JPH1130610A - Hop originated volatile aromatic component measuring method - Google Patents

Abstract

PROBLEM TO BE SOLVED: To analyze and determine the quantity of sesquiterpenes in a hop originated volatile aromatic component easily and correctly without being influenced by other volatile components usign a gas chromatography-mass spectrometry (GCS-MS) and a selective ion detecting method. SOLUTION: Sesquiterpenes in a hop originated volatile aromatic component existing in a drink with hop used as a part of material is detected using a GCS-MS and a selective ion detecting method. A sesquiterpene such as caryophyllene and humulene are named as the sesquiterpenes contained as the hop originated volatile aromatic component. Oxygen contained sesquiterpene with one or more oxygen atoms combined with the sesquiterpene, to be concrete, humulenol, humuladienone, and the like, can also be named. The sesquiterpenes can be correctly determined since volatilization caused by heating during operation is not much generated because of its relatively high boiling point.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to fermentable beverages such as beer and low-malt beer using hops as a part of raw materials, and wort, and relates to fermentable beverages and hops present in wort. The present invention relates to a method for simply and accurately measuring a volatile odor component.

[0002]

2. Description of the Related Art In beverages using hops such as beer and low-malt beer as a part of raw materials, the unique aroma and taste derived from hops are important factors that characterize such beverages. Various hop-derived aroma components in this beverage have been clarified, but it is not always clear at present whether any one of them substantially contributes to determine the aroma of the hop of the beverage. According to the study of the present inventors, hop-derived terpenes, that is, hydrocarbon terpenes and oxygen-containing terpenes may be involved, and the sensory evaluation results of beer or low-malt beer It is predicted from the results of comparative study.

[0003] Terpenes are, in principle, natural organic compounds which are biosynthesized by combining a plurality of isoprene units (C ₅ ), and more than 6,000 are known. They are,
Depending on the number of isoprene units, monoterpenes (C ₅ ×
2), sesquiterpenes (C ₅ × 3), diterpenes (C ₅ × 4) and the like.

The hop-derived terpenes include hydrocarbon terpenes and oxygen-containing terpenes as described above. The hydrocarbon terpenes are terpenes composed of a carbon atom and a hydrogen element. Examples of hydrocarbon terpenes contained in beer include monoterpenes such as myrcene, sesquiterpenes such as humulene, and diterpenes. Further, the oxygen-containing terpenes are those in which one or more oxygen atoms are bonded to the above-mentioned hydrocarbon terpenes. Specifically, an epoxy compound, an alcohol compound, a ketone compound, or the like can be given. All of these hop-derived terpenes have a strong scent, and the intensity is such that even humans can perceive it even in trace amounts on the order of ppm or less.

[0005] In particular, hydrocarbon terpenes are highly volatile and highly hydrophobic, so that they are almost volatilized and removed during the wort boiling, fermentation, and filtration steps and remain in the final product. The amount to do is insignificant. However, even with such a slight residual amount, the scent of terpenes is strong enough to be perceived by human olfaction. Oxygen-containing terpenes are more water-soluble than hydrocarbon terpenes, and therefore transfer a larger amount to the final product (ie, beer or low-malt beer), and are unique due to the balance of these volatile flavor components. It has a scent.

As a method of objectively analyzing the amount of these volatile odor components derived from hops, an analysis method using a gas chromatograph (hereinafter abbreviated as GC) can be considered.

However, when taking beer as an example of a beverage, volatile substances relating to the aroma of beer include not only the above-mentioned volatile aroma components derived from hops (eg, terpenes) but also fermentation metabolites produced by yeast. Volatile components (esters, alcohols, aldehydes, ketones, etc.). The volatile components of these fermentation metabolites are present in orders of magnitude higher than the volatile flavor components derived from hops. Therefore, when GC analysis of volatile substances in beer is performed, the peak waveform of the fermentation metabolite is large, and the small peak waveform of the volatile aroma component derived from hops is masked. It was difficult to analyze only the volatile odor components involved.

[0008] In identifying and quantifying each volatile component in these beers, extraction with an organic solvent or the like is performed to separate and separate the volatile components.
Although it is common to use a method of isolation, since the fermentation metabolite is overwhelmingly contained in a larger amount than the volatile aroma component derived from hops as described above, the desired hop-derived It was difficult to reliably isolate only the volatile odor components, and knowledge and skill were required to work efficiently.

[0009]

DISCLOSURE OF THE INVENTION The present inventors have made intensive studies to solve the above-mentioned problems of the prior art. As a result, among terpenes, which are substances involved as volatile odor components derived from hops in beverages, sesquiterpenes, in particular, remain in the final product in relatively large amounts and characterize hop-derived aroma. noticed.
That is, the present inventors have found that among the volatile aroma components derived from hops, sesquiterpenes have a high content in the terpenes contained in the final product, a relatively high boiling point, and fragment ions from fermentation metabolites. Note that the sesquiterpenes are selectively analyzed using gas chromatography-mass spectrometry (hereinafter abbreviated as GC-MS) and a selected ion detection method, noting that the sesquiterpenes are not easily affected by masking. By this, it was found that the content of sesquiterpenes was easily and accurately known, and that this could be used as a measure of the total amount of terpenes, which are volatile aroma components derived from hops. Thus, the present invention has been completed.

[0010] The present invention characterizes the hop-derived aroma (hop aroma) contained in a final product or an intermediate product among hop-derived volatile aroma components in a beverage using hop as a part of a raw material. Among the terpenes that have been, the amount of sesquiterpenes is easily and accurately analyzed and quantified without being affected by other volatile components, thereby estimating the amount of hop-derived aroma components in the product, It is an object of the present invention to provide a method capable of performing an objective evaluation without using a sensory test. In other words, an object of the present invention is to provide a method capable of measuring the quality and quantity of a hop-derived aroma component without using a sensory test.

[0011]

Means for Solving the Problems The present invention uses a GC-MS method for measuring a hop-derived volatile aroma component present in a beverage using hop as a part of a raw material,
The present invention also provides a method for measuring a hop-derived volatile odor component, which comprises detecting sesquiterpenes in a hop-derived volatile odor component using a selective ion detection method.

[0012]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below.
The method of the present invention is directed to beverages using hops as part of the raw material. Here, as a beverage using hops as a part of the raw material, for example, beer, low-malt beer, and the like can be given. It is needless to say that the present invention can be applied not only to the beverage as described above but also to wort before fermentation.

In the method of the present invention, when measuring the volatile odor components derived from hops present in a beverage using the hops as a part of the raw material as described above, the volatile fragrance components derived from hops may be sesquiscent. Terpenes are detected. Sesquiterpenes have a carbon skeleton represented by C ₁₅ H ₂₄ in principle, and have various structures such as linear, monocyclic, bicyclic, and tricyclic. The carbon skeleton alone contains more than 100 types and more than 1000 compounds, and is the largest group among terpenes.

Among them, sesquiterpenes such as caryophyllene, humulene, and farnesene are examples of sesquiterpenes contained as volatile aroma components derived from hops. In addition, oxygen-containing sesquiterpenes in which one or more oxygen atoms are bonded to those sesquiterpenes can be mentioned, and specific examples include humulenol, humladienone, humulene epoxide, and caryophyllene epoxide. it can.

Since sesquiterpenes have a relatively high boiling point [boiling point of about 250 ° C. (normal pressure)], volatilization due to heating during operation does not occur so much, so that accurate quantification is possible. Further, as described later, sesquiterpenes are hardly affected by masking of fragment ions derived from fermentation metabolites during mass spectrometry.

In the method of the present invention, sesquiterpenes in hop-derived volatile aroma components present in a beverage using hops as a part of a raw material are subjected to GC-MS and selected ion detection. It is characterized by using and detecting.

Specifically, a volatile component (sample) containing an organic component obtained from a hop-based beverage, which is a final product, is separated by GC and then subjected to mass spectrometry (MS). Detect and identify terpenes.

More specifically, for example, sesquiterpenes may be detected as follows. First, the volatile components are extracted from the beverage to be investigated. The extraction of the volatile component is performed, for example, by a solid phase extraction method. For example, the beverage to be investigated is placed in a sealed sample bottle with a septum cap, then the sample bottle with the beverage is immersed in a thermostat, and the volatile components in the beverage are converted to the gas phase in the container. Spread. In order to sufficiently diffuse the volatile components, it is desirable that the temperature of the thermostat is set to about 30 to 60 ° C. and the immersion is performed for about 5 to 30 minutes.

An adsorption needle having a surface coated with an organic component (volatile component) adsorption resin for a solid phase is inserted into the gas phase. In order to adsorb a sufficient amount of organic components (volatile components), the adsorption needle is set for 5 to 30 minutes, preferably 10 to 2 minutes.
It is desirable to insert it for about 0 minutes.

The adsorption needle which has sufficiently adsorbed the organic component (volatile component) is extracted and applied to a gas chromatograph-mass spectrometer (hereinafter abbreviated as GC-MS device).

Here, the GC-MS device comprises a GC unit and an MS.
And an apparatus as shown in FIG. The GC section is composed of a sample introduction section and a separation column, and is individually temperature-controlled. The MS section is maintained in a high vacuum, and includes an ionization section for ionizing a sample, an ion separation section for separating ions by mass number (m / z), and a detection section for separated ions. Consists of The recording data detected by the ion detection unit of the MS unit can be recorded and processed by a separately provided recording device and data processing device (not shown).

The extracted suction needle is first introduced into a sample introduction section in a GC section of a GC-MS apparatus. The temperature of the sample introduction section is individually controlled as described above, and is maintained at a high temperature. Specifically, it is maintained at about 200 to 280 ° C. As described above, since the temperature of the sample introduction portion is high, the organic component (volatile component) adsorbed on the resin of the adsorption needle is instantaneously vaporized and separated. The organic component (volatile component) vaporized and separated in the sample introduction section is subsequently supplied to a separation column, where the target substance in the volatile component is separated.

The target substance separated in the GC section is
It is introduced into the ionization section of the S section. In the ionization section, the volatile components are ionized by an electron impact ionization method. In this method, a molecular ion is generated from an organic component in the volatile component by colliding electrons (irradiating an electron beam) with the volatile component that has become a gas. By being hit by electrons, molecular ions are further cleaved,
Fragment ions are generated.

The fragment ions produced in the ionizing section are then input to the ion separating section, where these ions are sequentially separated according to the mass number (m / z). There are various types of separation methods in the ion separation unit, and typical examples include a magnetic field type, a quadrupole type, and an ion trap type. In the present invention, any method can be used, and the voltage and the magnetic field or the electric field of the ion separation unit are sequentially changed, so that the ions having a small mass number can be sequentially reached to the next ion detection unit. .

The ion detector records the amount of ions reached as an amount of electricity. Detection methods include mass spectrum detection, total ion detection, and selected ion detection.
In the present invention, selective ion detection is performed in order to selectively detect sesquiterpenes.

The present invention is characterized in that selective ion detection is performed in order to selectively detect sesquiterpenes.
By using the selected ion detection method and setting the mass number (m / z) of the ions to be detected in the range of 190 m / z to 240 m / z, only sesquiterpenes can be selectively detected and analyzed. it can.

The reason is as follows. The mass number of the cleavage ion demethylated from the molecular ion of the sesquiterpene, the cleavage ion dehydrated from the oxidized sesquiterpene, and the molecular ion or fragment ion of the compound having one or two oxidized oxygen atoms bonded thereto Are included in the range of the mass number (m / z).
On the other hand, the mass numbers of the fermentation metabolite molecular ions other than the terpenes and the fragment ions of the cleaved fragments thereof are not included in this range. Therefore, the mass number is 19
When the range is from 0 m / z to 240 m / z, only sesquiterpenes can be detected without fear of masking by metabolite molecular ions other than terpenes or their fragmented ions.

As described above, with respect to fermentable beverages such as beer and low-malt beer using hops as a part of the raw material, and wort, the fermentable beverage and the hop-derived volatile aroma present in the wort are used. Particularly, only sesquiterpenes among the components can be easily and accurately measured.

[0029]

EXAMPLES The present invention will be described below in more detail with reference to Examples, but the present invention is not limited thereto.

Experimental Example 1 First, a sample was extracted using a solid phase extraction method. That is, 20 ml of beer was placed in a sealed sample bottle with a septum cap, immersed in a thermostat at 40 ° C. for 15 minutes, and the volatile components in the beer were diffused into the gas phase in the container. Then, into this gas phase, an adsorption needle whose surface is coated with an adsorption resin is inserted, and after the organic component is adsorbed while being left inserted for 15 minutes, the adsorption needle is pulled out.
It was directly introduced into a GC-MS device as shown in FIG.
The GC conditions in the GC-MS device are as follows.

[GC conditions] Separation column: DB-1 df 1 μm × 0.25 mm × 30 m He: 15 psi, split / splitless injection port Valve switching: 3 minutes, INJ: 250 ° C. Transfer line: 270 ° C.・ Separation column temperature: 50 ° C, 1 minute → Heat at 5 ° C / min → 2
50 ° C, 3 minutes

The introduction of the suction needle was performed as follows.
That is, the adsorption needle was introduced into the sample introduction part of the GC part maintained at 250 ° C., and the organic component (volatile component) sample adsorbed on the resin of the adsorption needle was instantaneously vaporized and separated. . Next, the organic component (volatile component) sample that was vaporized and separated was supplied to a separation column in the GC section. The temperature of the separation column was initially maintained at 50 ° C. for 1 minute,
Heat to 250 ° C at a rate of 5 ° C per minute, 250 ° C
For 3 minutes.

The organic component (volatile component) sample separated and heated and vaporized in the GC section (sample introduction section and separation column) as described above was subsequently introduced into the high vacuum MS section. First, this organic component (volatile component) sample was subjected to MS
Introduced to the ionization part of the part, by electron impact ionization method,
The sample was ionized. Next, the fragment ions produced in the ionization section were input to the ion separation section, where the ions were separated according to the mass number (m / z), and the ions having the smaller mass number were sequentially reached to the ion detection section. .

The ion detector uses a selected ion detector (SIM) to reduce the mass number from 65 m / z to 255 m.
/ Z range and set the total ion chromatography (Total Ion Ch
romato; TIC). FIG. 2 shows the measurement results of the TIC in the range of a mass number of 65 m / z to 255 m / z. FIG. 2 is an ion chromatogram when beer head space analysis was performed.

As can be seen from the results shown in FIG. 2, a number of peaks (*) not observed with the cold wort were observed. Peaks (*) not observed in cold wort but observed in beer are fatty acids and esters which are fermentation metabolites. From this result, it can be seen that various types of organic components (volatile components) are included in the range of mass numbers of 65 m / z to 255 m / z. That is, it is difficult to examine only the volatile flavor components derived from hops from the chromatogram as shown in FIG. Therefore, the following Experimental Example 2 was performed in order to specify the range of the mass number (m / z) suitable for the measurement of terpenes, which are a kind of volatile aroma components derived from hops.

Experimental Example 2 In Experimental Example 1, the mass number of the SIM was changed to 67 m / z, 9
3m / z, 109m / z, 138m / z, 204m /
z or 220 m / z, and the same procedure as in Experimental Example 1 was performed, except that ions of each mass number were detected by the selected ion detection method.

Here, the mass number of 67 m / z means isoprene (C ₅ H ₈ , MW = 68) which is a basic skeleton of terpene.
It is a residue ion and corresponds to the mass number of the characteristic cleavage ion of terpenes. Further, the mass numbers 93 m / z, 109 m / z, and 138 m / z correspond to the mass numbers of ions that frequently appear when the terpene compound is cleaved. Mass number 204m / z
Is a molecular ion of sesquiterpene (C ₁₅ H ₂₄ ; MW = 204) and a cleavage ion deprived from oxygen-containing sesquiterpene such as humulene epoxide (MW = 204:
220-16). 220m /
z is a molecular ion (MW = 220: 204 + 16 (mass number of oxygen)) of a compound in which one oxygen atom is bonded to a sesquiterpene such as humuladienone or humulene epoxide.
Corresponds to the mass number of The chromatograms at each mass number are shown in FIGS.

The following can be seen from the results shown in FIGS. First, in FIGS. 3 to 6, peaks are found at the same positions as in FIG. These indicate the presence of fermentation metabolites such as fatty acids and esters. Therefore, as shown in FIGS. 3 to 6, the mass number of the SIM is 67 m / z, 93 m / z, 1
In the case of 09 m / z and 138 m / z, as in FIG. 2, the peaks of components other than the volatile aroma component derived from hops are affected, and these ions are used to generate volatile aroma components derived from beer hops. It is difficult to make an analysis. On the other hand, in FIGS. 7 and 8, no peak was observed as shown in FIGS. 2 and 3 to 6. Since the mass numbers 204 m / z and 220 m / z set in FIGS. 7 and 8 correspond to the mass numbers of sesquiterpenes, it is recognized that they indicate the presence of hop-derived sesquiterpenes.

Therefore, it can be seen that the mass number should be set in the range of 190 to 240 m / z in order to specifically analyze sesquiterpenes, which are a kind of volatile aroma components derived from hops. According to FIGS. 7 and 8, peaks such as those shown in FIGS. 2 and 3 to 6 are not recognized,
Instead, a peak was observed in the cold wort, that is, a peak based on hop-derived sesquiterpenes.

[0040]

According to the method of the present invention, among the volatile aroma components derived from hops in a beverage using hops as a part of the raw material, the fragrances derived from the hops are contained in the final product or the intermediate product, Of the terpenes that characterize hop aroma, the amount of sesquiterpenes can be easily and accurately analyzed and quantified without being affected by other volatile components. It is possible to estimate the amount of hop-derived aroma components and perform an objective evaluation without using a sensory test. According to the method of the present invention, the quality and quantity of the hop-derived aroma component can be measured without using a sensory test.

In other words, according to the method of the present invention, among the volatile fragrance components derived from hops in a beverage using hops as a part of the raw material, the content of the fragrance component recognized as a hop-derived fragrance is determined as follows: It can be compared and determined by the total area of the chromatographic peak waveforms, and since this determination is correlated with the sensory test, it is possible to control the hop-derived aroma in the beverage based on this determination method, It becomes possible to easily produce a beverage having hop flavor according to taste.

[Brief description of the drawings]

FIG. 1 is an explanatory diagram showing an example of a chromatograph-mass spectrometer (CG-MS device) used in the present invention.

FIG. 2 The mass number of beer is 65 to 255 m / z.
FIG. 5 is a diagram showing the results of measurement of total ion chromatography (TIC) when CG-MS was performed with the setting of.

FIG. 3 is a diagram showing the results of ion chromatography measurement when CG-MS is performed on beer with a mass number set to 67 m / z.

FIG. 4 is a view showing the results of ion chromatography measurement when CG-MS is performed on beer at a mass number of 93 m / z.

FIG. 5 is a diagram showing the results of ion chromatography measurement when CG-MS is performed on beer at a mass number of 109 m / z.

FIG. 6 is a view showing the results of ion chromatography measurement when CG-MS is performed on beer at a mass number of 138 m / z.

FIG. 7 is a diagram showing the results of ion chromatography measurement when CG-MS is performed on beer at a mass number of 204 m / z.

FIG. 8 is a diagram showing the results of ion chromatography measurement when CG-MS was performed on beer at a mass number of 220 m / z.

Claims (2)

[Claims] 1. A gas chromatograph-mass spectrometry method for measuring a hop-derived volatile aroma component present in a beverage using hops as a part of a raw material, and
A method for measuring hop-derived volatile odor components, comprising detecting sesquiterpenes in hop-derived volatile odor components using a selective ion detection method. 2. The method according to claim 1, wherein a fragment ion and a molecular ion having a mass of about 190 to 240 are detected as specific components in the volatile odor components derived from hops.