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GUIDES IN BIOCHEMISTRY EXPERIMENTSSBK3013

2012/2013

DEPARTMENT OF BIOLOGY

DR SHAKINAZ DESA

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Laboratory safety will be e!lai"e# #$ri"% t&e first !ra'ti'al sessio"(

THE REPORT CONTENT

Your report M)Yo!"#"t o$

1% Pr&t#&' !o% (2% E)per#*e!t t#t'e + ,r#te & t#t'e t-&t pre"e!t .our e)per#*e!t $or e)&*p'e( T-e

eter*#!&t#o! o$ t,o prote#! "&*p'e u"#! B#uret &! Lo,r. &""&.

3% A"tr&t + "u**&r. o$ t-e e)per#*e!t ,-#- "-ou' #!'ue oet#4e" r#e$#!trout#o! r#e$ *et-o" &!&'.5e re"u't" o!'u"#o!%

6% I!trout#o! 7 -#-'#-t o! t-e *! "uet o$ #!tere"t8% M&ter#&'" &! *et-o" 7 o !ot op. $ro* t-e *&!u&'% 9r#te #! .our o,!

"e!te!e"% You *&. u"e #&r&*" $'o, -&rt et%

:% Re"u't" 7 &!&'.5e t-e &t& &! .ou *&. pre"e!t t-e re"u't" #! $or* o$ -&rt"t&'e $#ure &! p-oto%

;% D#"u""#o! 7 &!",er t-e /our!&' .e&r p&e !u*er?"@ &! pu'#"-er%

A'' report" *u"t e "u*#tte #! t,o ,ee>" &$ter t-e pr&t#&'% Your #!"trutor ,#''

#!$or* .ou t-e "u#t&'e ue &te%

G$i#e Title

* )'i# Base E!eri+e"t

, Protei" assay

- E".y+e e!eri+e"t

/ E!eri+e"t o" 0ita+i" C

1 E!eri+e"t $si"% li!i#

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GUIDE *2 )CID B)SE EXPERIMENT

*( )'i# Base Titratio"

9e&> #" #$$ere!t $ro* "tro! e&u"e #t &!!ot #""o#&te

o*p'ete'. #! ,&ter% 9e&> "u- &" &et# &! or&!# &! e

p&rt#&''. #""o#&te% T-ere$ore t-ere #" o!'. "*&'' &*ou!t o$ ,e&>

*o'eu'e" #""o#&te ,-#'e #! "tro! &'' *o'eu'e" e)#"t &" H

&! A+ #! & "o'ut#o!% Due to t-#" propert. H

o!e!tr&t#o! #! ,e&>

epe!" o! t-e oe$$##e!t o$ e

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Met&o#s

1% F#'' #! 0%1M N&OH #! & urette2% T#tr&te 28 *' o$ t-e t-ree " "ep&r&te'. ,#t- N&OH%3% Me&"ure t-e pH e4er. t#*e .ou & 1 *' o$ N&OH%6% Reor .our $#!#!"%

Data a"alysis

1% P'ot & r&p- o$ pH 4er"u" 4o'u*e o$ &"e% D#"u"" t-e r&p-%2% Deter*#!e t-e pK& $or e&- $ro* t-e r&p-3% Ho, o .ou eter*#!e t-e pK&6% 9-. *u"t .ou -oo"e t-e po#!t o$ #!$'et#o! &t t-e r&p-8% 9-. #" t-e r&p- #$$ere!t $or e&-

,( )!!li'atio"2 Ma4i"% !H i"#i'ator

T-#" e)per#*e!t e)p'ore" t-e e)tr&t#o! o$ !&tur&' #!#&tor" $ro* o**o!

$'o,er" $ru#t" &! 4eet&'e" &! t-e pH &t ,-#- t-e"e !&tur&' #!#&tor"-&!e o'or% So*e #!#&tor "o'ut#o!" &! p&per" ,#'' #!#&te ot- &!

&! & &"e ,-#'e ot-er" &re "pe#$# to u"t o!e% I! t-#" '& .ou ,#'' e

*&>#! & te"t p&per t-&t ,#'' #!#&te t-e pre"e!e o$ or &"e% It ,or>"

o!'. ,-e! #t #" ,et ,#t- H2O% You &! *&>e .our o,! "&'e . #pp#! t-e

p&per #!to >!o,! pH "o'ut#o!% You &! u"e t-e o'or "&'e to o*p&re%

Materials

#% 0%1 M H' "o'ut#o! ?& =%3 *L o$ o!e!tr&te H' "o'ut#o! toe!ou- e#o!#5e or #"t#''e ,&ter to *&>e 1%0 L o$ "o'ut#o!@

##% 0%1 M N&OH "o'ut#o! ?#""o'4e 6%0 N&OH #! e!ou- e#o!#5e or#"t#''e ,&ter to *&>e 1%0 L o$ "o'ut#o!@

###% 2+prop&!o' ?#"oprop.' &'o-o'@ /&eto!e /e#o!#5e or #"t#''e ,&ter#4% Fru#t"/4eet&'e"/$'o,er" ?&r> o'or" &re pre$erre@%

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2:

Ca$tio" s&o$l# be $se# w&e" wor4i"% wit& sol$tio"s of &y#ro'&lori' a'i#

a"# so#i$+ &y#roi#e( Bot& 'a" be irritati"% to t&e s4i"( Go%%les +$st bewor" t&ro$%&o$t t&e e!eri+e"t( If it is "e'essary to &eat t&e etra't i"

or#er to 'o"'e"trate t&e i"#i'ator5 #o "ot &eat t&ose 'o"tai"i"% t&e

fla++able sol0e"ts5 ,6!ro!a"ol or a'eto"e5 o0er a" o!e" fla+e( Use a

water bat& or &ot !late(

Etra'ti"% t&e i"#i'ator

1% Se'et p'&!t p&rt" t-&t &re *o"t p#*e!te%For $'o,er" o!'. & $e, pet&'" &re !or*&''. !eee% For $ru#t" &!

4eet&'e" $#!e'. -oppe p#ee" "-ou' e u"e% "e o!'. t-&t port#o!

o$ t-e $ru#t or 4eet&'e t-&t #" *o"t p#*e!te%

2% A &out 10 *L o$ "o'4e!t &! *&er&te%3% You *&. 'e&4e #t o4er!#-t to e!-&!e t-e e)tr&t#o! #$ !ee""&r.%Sue"te "o'4e!t"( ,&ter 2+prop&!o' 80+80 ,&ter/2+prop&!o' *#)ture

or &eto!e% O!e o$ t-e"e "-ou' e&"#'. e)tr&t t-e o'or% I$ t-e e)tr&t"

&re 4er. #'ute t-e o'or &! e o!e!tr&te . -e&t#! t-e ope!e

& #! & ,&r* ,&ter &t- or . pour#! t-e e)tr&t #! & e&>er &!

e4&por&t#! o! & -ot p'&te%

6% F#'ter &! o''et t-e $#'tr&te $ro* t-e *&er&te p'&!t "&*p'e"%

Testi"% t&e !H ra"%e of t&e i"#i'ator

1% L&e' 13 te"t tue" $ro* 1 to 13%2% P'&e C%0 *L o$ #"t#''e or e#o!#5e ,&ter #! &'' te"t tue" e)ept 1

&! 13%

3% Prep&re "o'ut#o!" #! t-e r&!e #! t-e $o''o,#! *&!!er(&% P'&e 10%0 *L o$ 0%1 M H' #! te"t tue 1% ?pH 1@

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% Tr&!"$er 1%0 *L o$ 0%1 M $ro* te"t tue 1 to te"t tue 2&! *#) t-orou-'.% ?pH 2@

% Tr&!"$er 1%0 *L o$ "o'ut#o! $ro* te"t tue 2 to te"t tue3 &! *#) t-orou-'.% ?pH 3@

% o!t#!ue *&>#! t-e "er#&' #'ut#o!" . tr&!"$err#! 1%0 *L o$t-e *o"t ree!t'. #'ute "o'ut#o! to t-e !e)t te"t tue

u!t#' "#) "o'ut#o!" o$ pH 1 to : -&4e ee! prep&re% Be

"ure to *#) e&- t-orou-'. e$ore t-e tr&!"$er%

6% A 10%0 *L #"t#''e or e#o!#5e ,&ter to te"t tue ;% ?pH ;@8% Prep&re "o'ut#o!" o$ &"e #! t-e $o''o,#! *&!!er(

&% P'&e 10%0 *L o$ 0%1 M N&OH #! te"t tue 13% ?pH 13@% Tr&!"$er 1%0 *L o$ 0%1 M N&OH $ro* te"t tue 13 to te"t tue

12 &! *#) t-orou-'.% ?pH 12@

% o!t#!ue *&>#! "er#&' #'ut#o!" o$ t-e &"e o#! $ro* pH 12o,! to pH = . tr&!"$err#! 1%0 *L o$ t-e *o"t ree!t'.

#'ute &"# "o'ut#o! to t-e !e)t te"t tue &! *#)#!

t-orou-'. e&- t#*e%

:% L&e' t-e ,e''" o$ & "pot p'&te $ro* 1 to 13% Tr&!"$er & $e, rop" o$e&- o$ t-e "o'ut#o!" prep&re #! "tep" 3 6 &! 8 to t-e

orre"po!#! ,e'' #! t-e "pot p'&te%

;% A & rop or t,o o$ t-e $'o,er/$ru#t/4eet&'e e)tr&t #!#&tor toe&- ,e''% O"er4e t-e pH &t ,-#- t-e #!#&tor -&!e" o'or%

Testi"% t&e !H of ot&er li7$i#s

O!e t-e pH r&!e" o$ t-e #!#&tor" -&4e ee! eter*#!e t-e. &! e

u"e #! +&"e t#tr&t#o!" or to te"t t-e pH o$ -ou"e-o' -e*#&'"%

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GUIDE ,2 PROTEIN EXPERIMENT

*( Protei" )ssays

T-#" e)er#"e #!troue" "tue!t" to *et-o" o$ eter*#!#! prote#!

o!e!tr&t#o!"( &"or&!e &t 860 !* $or t-e B#uret &! ;80 !* $or t-e

Lo,r. &""&."%

Prep&re "o'ut#o!" o$ e'&t#! &t 1 2 3 6 8 &! : */*L #! ,&ter $or

B#uret &""&.% Prep&re 0%1 0%2 0%3 0%6 0%8 &! 0%: */L $or Lo,r. *et-o%

Pro'e#$re2

a8 Bi$ret assay2M#) 0%80 *L o$ prote#! ,#t- 2%80 *L o$ B#uret re&e!t% Me&"ure t-e

&"or&!e &t 860 !* &$ter 10 *#!ute"%

B#uret re&e!t(

i. Add, with stirring, 300 mL of 10% (w/v) NaOH to 500 mL of a soltion!ontaining 0.3% !o""#r slfat# "#ntah$drat# and 1.% sodim

"otassim tartarat#, th#n dilt# to on# lit#r.

ii. &h# r#ag#nt is sta'l# for a f#w months 't not a $#ar.iii. Adding on# gram of "otassim iodid# "#r lit#r and storing in th# dar

ma#s it last ind#finit#l$.

)1/92Me&"ure t-e &"or&!e o$ 1 *L &t 860 !*% P'ot t-e "t&!&r ur4e%

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GUIDE -2 EXPERIMENT ON EN;YME

E".y+e "o'ut#o!%

2% Io#!e "o'ut#o! #" prep&re . ! 8 pot&""#u* #o#e to 100 *',&ter% T-e #""o'4e pot&""#u* #o#e #" &e ,#t- 1 o$ #o#!e &!

#" &''o,e to #""o'4e%3% Prep&re & "t&!&r ur4e o$ A"or&!e ? 8C0 !*@ 4"%

o!e!tr&t#o! o$ & "t&r-/#o#!e *#)ture% "e t-e $o''o,#! t&'e &"

u#e%

Table *Pre!aratio" of star'&=io#i"e +it$re for sta"#ar# '$r0e #eter+i"atio"

Test

t$be

> +l star'&

of +%=+l

?ater

@+l8

Io#i"e

@+l8

Me&"ure t-e

&"or&!e

&t 8C0!*

1 0 C 1

2 0%01 1 1

3 0%028 1 1

6 0%08 1 1

8 0%1 1 1

: 0%3 1 1

; 0%8 1 1

= 0%; 1 1

C 1%0 1 1

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,( T&e effe't of s$bstrate 'o"'e"tratio"

E)per#*e!t o$ "t&r- -.ro'."#" #! #$$ere!t "u"tr&te o!e!tr&t#o!

e)per#*e!t *u"t e prep&re &" t-e $o''o,#! t&'e%

:$estio"2 ?&at will yo$ $se to !re!are t&e bla"4=refere"'eA

Table ,

Data a"alysis

1% &'u'&te "t&r- o!e!tr&t#o! $or e&- "&*p'e &$ter -.ro'."#" ?SF@t-rou- u"e o$ t-e "t&!&r ur4e% T-e #!#t#&' "t&r- o!e!tr&t#o! ?S0@

#" &'re&. >!o,!% S ?So@ 7 ?SF@

2% T-e 4e'o#t. ?r&te o$ #e"t#o!@ o$ t-e re&t#o! $or e&- "&*p'e &! e&'u'&te &"(

S= t @S96 S8 = *9 +i"$tes

3% Prep&re & t&'e "-o,#! r&te o$ -.ro'."#" ?@ &t #$$ere!t t-e "t&r-o!e!tr&t#o!"%

6% P'ot & M#-&e'#"+Me!te! r&p-%8% Prep&re & r&p- o$ 1/"t&r- o!e!tr&t#o! ?)+&)#"@ 4er"u" 1/r&te o$

#e"t#o! ?.+&)#"@% T-#" t.pe o$ re#pro&' r&p- #"p'&.#! e!5.*e

>#!et#" #" & L#!e,e&4er+Bur>e p'ot%

:% St&te t-e 4&'ue o$ *&)&! M#-&e'#" o!"t&!t K*$ro* .our r&p-%

Test

t$be

> +l star'&

ofx+%=+l

?ater

@+l8

)+ylase

@+l8

Io#i"e

@+l81 0 = 1 1

2 0%01 0 1 1 P'&e &'' te"t

3 0%028 0 1 I!u&te 1 tue" #! &!

6 0%08 0 1 e&- 1 #e &t-%

8 0%1 0 1 "&*p'e 1 Me&"ure t-e

: 0%3 0 1 &t 38 $or 1 &"or&!e &t

; 0%8 0 1 10 *#!ute" 1 8C0!*

= 0%; 0 1 1

C 1%0 0 1 1

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T-e .+#!terept o$ t-e L#!e+,e&4er Bur>e p'ot #" t-e re#pro&' o$ t-e

*&)#*u* 4e'o#t. o$ t-e re&t#o! ?*&)@% T-e )+#!terept #" t-e !e&t#4e

re#pro&' o$ t-e M#-&e'#" o!"t&!t% ?K*@%

-( T&e effe't of te+!erat$re

Prep&re &" t-e $o''o,#! $or t-e e)per#*e!t o$ #$$ere!t te*per&ture%

Table -

Data a"alysis

P'ot t-e L#!e,e&4er+Bur>e '#!e $or t-e re"u't o$ 20 2= 38 &! 60%

o*p&re &'' t-ree p'ot"%

9-&t &re t-e 4&'ue" o$ *&)&! K*$or &'' p'ot"

Test

t$be

> +l star'&

of +%=+l

?ater

@+l8

)+ylase

@+l8

Io#i"e

@+l8

1 0 = 1 1

2 0%01 0 1 I!u&te 1 P'&e &'' te"t

3 0%028 0 1 e&- 1 tue" #! &! #e

6 0%08 0 1 "&*p'e 1 &t-%

8 0%1 0 1 &t 20 2= 1 Me&"ure t-e

: 0%3 0 1 3860 $or 1 &"or&!e &t

; 0%8 0 1 10 *#!ute" 1 8C0!*

= 0%; 0 1 1

C 1%0 0 1 1

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/( T&e effe't of !H

Prep&re t-e $o''o,#! $or t-e e)per#*e!t u"#! #$$ere!t pH%

(CAUTION: DO NOT ADD THE ENZYME BEFORE ADDING BUFFER pH)

Table /

Data a"alysis

9-&t &re t-e 4&'ue" o$ $or &'' pH

o*p&re t-e 4e'o#t. $or e&- o$ t-e pH te"t

Te"t

tue

St&r-

0%8 */*'

2 *'

u$$er o$

pH*

A*.'&"e

?*'@

Io#!e

?*'@

1 8*' 6 1*' I!u&te 1 P'&e &'' te"t tue"2 8 8 1 e&- "&*p'e 1 #! &! #e &t-%

3 8 : 1 &t 38 1 Me&"ure t-e

6 8 ; 1 $or 10 1 A"or&!e &t

8 8 = 1 *#!ute" 1 8C0!*

: 8 C 1 1

; 8 10 1 1

'&!> 8 3 *' o$ H2O 1

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GUIDE /2 EXPERIMENT ON IT)MIN C

*( Meas$ri"% ita+i" C $si"% star'&6io#i"e test

9e ,#'' *e&"ure t-e &*ou!t o$ 4#t&*#! #! *&!. #$$ere!t t.pe" o$ $oo"%

T-e -e*#&' re&t#o! ,e ,#'' u"e to *e&"ure t-e &*ou!t o$ 4#t&*#! u"e"

o!e o$ #t" $u!t#o!" #! t-e o.% #t&*#! #!4o'4e" #! our e''" o)#&t#o!+

reut#o! re&t#o!"% #t&*#! &! re&t ,#t- #o#!e% T-ere$ore ,e ,#''

*e&"ure t-e &*ou!t o$ 4#t&*#! . ! #o#!e to our $oo e)tr&t"

u!t#' t-e 4#t&*#! &! #! !o *ore #o#!e%

Io#!e #! e)e"" o$ t-e 4#t&*#! ,#'' re&t ,#t- & "t&r- "o'ut#o! .ou

,#'' & to t-e e)tr&t to proue & 'u#"-+'&> o'or% T-e t#o! o$ &

-e*#&' to *e&"ure &!ot-er -e*#&' #" &''e & t#tr&t#o!%

Materials2

1% oo# so$r'es of 0ita+i" C( $or e)&*p'e u#e" e)tr&t#o! o$ p'&!t"$'o,er" $ru#t" r!" &! 4eet&'e" 4#t&*#! t&'et or

oo>e/tre&te $oo "&*p'e ?o#'e/re$r#er&te/r#''e@

2% Star'& sol$tio" @*F82M#) 1 "t&r- #! 100 *' o#'#! H2O% Bo#' $or o!e

*#!ute ,-#'e "t#rr#!% St#r u!t#' o*p'ete'. #""o'4e ?t-#" "o'ut#o! ,#''

e 'ou.@%

3% Io#i"e sol$tio"( M#) 0%: pot&""#u* #o#e #! 800 *' H2O% M#) 0%:

#o#!e #! 80 *' o$ et-.' &'o-o'% T-e"e t,o #o#!e "o'ut#o!" "-ou' e*#)e ,e'' e$ore o*#!#!% o*#!e t-e t,o #o#!e "o'ut#o!" &!

& &! t#o!&' 680 *' o$ H2O%

6% H.ro-'or# A# ?H'@ 1 M ?8 *'@

8% B'e!er

:% F#'ter/ -ee"e'ot-

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Pro'e#$re2

1% Prep&r#! t-e 4#t&*#! e)tr&t"(

#% -op $oo *&ter#&' #!to "*&'' p#ee" &! p'&e #!to'e!er%

##% A 100 *' o$ #"t#''e ,&ter to t-e 'e!er%###% B'e! u"#! t-e -#-e"t "pee u!t#' t-e *&ter#&' #"

t-orou-'. rou!%

#4% Str! t-e rou! e)tr&t4% Me&"ure 30 *' o$ t-e "tr!e e)tr&t #!to & 280 *'

Er'e!*e.er $'&"> or e&>er%

2% Me&"ur#! 4#t&*#! #! t-e $oo "&*p'e(

#% P'&e 30 *L o$ t-e $oo e)tr&t" "o'ut#o! #! & 280 *' $'&">or e&>er%

##% A 2 rop" o$ t-e 0%1 M H' to t-e $'&">%###% A 8 *' o$ t-e "t&r- "o'ut#o! to t-e $'&">%#4% F#'' & urette ,#t- t-e #o#!e "o'ut#o!%4% Reor t-e #!#t#&' 4o'u*e re!%

4#% A t-e #o#!e "o'ut#o! #! 1 *' #!re*e!t" to t-e $'&">,-#'e ",#r'#! t-e $'&">%

4##% A #o#!e u!t#' t-e "o'ut#o! "t&." 'ue+'&> $or 18"eo!"%

4###% Reor t-e 4o'u*e re! o! t-e urette%3% o*p&r#! oo>e $oo &! r&, $oo" 4#t&*#!

Doe" t-e ,&. .ou prep&re .our $oo &$$et t-e 4#t&*#! &4'&'e to

e #!e"te #t&*#! #" & ,&ter+"o'u'e 4#t&*#!% 9ou' oo>#!$oo . o#'#! #! ,&ter &$$et t-e 4#t&*#! o!te!t I$ 4#t&*#! #"

'o"t ur#! t-e oo>#! proe"" ,-ere oe" #t o 9-&t t.pe" o$

e)per#*e!t" ou' .ou e"#! to te"t .our -.pot-e"#" You ,#'' e

te"t#! .our -.pot-e"#" to eter*#!e #$ 4#t&*#! o!te!t #" -&!e

ur#! oo>#! or #$ #$$ere!t ,&." o$ $oo prep&r&t#o! .#e' #$$ere!t

&*ou!t" o$ 4#t&*#! %

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3:

#% Foo &! e prep&re &or#! to .our re&t#4#t.% Fore)&*p'e .ou &! o#' or "te&* or p'&e #! & $ree5er% You

&! &'"o prep&re t-e $oo . #$$ere!t e)po"#! t#*e to-e&t et%

##% -op $oo *&ter#&' #!to "*&'' p#ee" &! p'&e #!to'e!er%

###% Ot! .our &t& u"#! t-e "&*e *et-o #! pre4#ou""et#o!%

#4% Reor t-e 4o'u*e re! o! t-e urette%4% o*p&re t-e re'&t#4e &*ou!t" o$ &"or# pre"e!t #!

t-e "&*p'e" .ou &re te"t#!%

4#% o*p&re .our re"u't" ,#t- t-o"e o$ ot-er *e*er" o$ t-e'&""% 9-&t o t-e re"u't" "-o,

:$estio"s2

#% 9-&t u#e" or r#!>" -& t-e *o"t 4#t&*#! ##% D# t-e r#!>" -&4e t-e 4#t&*#! t-&t t-e. &4ert#"e o!

t-e '&e'"

###% 9-&t $oo "oure" -& t-e *o"t 4#t&*#! #4% 9-&t $&*#'#e" or roup" -& t-e *o"t 4#t&*#! 4% D# p'&!t" t-&t .ou o !ot !or*&''. e&t -&4e 4#t&*#!

4#% D# -e&t &$$et t-e 4#t&*#! o!te!t o$ $oo4##% D# -e&t #!re&"e or ere&"e t-e 4#t&*#! 'e4e'"

4###% 9-&t ,&. o$ $oo prep&r&t#o! ,ou' e t-e *o"t!utr#t#ou"

#)% Do .ou -&4e &!. #e&" !o, to et *ore 4#t&*#!" $ro* .our*e&'"

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,( )!!li'atio"2 Ma%i' ?riti"%

Materials

Be&>er

Io#!e

Le*o!/L#*e u#e

Noteoo> p&per

up

Art ru"-

Pro'e#$re2

STEP A( IODINE SOLTION

1% Pour 100 *' ,&ter #!to & 800*'+e&>er%2% A 10 *' o$ Io#!e to t-e ,&ter &! "t#r%

STEP B(

1% ut & "et#o! $ro* t-e !oteoo> p&per%2% T-e p&per *u"t $#t #!"#e & 800*'+e&>er

STEP ( ITAMIN SOLTION

1% SerSTEP D(

1% D#p t-e &rt ru"- #!to t-e 'e*o!/'#*e u#e2% 9r#te & *e""&e o! t-e p#ee o$ p&per%3% A''o, t-e u#e to r. o! t-e p&per%6% Su*er"e t-e p&per #! t-e #o#!e "o'ut#o! #! t-e o,'%

RESULTS

O"er4e .our $#!#!"% 9-. o .ou et t-e re"u't

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GUIDE 12 EXPERIMENT USING LIPID

*( Sa!o"ifi'atio" of tri%ly'eri#es

Tr#'.er#e" &re o*po"e o$ t-ree $&tt. " '#!>e to '.ero' . $&tt.

&.' e"ter" ?+O+O+R@% T-e $&tt. " *&. e "&tur&te ?!o ou'e

o!"@ or u!"&tur&te% L#

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3C

Materials2

Tr#'.er#e "&*p'e( e%% oo!ut o#' or! o#' p&'* o#' *&r&r#!e utter

So'4e!t ?1(1 et-&!o'/et-er@

0%8M KOH

P-e!o'p-t-&'e#!

0%8M HL

?)RNING2 er twi!# ,#t- 1 *' o$ "o'4e!t ?1(1 et-&!o'/et-er@ to o''et &''

re"#u&' *&ter#&'% A t-e ,&"- to t-e #"t#''&t#o! $'&">%3% A 28 *' o$ 0%8M KOH /et-&!o' "o'ut#o!6% Me&"ure t-e e)&t 4o'u*e o$ .our *#)ture% Set up & "eo! "."te*

&" & o!tro' ?or re$ere!e@ ,#t- 28 *' o$ t-e 0%8M KOH/et-&!o'

"o'ut#o! p'u" t#o!&' 1(1 et-&!o'/et-er "o'4e!t $or & $#!&' 4o'u*e

#e!t#&' to .our e)per#*e!t&' "o'ut#o!%

8% Set up & re$'u) o!e!"er o! e&- $'&"> &! p'&e #! o#'#! ,&ter&t- $or 30 *#!ute"% T-e -.ro'."#" ,#'' our ur#! t-#" per#o%

:% A''o, $'&">" to oo'% A t-ree rop" o$ #!#&tor "o'ut#o!?p-e!o'p-t-&'e#! 10 /L@ to ot- $'&">" &! t#tr&te ,#t- 0%8M H'

"o'ut#o!%

;% T-e *o'&r #$$ere!e et,ee! t-e &*ou!t o$ 0%8M H' re

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=% &'u'&te t-e ,e#-t ?*@ o$ KOH u"e to "&po!#$. t-e 1 "&*p'e%Deter*#!e t-e "&po!#$#&t#o! !u*er $or te"te "&*p'e"

C% Ot! t-e $#!&' re"u't" $ro* t-e ot-er roup" $or e&- "&*p'e &!prep&re & t&'e to "u**&r#5e t-e re"u't"%

"e t-e $o''o,#! t&'e to &""#"t .ou to &'u'&te t-e "&po!#$#&t#o! !u*er

Sa+!le I"itial

0ol$+e

of HCl

Bla"4

i"al

ol$+e

of HCl

Bla"4

I"itial

0ol$+e

of HCl

Sa+!le

i"al

ol$+e

of HCl

Sa+!le

Mol for

bla"4

@Molarity

ol$+e

HCl

bla"48

Mol for

sa+!le

@Molarity

ol$+e

HCl

sa+!le8

Mol of

rea'te#

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,( )!!li'atio"2 Ma4i"% Soa!

C)UTION2 Safety %o%%les +$st be war"e# t&ro$%&o$t t&is e!eri+e"t(T&is lab is i"te"#e# to be #o"e i" a s'&ool lab5 wit& a#$lt s$!er0isio"(

So#i$+ &y#roi#e is 0ery 'a$sti'5 a"# 'a" 'a$se se0ere b$r"s to t&e

s4i"5 es!e'ially w&e" &ot(

Materials2

=0 *' o$ : M N&OH "o'ut#o!

18 r&*" o$ $&t

;8 *' o$ #"t#''e ,&ter

300 *' -ot "o#u* -'or#e "o'ut#o!

100+*' r&u&te .'#!er

St#rr#! ro

600+*' e&>er

280+*' e&>er

To *&>e : *o'&r "o#u* -.ro)#e #""o'4e *K(, %ra+so$ N&OH #!

e!ou- ,&ter to *&>e & tot&' 4o'u*e o$ =0 *'%

T-#" #" u"t & "&tur&te "o'ut#o! o$ N&'%

Met&o#s2

1% Ot! /9 +lo$ : *o'&r N&OH &! *(1 %ra+so$ $&t2% P'&e /9 +lo$ t-e N&OH "o'ut#o! &! t-e $&t #! & 280+*' e&>er%3% He&t to o#' o4er t-e 'o,e"t $'&*e t-&t ,#'' "u"t! t-e o#'#! proe""%

St#r t-e *#)ture o!"t&!t'. to &4o# "p&tter#!% I$ "p&tter#! our"

re*o4e t-e $'&*e &! o!t#!ue "t#rr#! t-e *#)ture% Rep'&e t-e $'&*e&! o!t#!ue -e&t#! &$ter t-e "p&tter#! "top"%

6% o!t#!ue o#'#! &! "t#rr#! $or &out 20 *#!ute" or u!t#' #t &ppe&r"t-&t *o"t o$ t-e ,&ter -&" ee! e4&por&te%

8% T-e! &re$u''. & t-e re*!#! ,9 +lo$ N&OH "o'ut#o! &!o!t#!ue o#'#! $or &! t#o!&' 20 *#!ute" or u!t#' *o"t o$ t-e

,&ter -&" o#'e o$$% DO NOT LET IT BOIL DRY(

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:% A" t-e rue "o&p oo'" & ,&). "o'# "-ou' $or*% A to #t &out*,(1 +lo$ #"t#''e ,&ter &! &out 19 +lo$ -ot "&tur&te "o#u*

-'or#e "o'ut#o!%;% St#r t-e *#)ture re&>#! up 'u*p" ,#t- .our "t#rr#! ro%=% De&!t t-e ,&"- "o'ut#o! . pour#! #t t-rou- & ,#re "ree! ,-#-

,#'' tr&p "*&'' "o&p p&rt#'e"%

K( Repe&t t-e ,&"- proe"" t,#e% A$ter t-e $#!&' ,&"-#! pre"" t-e "o&pet,ee! t,o "-eet" o$ p&per to,e''#! to e)pe' &" *u- ,&ter &"

po""#'e

Que"t#o!"(

1% 9-&t #" t-e re'&t#o!"-#p et,ee! "&po!#$#&t#o! &! p-&"e ?'#