used as

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a first step in confirming

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progress of each
reaction by revealing

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the presence of product

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versus starting material.

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Tlc plates are coded in

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00:05:36,580 --> 00:05:38,515
a silica gel
stationary phase.

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And then a
deposited sample

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00:05:41,260 --> 00:05:42,310
of a reaction starting

187
00:05:42,310 --> 00:05:43,660
material is compared with

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00:05:43,660 --> 00:05:44,980
the deposited sample of

189
00:05:44,980 --> 00:05:47,155
a synthesized product
of the reaction.

190
00:05:47,155 --> 00:05:48,880
Bands will then
develop according to

191
00:05:48,880 --> 00:05:50,020
each compounds affinity

192
00:05:50,020 --> 00:05:51,325
with the stationary phase.

193
00:05:51,325 --> 00:05:53,080
So you can see here
that the absence

194
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of the band
corresponding with

195
00:05:54,370 --> 00:05:56,080
the starting
material which we

196
00:05:56,080 --> 00:05:57,220
deposited on the
left side of

197
00:05:57,220 --> 00:05:59,130
the plate labeled 1807.

198
00:05:59,130 --> 00:06:00,410
In addition to a newly

199
00:06:00,410 --> 00:06:01,970
developed higher
concentration band

200
00:06:01,970 --> 00:06:03,770
and the product
area of the play

201
00:06:03,770 --> 00:06:06,230
there on the
right labeled 824

202
00:06:06,230 --> 00:06:09,605
indicates a
completed reaction.

203
00:06:09,605 --> 00:06:12,080
So TLC is a valuable
tool as it's relatively

204
00:06:12,080 --> 00:06:13,490
inexpensive and takes

205
00:06:13,490 --> 00:06:15,755
approximately five
minutes to complete.

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00:06:15,755 --> 00:06:18,140
An additional analytical
step in assessing

207
00:06:18,140 --> 00:06:19,760
reaction progress
is the use of

208
00:06:19,760 --> 00:06:21,800
liquid chromatography,
mass spectrometry,

209
00:06:21,800 --> 00:06:24,410
or LC-MS, which
uses mass-to-charge

210
00:06:24,410 --> 00:06:25,760
ratios to identify and

211
00:06:25,760 --> 00:06:27,920
delineate between
chemical compounds.

212
00:06:27,920 --> 00:06:30,320
It's also a valuable
tool for evaluating

213
00:06:30,320 --> 00:06:32,600
the progress of
chemical reactions by

214
00:06:32,600 --> 00:06:34,040
using associated masses to

215
00:06:34,040 --> 00:06:35,660
identify remaining
regions in

216
00:06:35,660 --> 00:06:39,230
solution versus intended
synthesized product.

217
00:06:39,230 --> 00:06:41,360
In the lab, we use
liquid chromatography

218
00:06:41,360 --> 00:06:43,175
with photodiode
array detection,

219
00:06:43,175 --> 00:06:45,530
which offers additional
data related to

220
00:06:45,530 --> 00:06:47,210
the classification
of compounds

221
00:06:47,210 --> 00:06:49,429
and their absorbance
respectively.

222
00:06:49,429 --> 00:06:51,770
So DID is
especially valuable

223
00:06:51,770 --> 00:06:53,930
for the purposes of
this research project.

224
00:06:53,930 --> 00:06:55,340
As the intended product

225
00:06:55,340 --> 00:06:56,360
of each reaction will have

226
00:06:56,360 --> 00:06:59,300
not only a specific
mass-to-charge ratio,

227
00:06:59,300 --> 00:07:01,820
but also substantial
fluorescent properties

228
00:07:01,820 --> 00:07:03,950
and location of peaks
in the readings

229
00:07:03,950 --> 00:07:06,200
for both
mass-to-charge and DAD

230
00:07:06,200 --> 00:07:07,520
is an excellent way to

231
00:07:07,520 --> 00:07:08,750
narrow down the
location of

232
00:07:08,750 --> 00:07:11,000
highest product
concentration within

233
00:07:11,000 --> 00:07:12,050
the product sample as an

234
00:07:12,050 --> 00:07:13,760
analytical step to then

235
00:07:13,760 --> 00:07:15,800
be able to anticipate
its location in

236
00:07:15,800 --> 00:07:18,980
the next step of product
isolation via crass,

237
00:07:18,980 --> 00:07:22,290
excuse me, via flash
chromatography.

238
00:07:22,870 --> 00:07:28,325
Here you can see an
example digital reading

239
00:07:28,325 --> 00:07:30,920
that would be

240
00:07:30,920 --> 00:07:34,160
generated during
flash chromatography.

241
00:07:34,160 --> 00:07:36,890
Flash chromatography
also separate substances

242
00:07:36,890 --> 00:07:37,970
according to
their affinity

243
00:07:37,970 --> 00:07:39,245
with the stationary phase.

244
00:07:39,245 --> 00:07:41,915
However, this separation
is now preoperative.

245
00:07:41,915 --> 00:07:43,160
Isolating compounds

246
00:07:43,160 --> 00:07:44,704
from solution
into fractions,

247
00:07:44,704 --> 00:07:45,950
the most pure of

248
00:07:45,950 --> 00:07:47,480
which can then
be combined and

249
00:07:47,480 --> 00:07:51,180
worked up into a
purified final product.

250
00:07:52,270 --> 00:07:55,190
And here is what we
are left with after

251
00:07:55,190 --> 00:07:58,024
purification of the
isolated final compounds.

252
00:07:58,024 --> 00:08:00,020
I love the stark
contrast between

253
00:08:00,020 --> 00:08:02,660
the precursor solutions
and these here on

254
00:08:02,660 --> 00:08:04,550
the left are
previously synthesized

255
00:08:04,550 --> 00:08:06,125
fluorophores in solution

256
00:08:06,125 --> 00:08:07,730
pictured in natural light.

257
00:08:07,730 --> 00:08:09,290
And then on the right,

258
00:08:09,290 --> 00:08:10,430
those same fluorophores

259
00:08:10,430 --> 00:08:12,125
under laser excitation.

260
00:08:12,125 --> 00:08:15,690
I personally find
them quite beautiful.

261
00:08:16,930 --> 00:08:19,700
Near-infrared fluorophores
have been shown to

262
00:08:19,700 --> 00:08:20,300
highlight some of

263
00:08:20,300 --> 00:08:21,800
the most delicate
nervous tissue in

264
00:08:21,800 --> 00:08:23,870
the peripheral nervous
system of mammals with

265
00:08:23,870 --> 00:08:26,210
high contrast against
background tissues,

266
00:08:26,210 --> 00:08:27,710
nerves which would
be invisible in

267
00:08:27,710 --> 00:08:30,635
white light become
brightly illuminated.

268
00:08:30,635 --> 00:08:34,865
Following successful
synthesis of 842 in 44,

269
00:08:34,865 --> 00:08:36,110
optical properties of

270
00:08:36,110 --> 00:08:37,370
fluorescence and
absorption are

271
00:08:37,370 --> 00:08:39,170
evaluated by
digital processing

272
00:08:39,170 --> 00:08:40,910
of the isolated
product sample.

273
00:08:40,910 --> 00:08:42,515
As previously stated,

274
00:08:42,515 --> 00:08:44,450
near infrared
emission is ideal.

275
00:08:44,450 --> 00:08:46,970
So this slide shows
photographs in

276
00:08:46,970 --> 00:08:48,380
conventional
white light and

277
00:08:48,380 --> 00:08:49,820
then fluorescent
images and

278
00:08:49,820 --> 00:08:52,310
corresponding fluorescence
intensity through

279
00:08:52,310 --> 00:08:53,749
cross-sectional profiles

280
00:08:53,749 --> 00:08:54,920
for visible and Miss have

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00:08:54,920 --> 00:08:56,270
oxygen for there on

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00:08:56,270 --> 00:08:58,850
the left and near
infrared emissive oxygen

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00:08:58,850 --> 00:09:00,770
for a on the rate.

284
00:09:00,770 --> 00:09:02,840
This collection
of images really

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00:09:02,840 --> 00:09:05,270
outlines the efforts of
this project for me.

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00:09:05,270 --> 00:09:06,560
Taking fluorophore,

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00:09:06,560 --> 00:09:08,210
which naturally
lend themselves to

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00:09:08,210 --> 00:09:10,730
nerve specific application
and fine tuning

289
00:09:10,730 --> 00:09:12,290
them to be ideal
candidates

290
00:09:12,290 --> 00:09:14,490
for clinical application.

291
00:09:15,370 --> 00:09:17,315
The time in this report,

292
00:09:17,315 --> 00:09:19,160
this project is ongoing.

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00:09:19,160 --> 00:09:20,570
The synthesis has been

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00:09:20,570 --> 00:09:22,130
successfully completed now

295
00:09:22,130 --> 00:09:25,730
for LG W 842 and
40 for both,

296
00:09:25,730 --> 00:09:27,740
which is super exciting.

297
00:09:27,740 --> 00:09:29,285
And in the time to come,

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00:09:29,285 --> 00:09:31,340
data as previously
outlined,

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00:09:31,340 --> 00:09:33,080
will be collected
for each compound.

300
00:09:33,080 --> 00:09:34,490
And in the case of either

301
00:09:34,490 --> 00:09:36,290
show traits which
highlight them as

302
00:09:36,290 --> 00:09:37,460
potential candidates for

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00:09:37,460 --> 00:09:38,960
future clinical
application

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00:09:38,960 --> 00:09:40,670
in image guided surgery.

305
00:09:40,670 --> 00:09:42,380
Research well
progress forward

306
00:09:42,380 --> 00:09:43,880
with systemic
administration in

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00:09:43,880 --> 00:09:46,250
rodents and eventually
larger mammals

308
00:09:46,250 --> 00:09:48,215
preceding that
clinical trials.

309
00:09:48,215 --> 00:09:49,730
Thank so much for

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00:09:49,730 --> 00:09:50,870
your attention and have

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00:09:50,870 --> 00:09:53,130
a wonderful afternoon.