Evolution of Fructans in Aguamiel (Agave Sap) During the Plant Production Lifetime

Abstract

Aguamiel is the sap collected from agave, while pulque is the result of the natural fermentation of aguamiel. Despite its ancestral origin and numerous publications on pulque production, little is known about the evolution and concentration of sugars and fructo-oligosaccharides in aguamiel, either during its daily accumulation or through the agave production lifetime. In this study, we examined aguamiel composition in three agave plants during their productive lifetime (4 to 9 months). After each collection, the agave pine is scraped to induce aguamiel to flow into an internally created cavity (cajete), producing a residual bagasse (metzal). We found that the concentration of agave fructans and sucrose, as well as the fructan profile, change during the aguamiel production process. During the daily collection, a small amount of agave fructans released from the pine by scraping is drawn into the cajete with the first milliliters of sap where it is then diluted with the inflow of aguamiel. The main component of aguamiel is the sucrose produced in high concentration in the leaves through photosynthesis and then hydrolyzed in the cajete as aguamiel accumulates. We also describe how the fructan profile changes during the accumulation of aguamiel in the cajete. In addition to the varying amount of sucrose that is hydrolyzed in the aguamiel accumulated, we found that fructo-oligosaccharides are either diluted, consumed, or hydrolyzed, depending on the plant and its production stage, thus yielding different fructan profiles. New fructo-oligosaccharides are, in some cases, synthesized by bacteria present in aguamiel. These profiles were also observed in aguamiel collected from ten different plants in the same production region. We also found that a considerable amount of agave fructans is lost in metzal (bagasse), the agave material that is scraped and thrown away twice a day during the production process.

Keywords: aguamiel; fructans; fructo-oligosaccharides; metzal; pine; pulque; scraped; sucrose.

Figure 1

Sugar content in aguamiel at different collection times (0, 3.5, and 7 h after scraping) during a single day collection: sucrose (A,C) and fructo-oligosaccharide profile (B,D). Scraping as usual before collection (A,B) or scraping before each aguamiel sample was collected (C,D). Sucrose was measured by HPLC; FOS was measured by HPAEC-PAD. The HPAEC-PAD chromatograms (B,D) for plant P2 are shown as an example. Statistical differences: p-value for sucrose concentration in the traditional process for the three plants (P1, p = 0.968; P2, p = 0.111; and P3, p = 0.012). When the plant was scraped before sampling (P1, p = 0.779; P2, p = 0.005; and P3, p = 0.73). The cajete was previously emptied before sampling to avoid dilution.

Figure 2

Changes in the concentration of sucrose, fructans, and the fructan profile in aguamiel exuded during the 3 h following scraping. Aguamiel samples were always collected in a previously emptied cajete to avoid dilution with accumulated aguamiel. Only three HPAEC profiles are shown.

Figure 3

Examples of the three different FOS profiles observed in accumulated aguamiel. HPAEC-PAD oligosaccharide profiles in fresh (immediately after scraping) and accumulated (10 h) aguamiel. (A) Type 1. Synthesis of oligosaccharides: A. mapisaga in the early productive stage (B) Type 2. Agavin-like profile and synthesis of new oligosaccharides: A. mapisaga in intermediate-late productive stages, and (C) Type 3. Fructan dilution, A. mapisaga in an intermediate-late productive stage. See explanation in the text.

Figure 4

HPAEC-PAD oligosaccharide profile in samples of accumulated aguamiel before and after enzymatic treatment to identify the chemical nature of oligosaccharides. (A) Treatment with Fuctozyme. (B) Treatment with Dextranase. The analyses correspond to samples of accumulated aguamiel from plants P1 and P2 (A. mapisaga).

Figure 5

Changes in sucrose and fructan concentration, as well as in the FOS profile, in fresh aguamiel collected after scraping [(A,C), respectively] and metzal [(B,D), respectively] during the agave productive lifetime. Statistical differences: p-values for sucrose concentration (P1, p < 0.0005; P2 p < 0.0005; and P3 p < 0.0005), while for fructan concentration (P1, p < 0.0005; P2 p < 0.0005; and P3 p < 0.0005).

Figure 6

Fructan molecular weight distribution as determined by Gel Permeation Chromatography (GPC) in (A) fresh aguamiel and (B) metzal during the agave productive lifetime as well as in (C) Stem sections of reference plant (S3.2 = Upper stem section; S4 = section 4; S5= section 5 and S6 = lower stem section).

Figure 7

Schematics of the distribution of simple sugars and fructans in an A. salmiana pine. Fructan concentration of the center sample: S1:268.5, S2:260.3, S3.1:380.6, S3.2:307.8, S4:397.9, S5:531.6, S6: 565.9 mg/g of dry mass. Simple sugars concentration of the center sample S1:118.1, S2:67.9, S3.1:67.8, S3.2:39.9, S4:49.8, S5:28.5, S6:18.5 mg/g of dry mass.