Development of Continuous-Mode Separation Processes for High-Purity Recoveries of Neoagarohexaose and Galactotriose from Biomass-Derived Oligosaccharides

Abstract

Recently, it has been discovered that neoagarohexaose (NA6) contained in neoagarooligosaccharides (NAO), which originate from the β-agarase hydrolysis of agarose in red algae, has various medical effects. This has led to growing interest in establishing an industrial-utilizable process for high-purity separation of NA6 from NAO, which has, however, yet to be realized. To address this issue, we attempt to develop an efficient simulated-moving-bed (SMB) process for continuous-mode separation of NA6 from NAO with high purity and high yield, which was named “NA6-SMB”. It was found first that the most suitable adsorbent for the targeted SMB separation was the ion-exclusion resin of sodium form with 4% crosslinkage, and its proper operating temperature was 60 ℃. For the selected absorbent with the aforementioned properties, the intrinsic parameters of all NAO components were determined. Using the resultant parameter values, the configuration and operation parameters of the NA6-SMB were designed such that the high-purity separation of NA6 could be realized and kept stable throughout a long period of SMB operation while maximizing throughput. The separation efficiency of such designed NA6-SMB was first validated by model simulations, and then verified by experiments, which confirmed that the NA6 product of 98.7% purity could be obtained continuously throughout the long-term NA6-SMB processing while keeping its loss below 1%. Finally, an additional SMB was developed to treat the non-product stream of the NA6-SMB for the purpose of producing high-purity NA4 (> 99%), which has been considered worthy of testing its medical efficacy and physiological activation in large quantity. As the next stage of research, we aimed at accomplishing comprehensive optimization of NA6-SMB, which was carried out such that the throughput of the NA6-SMB could be maximized to the maximum extent possible while keeping the utilization of its adsorbent at the highest reachable level. This task was carried out on the basis of a standing-wave-design method. It was found first that in a conventional frame where the same column length was set to be used in Ring I and Ring II under the column configuration of 2–2–2–2 in each ring, the separation sequence based on removing a larger-size impurity in Ring I and then smaller-size impurities in Ring II (S1 mode) led to higher throughput than the opposite separation sequence (S2 mode). By contrast, if the column lengths of the two rings were allowed to be set differently, the bed utilization of the S2 mode could increase more than that of the S1 mode, thereby causing the S2 mode to surpass the S1 mode in throughput. In addition, if the column configuration of each ring was allowed to be varied, the level of attainable throughput could be increased by allotting more columns in the enrichment zones containing the front and rear of product solute band in Ring I. Furthermore, it was confirmed that the simultaneous application of the aforementioned two methods (i.e., using different column lengths in two rings and placing more columns in the enrichment zones of Ring I) had an additional upward effect on each other. As a result, the optimized NA6-SMB under such frame, whose column length and configuration were 13.33 cm and 2–4–3–3 in Ring I and 18.67 cm and 2–3–5–2 in Ring II, could achieve 46% higher throughput than that under the conventional frame, and further lead to 133% higher throughput than the initial NA6-SMB. Galactotriose (G3), which is contained in a crude galactooligosaccharides (GOS) mixture coming from enzymatic transgalactosylation reactions, is known to have potential as a high-grade prebiotic ingredient. This has aroused an interest in the high-purity recovery of G3 from a crude GOS mixture, which included monosaccharides (galactose and glucose), disaccharides (G2), and GOS components with different degrees of polymerization (G3, G4, and G5). To settle this problem, we aimed to develop an efficient simulated-moving-bed process for high-purity recovery of G3 from the crude GOS mixture. It was found first that the appropriate column temperature and ionic form of the adopted adsorbent in consideration of both G3-G2 and G3-G4 resolutions were 65 ℃ and K+ form respectively. Under such adsorbent conditions, the equilibrium constants and mass-transfer para- meters of the crude mixture components were determined and then exploited in the design of the G3-recovery SMB, which was carried out with a priority focus on the stable maintenance of high purity for G3 product throughout the SMB operation. The separation performance of such designed SMB was verified on the bases of the column profiles from simulations and the outlet- concentration data from the relevant SMB experiment. Overall, it was confirmed that the designed SMB could recover G3 from the crude GOS mixture in a continuous mode with a purity of more than 98.9% while keeping the overall loss of G3 below 7.3%. |최근 홍조류에 함유된 아가로스의 β-아가레이즈 가수분해에서 유래한 네오아가로올리고당(NAO)에 함유된 네오아가로헥사오스(NA6)가 다양한 의학적 효능을 갖고 있다는 사실이 밝혀졌다. 이로 인해 NAO에서 NA6의 고순도 분리를 위한 산업 규모의 분리공정에 대한 관심이 높아지고 있으나, 아직 개발된 바 없다. 이 문제를 해결하기 위해 "NA6-SMB"라는 이름의 고순도 및 고수율 연속 분리에 적합한 SMB(simulated-moving-bed)공정을 개발했다. 표적물질 분리에 가장 적합한 흡착제는 4% 가교결합을 갖는 Na+ 형태의 이온교환수지이며, 최적 운전 온도는 60℃인 것이 처음으로 밝혀졌다. 해당 분리 조건에서 모든 NAO 구성 물질의 고유 매개변수 값을 조사했다. 조사한 매개변수 값을 사용한 모델 시뮬레이션을 바탕으로 NA6의 고순도를 유지하며, 처리량을 최대화하고, 장기간 운전에 안정성을 유지하는 NA6-SMB를 설계하고 운전변수 값을 확보했다. 이렇게 설계된 NA6-SMB의 분리 효율을 먼저 모델 시뮬레이션을 통해 검증한 후, 실험을 통해 NA6를 순도 98.7% 이상, 1% 미만의 손실로 생산할 수 있음을 검증했다. 마지막으로 NA6-SMB의 부산물로부터 의학적 효능과 생리 활성을 시험할 가치가 있다고 판단되는 고순도 NA4(>99%)를 생산하기 위한 NA4-SMB를 추가로 개발했다. 이어지는 연구에서, NA6-SMB의 처리량을 최대화하면서 흡착제 활용도를 최대로 유지하며 운전변수와 공정변수의 포괄적인 최적화를 시도하였다. 탠덤 SMB의 각 링을 구성하는 컬럼의 열 길이와, 각 링을 구성하는 컬럼의 구성을 두가지 공정시스템변수로 설정하고, 각 변수에 대해 정상파설계방법을 기반으로 하는 SMB 최적화도구를 사용하여 공정변수와 운전변수를 포괄적으로 최적화했다. 최종적으로 기존 NA6-SMB 대비 2.3배 높은 처리량을 확보할 수 있었다. 갈락토트리오스(G3)는 효소에 의한 갈락토실화 반응에서 나오는 미정제 갈락토올리고당혼합물(GOS)에 함유되어 있으며 고급 프리바이오틱 성분으로서 높은 가치가 있는 것으로 알려져 있다. 단당류(갈락토스 및 글루코스), 이당류(G2) 및 중합도가 다른 GOS 성분(G3, G4 및 G5)을 포함하는 미정제 GOS 혼합물로부터 G3의 고순도 회수를 목표로 하는 G3-SMB공정을 개발했다. G3-G2와 G3-G4 분해능을 모두 고려할 때, 운전 온도 65℃에서 K+ 형태의 이온교환수지가 적합한 흡착제임을 확인했다. 해당 분리 조건에서 미정제 GOS 혼합물 성분의 고유 매개변수를 결정하여 G3의 고순도를 안정적으로 유지하는데 중점을 둔 G3-SMB의 설계에 활용했다. 설계한 G3-SMB공정은 미정제 GOS 혼합물로부터 7.3% 미만의 손실률을 유지하며 순도 98.9% 이상의 G3를 생산할 수 있음을 확인하였다.