Attachment  
 
Iso-α-bitter Acids  
イソアルファー苦味酸  
 
Compositional Specifications  
Definition  Iso-α-bitter Acids are obtained by the flowers  of the hops  (Humulus lupulus  L.). They  
consist mainly of isohumulones.  
Content   Iso-α-bitter Acids contain not less than 20.0% of iso-α-bitter acids. 
Description   Iso-α-bitter Acids occur as yellow -brown liquids. They  have a characteristic odor  and 
a strongly  bitter taste . 
Identification  
Analyze the test solution  and the standard solution prepared as directed in the Assay by liquid 
chromatography using  the operating conditions specified in the Assay. Three of the peaks from the 
test solution correspond to the retention times of main peaks from the standard solution, respectively.  
Purity  
(1) Lead   Not more than 2 μg/g as Pb ( 2.0 g, Method 1, Control Solution : Lead Standard Solution 
4.0 mL, Flame Method).  
(2) Arsenic   Not more than 1.5 μg/g as As (1.0 g, Method 4, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Assay  
Test Solution   Weigh accurately about  0.1 g of the sample , and dissolve it in a 500 : 1 mixture 
of methanol/ phosphoric acid TS  (0.1 mol/L) to make  exactly  100 mL. To exactly 10 mL of this solution , 
add a 500 : 1 mixture of methanol/ phosphoric acid TS (0.1 mol/L) to make exactly  50 mL. If the 
solution  is turbid,  filter it through with a  membrane filter (0.45 μm pore size) . 
Standard Solution   Weigh accurately about  50 mg of iso-α-bitter acids for assay.  Add a 500 : 1 
mixture of methanol/ phosphoric acid TS  (0.1 mol/L)  to make  exactly  100 mL. To exactly 10 mL of 
this solution , add a 500 : 1 mixture of methanol/ phosphoric acid TS (0.1  mol/L)  to make exactly  50 
mL. 
Procedure   Analyze 10  μL each  of the test solution and the standard solution  by gas 
chromatography using the operating conditions given belo w. For the standard solution, three main 
peaks appear in the order of isocohumulone , isohumulone , and isoadhumulone. Sum  up the peak 
areas of all the peaks  between  the retention time s of isocohumulone and  isoadhumulone  in the test 
solution, and d etermine the content  of iso-α-bitter acids by the formula:  Content  (%) of iso-𝛼-bitter  acids  = a ×b × AA
M × AS×1000   
a = weight  (mg) of iso-α-bitter acids for assay . 
b = content ( %) of iso-α-bitter acids in iso-α-bitter acids for assay . 
AA = total  area of all the peaks between  the retention time s of isocohumulone and 
isoadhumulone in the test solution.  
AS = total area of  the three main  peaks  from  the standard solution.  
M = weight  (g) of the sample.  
 
Operating conditions  
Detector: Ultraviolet spectrophotometer (wavelength: 2 70 nm).  
Column: A stainless steel tube (4.6 mm internal diameter and 25 cm length).  
Column packing material: 5 -μm octadecylsilanized silica gel for liquid chromatography.  
Column temperature: 35 C. 
Mobile phase: A 75 : 24 : 1 mixture of methanol /water /phosphoric acid . 
Flow rate:  1 mL/min. 
 
Reagents, Solutions, and Other Reference Materials  
Iso-α-bitter Acids  for Assay  An international calibration standard (DCHA -Iso) with a known 
concentration.  It is a mixture of isohumulone , isoadhumulone , and isocohumulone and their 
isomers.  Use the content  (%) of the total iso-α-bitter acids as the content  (%) of iso-α-bitter acids. 
Phosphoric Acid TS (0.1 mol/L)  Add water to 11.5 g of phosphoric acid to make 1000  mL. 
 
  Higher Fatty Acid ( Caprylic Acid)  
高級脂肪酸（カプリル酸）  
 
Compositional Sp ecifications  
 
Substance  Name   Higher Fatty Acid ( Caprylic Acid) 
Definition   Higher Fatty Acid ( Caprylic Acid)  is one of the higher fatty acids                                                                                                                                                                                
(substances that are obtained by hydrolyzing animal or vegetable fats or oils or hydrogenated animal 
or vegetable fats or oils).  It con sists mainly of caprylic acid.  
Content  Higher Fatty Acid ( Caprylic Acid)  contains not less than 50.0% of caprylic acid (C 8H16O2 
= 144.21).  
Description   Higher Fatty Acid ( Caprylic Acid)  occurs as a col orless to light yellow liquid or a white 
to bright grayish yellow paste.  
Identification  Analyze the test solution and the standard solution prepared as directed in the Assay 
by gas chromatography using the operating conditions specified  in the Assay. The retention time of 
the main peak from the test solution corresponds to that of the peak of methyl caprylate from  the 
standard solution.  
Iodin e Value   Not more than 0.5.  
Purity   
(1) Acid value   380−395.  
(2) Lead   Not more than 2 μg/g as Pb (2.0 g, Method 1, Control Solution: Lead Standard Solution 
4.0 mL, Flame Method).   
(3) Arsenic   Not more than 3 μg/g as As  (0.50 g, Method 3, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Residue on Ignition  Not more than 0.1%.  
Assay   
Test Solution  Weigh 20 mg of Higher Fatty Acid ( Caprylic Acid) , and transfer it into a small 
conical flask fitted with a reflux condenser. Add 5 mL of boron trifluoride−methanol TS, shake, and 
heat for about 10 minutes until it  dissolves. Add through the condenser 4 mL of hexane, and heat 
for 10 minutes. After cooling, add 20 mL of a saturate solution of sodium chloride, shake, and leave 
to stand to allow the solution to separate into two layers. Transfer 2 mL of the separated h exane 
layer into another flask through about 0.2 g of sodium sulfate previously washed with hexane. To 1 
mL of this solution, add h exane to make 10 mL, and shake.  
Standard Solution   Dissolve 10 mg of methyl caprylate by a dding 5 mL of hexane . 
Procedure   Analyze  1 L each  of the test solution and the standard solution by gas chromatography using the operating conditions given below. Determine the peak area (A A) of methyl 
caprylate and the total peak area (A T) of all the fatty acid esters (total peak area of all peaks detected)  
in the test solution,  and calculate the cap rylic acid content in the fatty acid fraction of the sample by 
the following formula. Identification of methyl caprylate should be confirmed by comparing the 
retention time of the main peak from the test solution with that of methyl caprylate in the standard 
solution . The chromatography should be continued for 1.5 times the retention time of methyl 
caprylate, and the main solvent peak should be excluded from the measurement.  
             Cont ent (%) of caprylic  acid = AA
AT ×100 
Operating conditions  
Detector: Flam e ionization detector . 
Column: A fused silica tube (0.25 mm internal diameter and 50 m length) coated with a 0.2 -
m thick layer of 100% cyanopropylpolysilo xane for gas chromatography.  
Column temperature: 180°C.  
Injection port temperature: 250°C.  
Detector temperature: 250°C.  
Carrier gas: Helium.  
Flow rate: A constant rate of about 1.0 mL/min.  
    Injection method: Splitless.  
 
Reagents, Solutions, and Other Reference Materials  
Methyl Caprylate   See Methyl Octanate . 
Methyl Octanate   C9H18O2  [111-11-5]  A colorless , clear liquid . 
Refractive index  nD20: 1.415 −1.420 . 
Density  0.874−0.880 g/mL (20°C).  
 
  Higher Fatty Acid ( Capric Acid)  
高級脂肪酸（カ プリン酸）  
 
Compositional Sp ecifications  
 
Substance  Name   Higher Fatty Acid ( Capric Acid) 
Definition   Higher Fatty Acid ( Capric Acid)  is one of the higher fatty acids  (substances that are 
obtained by hydrolyzing animal or vegetable fats or oils or hydrogenated animal or vegetable fats or 
oils ).  It consist s mainly of capric acid.  
Content  Higher Fatty Acid ( Capric Acid)  contains not less than 50.0% of capric acid (C 10H20O2 = 
172.26).  
Description   Higher Fatty Acid ( Capric Acid)  occurs as white to bright grayish yellow flakes, powder, 
or granules or as a waxy lump.  
Identification  Analyze the test solution and the standard solution prepared as directed in the Assay 
by gas chromatography usin g the operating conditions specified  in the Assay. The retention time of 
the main peak from the test solution corresp onds to that of the peak of methyl caprate from the 
standard solution.  
Iodin e Value   Not more than 0.5.  
Purity  
(1) Acid value   321−333.  
(2) Lead   Not more than 2 μg/g as Pb (2.0 g, Method 1, Control Solution: Lead Standard Solution 
4.0 mL, Flame Method).  
(3) Arsenic   Not more than 3 μg/g as As  (0.50 g, Method 3, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Residue on Ignition  Not more than 0.1%.  
Assay  
Test Solution  Weigh 20 mg of Higher Fatty Acid ( Capric Acid) , and transfer it into a small 
conical flask fitted with a reflux condenser. Add 5 mL of boron trifluoride−methanol TS, shake, and 
heat for about 10 minutes until it dissolves. Add through the condenser 4 mL of hexane, and heat 
for 10 minutes. After cooling, add 20 mL of a saturat e solution of sodium chloride, shake, and leave 
to stand to allow the solution to separate into two layers. Transfer 2 mL of the separated hexane 
layer into another flask through about 0.2 g of sodium sulfate previously washed with hexane. To 1 
mL of this solution, add h exane to make 10 mL, and shake.  
Standard Solution  Dissolve 10 mg of methyl caprate  by adding 5 mL of hexane.  
Procedure  Analyze 1  L each  of the test solution  and the standard solution by gas chromatography using the operating conditions gi ven below. Determine the peak area (A A) of methyl 
caprate and the total peak area (A T) of all the fatty acid esters (total peak area of all peaks detected)  
in the test solution, and  calculate the cap ric acid content in the fatty acid fraction of the sample by 
the following formula. Identification of methyl caprate should be confirmed by comparing the 
retention time of the main peak from the test solution with that of methyl caprate in the standard 
solution. The chromatography should be continued for 1.5 times the retention time of methyl caprate, 
and the main solvent peak should be excluded from the measurement.  
             Content  (%) of capric  acid = AA
AT ×100 
Operating conditions  
Detector: Flam e ionization detector . 
Column: A fused silica tube (0.25 mm internal diameter and 50 m length) coated with a 0.2 -
m thick layer of 100% cyanopropylpolysiloxane for gas chromatography.  
Column temperature: 180°C.  
Injection port temperature: 250°C.  
Detector temperature: 250°C.  
Carrier gas: Helium.  
Flow rate: A constant rate of about 1.0 mL/min.  
Injection method: Splitless.  
 
Reagents, Solutions, and Other Reference Materials  
Methyl Caprate   See Methyl Decan oate. 
Methyl Deca noate  C11H22O2  [110-42-9]  A colorl ess, clear liquid . 
Refractive index  nD20: 1.424−1.427 . 
Specific gravity  d2020: 0.872−0.876.  
 
  Higher Fatty Acid ( Stearic Acid)  
高級脂肪酸（ ステアリン酸）  
 
Compositional Sp ecifications  
 
Substance  Name   Higher Fatty Acid ( Stearic Acid) 
Definition   Higher Fatty Acid ( Stearic Acid)  is one of the higher fatty acids  (substances that are 
obtained by hydrolyzing animal or vegetable fats or oils or hydrogenated ani mal or vegetable fats or 
oils ) . It consists mainly of stearic acid.  
Content  Higher Fatty Acid ( stearic Acid)  contains not less than 50.0% of stearic acid (C 18H36O2 = 
284.48).  
Description   Higher Fatty Acid ( Stearic Acid)  occurs as white to bright grayish yellow flakes, 
powder,  or granules or as a waxy lump.  
Identification  Analyze the test solution and the stand ard solution prepared as directed in the Assay 
by gas chromatography usin g the operating conditions specified  in the Assay. The retention time of 
the main peak from the test solution corresponds to that of the peak of methyl stearate from the 
standard solu tion.  
Iodin e Value   Not more than 4.0.  
Test Solution   Weigh  accurately about 1 g of Higher Fatty Acid ( Stearic Acid) into a 500 -mL 
stoppered flask, and dissolve it in 20 mL of a 1 : 1 mixture of cyclohexane /chloroform.  
Procedure   Proceed as directed under  Iodine  Value in the Fats and Related Substances Tests.  
Purity  
(1) Acid value   194−210.  
(2) Lead   Not more than 2 μg/g as Pb (2.0 g, Method 1, Control Solution: Lead Standard Solution 
4.0 mL, Flame Method).   
(3) Arsenic   Not more than 3 μg/g as As  (0.50 g,  Method 3, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Residue on Ignition  Not more than 0.1%.  
Assay  
Test Solution  Weigh 20 mg of Higher Fatty Acid ( Stearic Acid) , and transfer it into a small 
conical flask fitted with a reflux condenser. Add 5 mL of boron trifluoride−methanol TS, shake, and 
heat for about 10 minutes until it dissolves. Add through the condenser 4 mL of hexane, and heat 
for 10 minutes. After cooling, add 20 mL of a saturate solution of sodium chloride, shake, and  leave 
to stand to allow the solution to separate into two layers. Transfer 2 mL of the separated hexane 
layer into another flask through about 0.2 g of sodium sulfate previously washed with hexane. To 1 mL of this solution, add hexane to make 10 mL, and s hake.  
Standard Solution  Dissolve 10 mg of methyl stearate by adding 5 mL of hexane.  
Procedure  Analyze 1  L each  of the test solution  and the standard solution by gas 
chromatography using the operating conditions given below. Determine the peak area (A A) of methyl 
stearate and the total peak area (A T) of all the fatty acid esters (total peak area of all peaks detected)  
in the test solution, and calculate the stearic acid content in the fatty acid fraction of the sample by 
the following formula.  Identificat ion of methyl stearate should be confirmed by comparing the 
retention time of the main peak from the test solution with that of methyl stearate in the standard 
solution. T he chromatography should be continued for 1.5 times the retention time of methyl 
stearate, and the main solvent peak should be excluded from the measurement.  
             Content  (%) of stearic  acid = AA
AT ×100 
Operating conditions  
Detector: Flam e ionization detector . 
Column: A fused silica tube (0.25 mm internal diameter and 50 m length)  coated with a 0.2 -
m thick layer of 100% cyanopropylpolysiloxane for gas chromatography.  
Column temperature: 180°C.  
Injection port temperature: 250°C.  
Detector temperature: 250°C.  
Carrier gas: Helium.  
Flow rate: A constant rate of about 1.0 mL/min.  
Injection method: Splitless.  
  Higher Fatty Acid ( Palmitic Acid)  
高級脂肪酸（ パルミチン酸）  
 
Compositional Sp ecifications  
 
Substance  Name   Higher Fatty Acid (Palmitic  Acid) 
Definition   Higher Fatty Acid ( Palmitic Acid)  is one of the higher fatty acids  (substances that are 
obtained by hydrolyzing animal or vegetable fats or oils or hydrogenated animal or vegetable fats or 
oils).  It consists mainly of palmitic acid.  
Content  Higher Fatty Acid ( Palmitic Acid)  contains not less than 50.0% of palmitic acid (C16H32O2 
= 256.42 ). 
Description   Higher Fatty Acid ( Palmitic Acid)  occurs as white to bright grayish yellow flakes, 
powder, or granules or as a waxy lump.  
Identification   Analyze the test solution and the standard solution prepared  as directed  in the Assay 
by gas chromatography using the operating conditions specified in the Assay. The retention time of 
the main peak from the test solution corresponds to that of the peak of methyl palmitate from the 
standard solution.  
Iodin e Value   Not more th an 2.0.  
Test Solution   Weigh  accurately about 1 g of Higher Fatty Acid ( Palmitic Acid) into a 500 -mL 
stoppered flask, and dissolve it in 20 mL of a 1 : 1 mixture of cyclohexane /chloroform.  
Procedure  Proceed as directed under Iodine  Value in the Fats and R elated Substances Tests.  
Purity  
(1) Acid value   212−222.  
(2) Lead   Not more than 2 μg/g as Pb (2.0 g, Method 1, Control Solution: Lead Standard Solution 
4.0 mL, Flame Method).  
(3) Arsenic   Not more than 3 μg/g as As  (0.50 g, Method 3, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Residue on Ignition   Not more than 0.1%.  
Assay  
Test Solution  Weigh 20 mg of Higher Fatty Acid ( Palmitic Acid) , and transfer it into a small 
conical flask fitted with a reflux condenser. Add 5 mL of boron trifluoride−methanol TS, shake, and 
heat for about 10 minutes until it dissolves. Add through the condenser 4 mL of hexane, and heat 
for 10 minutes. After cooling, add 20 mL of a saturate solution of sodium chloride, shake, and leave 
to stand to allow the solution to separate into two layers. Transfer 2 mL of the separated hexane 
layer into another flask through about 0.2 g of sodium sulfate previously washed with hexane. To 1 mL of this solution, add h exane to make 10 mL, and shake.  
Standard Solution  Dissolve 10 mg of methyl palmitate  by adding 5 mL of hexane.  
Procedure  Analyze 1  L each  of the test solution  and the standard solution by gas 
chromatography using the operating conditions given below. Determine the peak area (A A) of methyl 
palmitate and the total peak area (A T) of all the fatty acid esters (total peak area of all peaks 
detected)  in the test solution, and calculate the palmitic acid content in the fatty acid fraction of the 
sample by the following formula. Identification of methyl palmitate sh ould be confirmed by 
comparing the retention time of the main peak from the test solution with that of methyl palmitate 
in the standard solution.  The chromatography should be continued for 1.5 times the retention time 
of methyl palmitate, and the main solvent peak should be excluded from the measurement.  
             Content  (%) of palmitic  acid = AA
AT ×100 
Operating conditions  
Detector: Flam e ionizat ion detector . 
Column: A fused silica tube (0.25 mm internal diameter and 50 m length) coated with a 0.2 -
m thick layer of 100% cyanopropylpolysiloxane for gas chromatography.  
Column temperature: 180°C.  
Injection port temperature: 250°C.  
Detector temperatur e: 250°C.  
Carrier gas: Helium.  
Flow rate: A constant rate of about 1.0 mL/min.  
Injection method: Splitless.  
 
  Higher Fatty Acid ( Behenic Acid)  
高級脂肪酸（ ベヘニン酸）  
 
Compositional Sp ecifications  
 
Substance  Name   Higher Fatty Acid (Behenic  Acid) 
Definition   Higher Fatty Acid ( Behenic Acid)  is one of the higher fatty acids  (substances that are 
obtained by hydrolyzing animal or vegetable fats or oils or hydrogenated animal or vegetable fats or 
oils).  It consists mainly of behenic acid.  
Content  Higher Fatty Acid ( Behe nic Acid)  contains not less than 50.0% of behenic acid (C 22H44O2 
= 340.58).  
Description   Higher Fatty Acid ( Behenic Acid)  occurs as white to bright grayish yellow flakes, 
powder,  or granules or as a waxy lump.  
Identification  Analyze the test solution and the standard solution prepared as directed in the 
Assay by gas chromatography using the operating conditions specified  in the Assay. The retention 
time of the main peak from the test solution corresponds to that of the peak of  methyl behenate from 
the standard solution.  
Iodin e Value   Not more than 3.0.  
Test Solution   Weigh  accurately about 1 g of Higher Fatty Acid ( Behenic Acid) into a 500 -mL 
stoppered flask, and dissolve it in 20 mL of a 1 : 1 mixture of cyclohexane /chloroform . 
Procedure  Proceed as directed under Iodine  Value in the Fats and Related Substances Tests.  
Purity  
(1) Acid value   160−175.  
(2) Lead   Not more than 2 μg/g as Pb (2.0 g, Method 1, Control Solution: Lead Standard Solution 
4.0 mL, Flame Method).  
(3) Arsenic   Not more than 3 μg/g as As  (0.50 g, Method 3, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Residue on Ignition  Not more than 0.1%.  
Assay  
Test Solution  Weigh 20 mg of Higher Fatty Acid ( Behenic Acid) , and transfer it into a small 
conical flask fitted with a reflux condenser. Add 5 mL of boron trifluoride−methanol TS, shake, and 
heat for about 10 minutes until it dissolves. Add through the condenser 4 mL of hexane, and heat 
for 10 minutes. After cooling, add 20 mL of a saturate solu tion of sodium chloride, shake, and leave 
to stand to allow the solution to separate into two layers. Transfer 2 mL of the separated hexane 
layer into another flask through about 0.2 g of sodium sulfate previously washed with hexane. To 1 mL of this soluti on, add hexane to make 10 mL, and shake.  
Standard Solution  Dissolve 10 mg of methyl behenate  by adding 5 mL of hexane.   
Procedure  Analyze 1  L each of the test solution and the standard solution by gas 
chromatography using the operating conditions given below. Determine the peak area (A A) of methyl 
behenate  and the total peak area (A T) of all the fatty acid esters (total peak area of all peaks detected)  
in the test solution, and calculate the behenic acid content in the fatty acid fraction of the sample by 
the following formula. Identification of methyl behenate should be confirmed by comparing the 
retention time of the main peak from the test solution w ith that of methyl behenate in the standard 
solution.  The chromatography should be continued for 1.5 times the retention time of methyl 
behenate, and the main solvent peak should be excluded from the measurement.  
             Content  (%) of behenic  acid = AA
AT ×100 
Operating conditions  
Detector: Flam e ionization detector . 
Column: A fused silica tube (0.25 mm internal diameter and 50 m length) coated with a 0.2 -
m thick layer of 100% cyanopropylpolysiloxane for gas chromatography.  
Column temperature: 180°C.  
Injection port temperature: 250°C.  
Detector temperature: 250°C.  
Carrier gas: Helium.  
Flow rate: A constant rate of about 1.0 mL/min.  
    Injection method: Splitless.  
 
Reagents, Solutions, and Other Reference Materials  
Methyl Behenate   See Methyl Docosanoate . 
Methyl Do cosanoate  C23H46O2  [929-77-1]  A colorless  crystalline powder . 
Melting point  53−56°C . 
 
  Higher Fatty Acid ( Myristic Acid)  
高級脂肪酸（ ミリスチン酸）  
 
Compositional Sp ecifications  
 
Substance  Name   Higher Fatty Acid (Myristic Acid)  
Definition   Higher Fatty Acid (Myristic Acid)  is one of the higher fatty acids  (substances that are 
obtained by hydrolyzing animal or vegetable fats or oils or hydrogenated animal or vegetable fats or 
oils).  It consists mainly of myristic  acid.  
Content  Higher Fatty Acid (Myristi c Acid)  contains not less than 50.0% of m yristic  acid (C 14H28O2 
= 228.38).  
Description   Higher Fatty Acid (Myristic  Acid)  occurs as white to bright grayish yellow flakes, 
powder, or granules or as a waxy lump.  
Identification  Analyze the test solution and the standard solution prepared as directed in the Assay 
by gas chromatography using the operating conditions  specified  in the Assay. The retention time of 
the main peak from the test solution corresponds to that of the peak of  methyl myristate from the 
standard solution.   
Iodin e Value   Not more than 1.0 . 
Purity  
(1) Acid value   240−250.  
(2) Lead   Not more than 2 μg/g as Pb (2.0 g, Method 1, Control Solution: Lead Standard Solution 
4.0 mL, Flame Method).  
(3) Arsenic   Not more than 3 μg/g as As  (0.50 g, Method 3, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Residue on Ignition  Not more than 0.1%.  
Assay  
Test Solution  Weigh 20 mg of Higher Fatty Acid (Myristic Acid) , and transfer it into a small 
conical flask fitted with a reflux condenser. Add 5 mL of boron trifluoride−methanol TS, shake, and 
heat for about 10 minutes until it dissolves. Add through the condenser 4 mL of hexane, and heat 
for 10 minutes. After cooling, add 20 mL of a saturate solu tion of sodium chloride, shake, and leave 
to stand to allow the solution to separate into two layers. Transfer 2 mL of the separated hexane 
layer into another flask through about 0.2 g of sodium sulfate previously washed with hexane. To 1 
mL of this soluti on, add hexane to make 10 mL, and shake.  
Standard Solution  Dissolve 10 mg of methyl myristate by adding 5 mL of hexane.  
Procedure  Analyze 1  L each  of the test solution and the standard solution by gas chromatography using the operating conditions given below. Determine the peak area (A A) of methyl 
myristate and the total peak area (A T) of all the fatty acid esters (total peak area of all peaks 
detected)  in the test solution, and calculate the myristic acid content in the fatty acid fraction of the 
sample by the following formula. Identification of methyl myristate should be confirmed by 
comparing the retention time of the main peak from the test solution  with that of methyl myristate 
in the standard solution . The chromatography should be continued for 1.5 times the retention time 
of methyl myristate, and the main solvent peak should be excluded from the measurement.  
             Content  (%) of myristic  acid = AA
AT ×100 
Operating conditions  
Detector: Flam e ionization detector . 
Column: A fused silica tube (0.25 mm internal diameter and 50 m length) coated with a 0.2 -
m thick layer of 100% cyanopropylpolysiloxane for gas chromatography.  
Column temperature: 180°C . 
Injection port temperature: 250°C.  
Detector temperature: 250°C.  
Carrier gas: Helium.  
Flow rate: A constant rate of about 1.0 mL/min.  
    Injection method: Splitless.  
 
Reagents, Solutions, and Other Reference Materials  
Methyl Myristate   See Methyl Tetradecanoate . 
Methyl Tetrad ecanoate   C15H30O2  [124-10-7]  A colorless , transparent liquid . 
Refractive index  nD20: 1.434 −1.438 . 
Specific gravity  d2020: 0.853 −0.873.  
 
  Higher Fatty Acid ( Lauric Acid)  
高級脂肪酸（ ラウリン酸）  
 
Compositional Sp ecifications  
 
Substance  Name   Higher Fatty Acid (Lauric Acid)  
Definition   Higher Fatty Acid (Lauric Acid)  is one of the higher fatty acids  (substances that are 
obtained by hydrolyzing animal or vegetable fats or oils or hydrogenated animal or vegetable fats or 
oils).  It consists mainly of  lauric  acid.  
Content  Higher Fatty Acid (Lauric Acid)  contains not less than 50.0% of l auric  acid (C 12H24O2 = 
200.32).  
Description   Higher Fatty Acid (Lauric  Acid)  occurs as white to bright grayish yellow flakes, powder, 
or granules or as a waxy lump.  
Identification  Analyze the test solution and the standard solution prepared  as directed  in the Assay 
by gas chromatography using the operating conditions specified  in the Assay. The retention time of 
the main peak from the test solution corresponds to that of the peak of methyl laurate from the 
standard solution.  
Iodin e Value   Not more than 1.0.  
Purity  
(1) Acid value   275−285.  
(2) Lead   Not more than 2 μg/g as Pb (2.0 g, Method 1, Control Solution: Lead Standard Solution 
4.0 mL, Flame Method).  
(3) Arsenic   Not more than 3 μg/g as As  (0.50 g, Method 3, Standard Color: Arsenic Standard 
Solution 3.0 mL, Apparatus B).  
Residue on Ignition   Not more than 0.1%.  
Assay  
Test Solution  Weigh 20 mg of Higher Fatty Acid ( Lauric  Acid) , and transfer it into a small 
conical flask fitted with a reflux condenser. Add 5 mL of boron trifluoride−methanol TS, shake, and 
heat for about 10 minutes until it dissolves. Add through the condenser 4 mL of hexane, and heat 
for 10 minutes. After cooling, add 20 mL of a saturate solution of sodium chloride, shake, and leave 
to stand to allow the solution to separate into two layers. Transfer 2 mL of the separated he xane 
layer into another flask through about 0.2 g of sodium sulfate previously washed with hexane. To 1 
mL of this solution, add hexane to make 10 mL, and shake.  
Standard Solution  Dissolve 10 mg of methyl laurate by adding 5 mL of hexane.  
Procedure  Analy ze 1  L each of the test solution and the standard solution by gas chromatography using the operating conditions given below. Determine the peak area (A A) of methyl 
laurate and the total peak area (A T) of all the fatty acid esters (total peak area of all p eaks detected)  
in the test solution, and calculate the lauric acid content in the fatty acid fraction of the sample by 
the following formula. Identification of methyl laurate should be confirmed by comparing the 
retention time of the main peak from the tes t solution with that of methyl laurate in the standard 
solution . The chromatography should be continued for 1.5 times the retention time of methyl laurate, 
and the main solvent peak should be excluded from the measurement.  
             Content  (%) of lauric  acid = AA
AT ×100 
Operating conditions  
Detector: Flam e ionization detector . 
Column: A fused silica tube (0.25 mm internal diameter and 50 m length) coated with a 0.2 -
m thick layer of 100% cyanopropylpolysiloxane for gas chromatography.  
Column tem perature: 180°C.  
Injection port temperature: 250°C.  
Detector temperature: 250°C.  
Carrier gas: Helium.  
Flow rate: A constant rate of about 1.0 mL/min.  
Injection method: Splitless.  
 
  Quicklime  
生石灰  
 
Compositional Specifications  
Definition  Quicklime  is obtained by calcinating limestones . It consists  mainly of calcium oxide.  
Content   Quicklime, when ignited, contains not less  than 9 3.0% of calcium oxide (CaO = 56.08).  
Description   Quicklime  occurs as  white to grayish -white lumps, granules, or powder . 
Identification  
(1) Moisten  1 g of Quicklime  with water. It generates heat.  Then add 5 mL of water. The 
resulting suspension is alkali.  
(2) To 1 g of Quicklime , add 20 mL of water and 6 mL of acetic  acid (1 in 3) , and  filter.  The 
filtrat e responds to a ll the tests for Calcium Salt as described in the Quantitative Tests.  
Purity  
(1) Hydrochloric acid -insoluble substances   Not more than 1.0%. 
Weigh accurately a crucible -type glass filter  (1G4) , previously dried at 105°C for 30 minutes 
and cooled in a desiccator . Weigh 5.0 g of Quicklime , add 100 mL of water, then add hydrochloric  
acid dropwise while shaking until the sa mple no longer dissolves, and boil. After cooling, a dd 
hydrochloric acid  to acidify if necessary.  Filter through with the previously we ighed glass filter . 
Wash the residue well  on the filter with water until the washings are free of chloride. Dry the 
residue with  the filter at 105°C for 1 hour.  Allow to c ool in a desiccator , and weigh accurately . 
(2) Carbonate   Weigh  2.0 g of Quicklime , add 50 mL  of water,  shake  thoroughly, and add 
25 mL  of diluted hydrochloric acid (1 in 4).  It does not  effervesce  vigorously.  
(3) Lead   Not more than 5 μg/g as Pb ( 0.80 g, Method 5, Control Solution: Lead  Standard 
Solution 4.0 mL, Flame Method).  
Sample S olution   Weigh the specified amount of Quicklime  into a crucible.  Heat it 
gradually to carbonize, and ignite at 500 °C with  a lid. Add  20 mL of  diluted  hydrochloric  acid (1 
in 4)  to the residue , cover with a watch glass,  and boil gently  for 15 minutes.  After c ooling, add 
30 mL of water.  If the sample does not dissolve, evaporate to dryness, add 20 mL of  diluted  
hydrochloric  acid (1 in 4)  to the residue , and boil gently  for 5 minutes.  After c ooling, add 30 mL 
of water . 
In Method 5 of the Lead Limit Test,  use 50 mL of a solution of diammonium hydrogen  citrate 
(1 in 2) and 1 mL of bromothymol blue TS as the indicator instead of thymol blue TS , and then 
add ammonia solution  until the color of the solution changes from  yellow to yellow -green.  
(4) Alkali metals and magnesium   Not more than 6.0%. Weigh  accurately  about 0.5 g of Quicklime , add 30 mL of water and 15 mL of diluted  
hydrochloric acid (1 in 4) to dissolve it . Heat the solution, and boil for 1 minute. Immediately 
add 40 mL of  a solution of oxalic acid dihydrate (3 in 50), agitate, and add 2 drops of methyl red 
TS, then add ammonia sol ution  dropwise  until the solution exhibits a yellow color to precipitate 
the calcium.  Heat this solution on a water bath for 1 hour, cool, and add water to make 100 mL . 
Mix well, and filter . Transfer 50 mL of the filtrate into a platinum crucible, previously ignited 
at 800°C for 30 minut es, cooled in a desiccator, and weighed accurately. Add 0.5 mL of sulfuric 
acid, evaporate to dryness , ignite at 800°C to constant weig ht, and weigh the crucible to  
determine the weight of the residue.  
(5) Barium  Not more than 300 μg /g as Ba.  
Test Solution   Weigh 1. 50 g of Quicklime , and add water to make  it muddy . Add 20 mL of  
diluted  hydrochloric acid (1 in 4) to dissolve it,  and add water to make 30 mL. Filter  the solution , 
and use 20 mL of the filtrate  as the test solution.  
Procedure   To the test solution , add 2 g of sodium acetate , 1 mL of diluted  acetic acid  (1 in 
20), and 0.5 mL of  potassium chromate solution  (1 in 20), and  allow to stand for 15 minutes.  The 
solution is not more  turbid than a control solution  prepared as follows:  To 0.30 mL of Barium 
Standard Solution, add  water to make 20  mL, and then treated in the same manner as for the 
test solution.  
(6) Arsenic   Not mor e than 3 μg/g as As (0.50 g, Standard Color: Arsenic Standard  Solution 
3.0 mL, Apparatus B).  
Test Solution   Dissolve the specified amount of Quicklime  in 8 mL of diluted  hydrochloric 
acid (1 in 4).  
Loss on Ignition   Not more than 10.0 % (800°C, constant wei ght).  
Assay   Weigh accurately about 1.5 g of Quicklime , previously ignited, add 30 mL of  diluted 
hydrochlo ric acid (1 in 4) to dissolve , and add water to make exactly 250 mL.  Use the resulting 
solution as the test solution. Proceed as directed under Method  1 in Calcium Salt Determination.  
Each mL of 0.05 mol/L disodium dihydrogen ethylenediaminetetraacetate = 2.804  mg of CaO . 
 
  Ethyl Acetoacetate  
アセト酢酸エチル  
 
 
The compositional specifications for Ethyl Acetoacetate were  revised as follows : 
 
 
Reagents, Solutions, and Other Reference Materials  
Bromocresol  Purple   C21H16Br2O5S [K8841,  Special Grade] [115 -40-2] 
Bromocresol  Purple  TS  Dissolve 50 mg of  bromocresol purple in 100 mL of ethanol (95).  Filter if 
necessary.  Revised regulation  Previous  regulation  
Purity   
Acid value   Not more than 5.0 (Flavoring 
Substances Tests). To confirm the endpoint 
with  an indicator , use b romocresol  purple  
TS. The endpoint is when the color of the 
solution changes from yellow  to bluish  
purple . Purity  
Acid value   Not more than 5.0  
(Flavoring Substances Tests).  